2005 FETAL AND NEONATAL PHYSIOLOGICAL SOCIETY · 2005 FETAL AND NEONATAL PHYSIOLOGICAL SOCIETY...

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Transcript of 2005 FETAL AND NEONATAL PHYSIOLOGICAL SOCIETY · 2005 FETAL AND NEONATAL PHYSIOLOGICAL SOCIETY...

Page 1: 2005 FETAL AND NEONATAL PHYSIOLOGICAL SOCIETY · 2005 FETAL AND NEONATAL PHYSIOLOGICAL SOCIETY Quick Program Guide 32nd ANNUAL FETAL AND NEONATAL PHYSIOLOGY SOCIETY JEFFREY ROBINSON
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2005 FETAL AND NEONATAL PHYSIOLOGICAL SOCIETY

Quick Program Guide 32nd ANNUAL FETAL AND NEONATAL PHYSIOLOGY SOCIETY JEFFREY ROBINSON SYMPOSIUM

(Begins with the Geoffrey Dawes lecture Sept 27th) SUNDAY

September 25th MONDAY

September 26th TUESDAY

September 27th WEDNESDAY September 28th

0730 Registration: FNPS or Combined FNPS/JSR 0815 WELCOME

Epidemiology 0825 WELCOME 0830-0910 Rajan Yajnick (India)

0910-0950 David Henderson-Smart (AUS)

0830-1030 8 Free Orals 8 Free Orals 7 Free Orals 0845-1030 0950-1020 Free oral communications 1 R Painter, M Schwab

1030-1100 MORNING TEA MORNING TEA MORNING TEA 1020-1100 MORNING TEA HPA / Programming / Placenta 1100-1140 Roger Smith (AUS)

1100-1300 8 Free Orals 8 Free Orals 8 Free Orals 1140-1220 Les Myatt (USA)

1220-1305 Free oral communications 2 J Bromfield, A Sferruzzi-Perri, M Tare

1300-1530 LUNCH & POSTERS LUNCH 1300-1345

LUNCH 1300-1430 AGM 1400-1430 1305-1500

LUNCH & POSTERS

Sponsored by DSL

Traditional Sporting Event Begins at 1400h 8 Free orals (1430-1630) Fetus / Development / Growth Factors

1530-1600 AFTERNOON TEA 1500-1540 Jane Harding (NZ)

Registration for attendees ofJSR Program only 1540-1620 David Hill (Canada)

1600-1745 7 Free Orals AFTERNOON TEA 1630-1700 1620-1705 Free oral communications 3

M Wallace, H Budge, B Muhlhausler

1800 Drinks & Nibbles Geoffrey Dawes Lecture

Jeffrey Robinson (1700-1800)

1705-1800 Round Table Discussion with drinks

1830-2000 DATA BLITZ (Selected Posters) Eugenie Lumbers (AUS) / Peter Gluckman (NZ) /

John Challis (Canada) DINNER at 2000h DINNER at 1900h

Combined FNPS/JSR GALA DINNER

1930h drinks for 2000h 1900 JSR SYMPOSIUM DINNEREND OF FNPS PROGRAM

END OF JSR PROGRAM

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Contents Quick Program Guide................................................................................................................... 2

Contents........................................................................................................................................ 3

Organising and Scientific Committees......................................................................................... 4

FNPS Mission Statement ............................................................................................................. 5

Geoffrey Dawes Lecture Fund ..................................................................................................... 6

Howard Florey Research Fund..................................................................................................... 7

Previous Meetings of the FNPS ................................................................................................... 8

Fetal and Neonatal Physiological Society 31ST Annual Business Meeting Minutes 2004 .......... 9

33RD FNPS Conference Cambridge 2006 .................................................................................. 11

34TH FNPS Conference Japan 2007 ........................................................................................... 12

35TH FNPS Conference The Netherlands 2008.......................................................................... 13

Sponsorship and Support............................................................................................................ 14

Student Prizes............................................................................................................................. 15

Maps of Glenelg and Adelaide................................................................................................... 16

Things to do while in Adelaide .................................................................................................. 18

Notes for presentations............................................................................................................... 19

Program Outline ......................................................................................................................... 20

Detailed Program........................................................................................................................ 21

Sunday September 25, 2005................................................................................................... 21

Monday September 26, 2005 ................................................................................................. 31

Tuesday September 27, 2005 ................................................................................................. 34

JEFFREY ROBINSON SYMPOSIUM ..................................................................................... 38

Tuesday September 27, 2005 ................................................................................................. 38

Wednesday September 28, 2005 ............................................................................................ 38

Plenary and Keynote Speakers................................................................................................... 41

32ND FNPS 2005 ORAL ABSTRACTS .................................................................................... 42

32ND FNPS 2005 DATA BLITZ ABSTRACTS........................................................................ 83

32ND FNPS 2005 POSTER ABSTRACTS ................................................................................ 92

JEFFREY ROBINSON SYMPOSIUM ABSTRACTS ........................................................... 110

List of Delegates....................................................................................................................... 130

List of Presenting Authors........................................................................................................ 138

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2005 FETAL AND NEONATAL PHYSIOLOGICAL SOCIETY

Organising and Scientific Committees

Organised by: FETAL AND NEONATAL PHYSIOLOGICAL SOCIETY

FNPS Board Members

Laura Bennet, New Zealand (Chair)

Carlos Blanco, The Netherlands

Dino Giussani, United Kingdom

Mark Hanson, United Kingdom

Richard Harding, Australia

Brian Koos, USA

Anibal Llanos, Chile

Carina Mallard, Sweden

Emanuela Marinoni, Italy

Karel Marsál, Sweden

Tim Moss, Australia

Julian T Parer, USA (Scribe)

Dan Rurak, Canada

Gerard Visser, The Netherlands

Charles Wood, USA

Convenor:

Cathie Coulter

Scientific Committee:

Janna Morrison

Paul Colditz

Karen Gibson

Richard Harding

Caroline McMillen

David Walker

John Newnham

Mary Wlodek

Karen Moritz

JSR Symposium Convenors

Kathy Gatford

Julie Owens

Local Organising Committee

Julia Pitcher

Miles De Blasio

Sanita Grover

Olivia Wyss

Amanda Sferruzzi-Perri

Natasha Campbell

Audio Visual: Mark Grover

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FNPS Mission Statement

The FNPS stimulates discussion and exchange of ideas between physiologists, obstetricians and

neonatologists. The FNPS considers an informal gathering and presentations of new and preliminary

data, especially by investigators in training, essential to achieve goals.

The society was founded in 1974 during an informal meeting in Oxford. Professor Geoffrey Dawes

(1918-1996) and Dr. Gerhard (Bo) Gennser took the initiative and were made honorary members of

the society in 1995. The name of the annual conference (and society) has changed several times,

reflecting the widening scope of the society.

1974-80 Conference on Fetal Breathing

1981-83 International Conference on Fetal Breathing and other Movements

1984-95 Society for the Study of Fetal Physiology

1996-date Fetal and Neonatal Physiological Society

Over the years the society has maintained its informal character and a lack of rigid structures. Those

who have attended at least one of the previous three meetings are members of the society and will be

informed about the next meeting. Abstracts for the annual meeting are requested two months before

the meeting and are compiled in the Book of Abstracts to encourage recent and preliminary data to

be presented.

The Organizational Coordinator will be selected by the Organizational Committee and shall serve

for three years. The Organizational Committee shall consist of representatives from Africa, Asia,

Australia, Canada, continental Europe, South America, the United Kingdom and the United States of

America and shall be selected by the Committee.

The annual meeting will be held in Europe, North America and the Southern Hemisphere, in June-

September, as determined by the Organizational Committee. Approximately half of the meetings

will be held in Europe. The costs for registration and accommodation should not exceed US$500

(shared room) for a three-day meeting. Any residual funds from prior meetings shall be passed on

to the coordinator for the next meeting. Audit will not be required if the residual funds are less than

10,000 US$. The (local) Organising Committee shall have the right to solicit funds in the name of

the Society from organisations for the purpose of providing financial support for students and

fellows-in-training to attend the meeting of the Society.

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Geoffrey Dawes Lecture Fund

1. The fund is established with excess moneys from the 1998 meeting. It is placed in a safe,

interest bearing account under the above name. It is an educational fund and has tax-exempt

status.

2. The primary intention of the establishment of the fund is to financially support the Geoffrey

Dawes Lecture at the Annual Meeting.

3. The disbursement of interest from the account will be as follows:

a) 5% of interest will be retained for account expenses and hedge against inflation.

b) The remainder of the annual interest will be available to the local organizing committee

for support of the Geoffrey Dawes Lecture of the following year. Unused money will be

returned to the fund. Expenses for the lectureship above and beyond the interest money

available will be the responsibility of the local committee.

c) The annual interest may be used for purposes other than that outlined in 3(b), but must be

approved by a majority vote of the committee.

c) Check from the account will require 2 signatures of the FNPS Committee.

4. The fund may be added to (in addition to 3(a)) from the following a) sources:

a) Donation to the Society.

b) Solicitations from other organizations or businesses.

c) Money in excess of US$ 3,000 is available from the prior meeting.

Generally US$ 3,000 is available from the prior meeting for passing on to the next local

committee.

Previous Geoffrey Dawes Lecturers

1999 Christine L Muimmery, the Netherlands

2000 David Barker, FRS, UK

2001 Ruud Kleinpaste, NZ

2002 Anthony Carter, Denmark

2003 Peter Gluckman, NZ

2004 Angelo Vescovi, Italy

2005 Jeffrey Robinson, Australia

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Howard Florey Research Fund The Florey Medical Research Fund is the fundraising and public affairs activity of the Medical

Foundation of the University of Adelaide in respect to its role in support of the Adelaide Medical

School. All funds raised by The Florey Medical Research Fund are deposited with The Medical

Foundation and administered by it.

The Florey Medical Research Fund is named to honour the memory of The Adelaide Medical

School's most famous graduate, Howard Florey, who received the Nobel Prize for his work in

developing penicillin. Howard Florey's development of penicillin for clinical use ushered in the

wonderful new era of antibiotics and changed the course of human history.

Florey was born, raised and educated in Adelaide, at Kyre School (now Scotch College) and St

Peter's College before graduating from Adelaide University Medical School in 1921. His hard work

and natural talents earned him bursaries to fund his education and a Rhodes Scholarship took him

overseas to pursue his destiny. Florey's host of honours and achievements included a Nobel Prize

award, presidency of the Royal Society and the inaugural Chancellorship of the Australian National

University, which he helped to establish at the request of the Australian Government. "It is hard for

us nowadays to imagine the world before antibiotics. Death from infection was commonplace and

surgical procedures were fraught with danger."

Continuing the Florey tradition, Adelaide University Medical School has research personnel of the

highest calibre engaged on projects of major international standard and importance. Sadly, however,

traditional funding sources are coming under threat and there is a growing critical need for new

donation support to maintain work. The Florey Medical Research Fund has been established to help

meet this challenge. People can help by becoming regular supporters of annual medical research

projects and/or making a bequest commitment in their Wills.

The Florey Medical Research Fund need and welcomes bequest of all sizes.

The fund supports a wide range of medical research projects at The Adelaide Medical School, which

has provided excellence in teaching and research for over a hundred years. All the projects

undertaken are of international standard.

http://www.florey.adelaide.edu.au/index.html

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Previous Meetings of the FNPS

1974 Oxford, United Kingdom

1975 Oxford, United Kingdom

1976 Malmö, Sweden

1977 Oxford, United Kingdom

1978 Nijmegen, The Netherlands

1979 Paris, France

1980 Oxford, United Kingdom

1981 Maastricht, The Netherlands

1982 London, Canada

1983 Malmö, Sweden

1984 Oxford, United Kingdom

1985 Haifa, Israel

1986 Banff, Canada

1987 Groningen, The Netherlands

1988 Cairns, Australia

1989 Reading, United Kingdom

1990 Pacific Grove, USA

1991 De Eemhof, The Netherlands

1992 Niagara-on-the-Lake, Canada

1993 Plymouth, United Kingdom

1994 Palm Cove, Australia

1995 Malmö, Sweden

1996 Arica, Chile

1997 S.Margherita Ligure, Italy

1998 Lake Arrowhead, USA

1999 Vlieland, The Netherlands

2000 Southampton, United Kingdom

2001 Auckland, New Zealand

2002 Prague, Czech Republic

2003 Banff, Canada

2004 Castelvecchio Pascoli (Tuscany), Italy

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Fetal and Neonatal Physiological Society 31ST Annual Business Meeting Minutes 2004

Tuesday,September 14, 2004

Il Ciocco International Centre Castelvecchio Pascoli (Tuscany) Italy

O1. The minutes of the previous meeting held in Banff, Canada in July 2003 were accepted

with a minor modification. Dr Parer explained that Item 5, regarding the Dawes Fund has been temporarily held up but progress is being made.

O2. Enthusiastic thanks were given to Romolo Di Iorio and Emanuela Marinoni for the

incredibly successful meeting this year in Tuscany.

The registration is amongst the highest we have had and both the scientific content and social aspects were considered of the highest order by the registrants. The people present confirmed this by vociferous and prolonged acclimation to the major organizers.

O3. The website is up and running and all agreed that it is serving a major purpose for the society.

O4. Changes in Board membership: Bryan Richardson has stepped down from the Board after

a long and fruitful association. He wanted a further Canadian member to contribute to the success of the society, and Dan Rurak has accepted the position.

Further changes in the Board composition are expected within the next year.

O5. Student Prizes: Awards for the Best Oral Presentation were shared between Monica Longo for her presentation on Nitric Oxide knockout mice and Candice Rodricks for her presentation on the effect of hypoxia on the chick brain.

Two equal prizes were also awarded for the poster presentations, to Lindsay Patrick on the time course of maternal cytokine responses in fetal brain injury and to Gioia Alvino for her paper on antioxidant vitamins in IUGR pregnancies. Certificates and awards were given to each of the students amidst widespread applause.

O6. Future meetings have firmed up somewhat. The meeting for 2005 will be held at the Stamford Grand, Glenelg, South Australia, from September 25- September 28, 2005. This meeting will be combined with a full day symposium to celebrate the career of Jeffrey Robinson. Following the meeting, there will be a further meeting in Fremantle, Western Australia on Preterm Delivery. It will be held from Friday, September 30th to Monday, October 3, 2005.

The meeting for 2006 will be held in Cambridge, UK, from the 17th – 20th of September 2006. The meeting for 2007 is planned for Sendai, Japan, at the Hotel Sendai Plaza, from September 2nd-5th, 2007.

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The meeting for 2008 is being pursued vigorously by Jan Nijuis of Maastricht, Holland, and Chile has also requested the venue for that year. With respect to 2009, it is felt that the meeting should move back to the Western Hemisphere and both the U.S. and Uruguay are requesting the meeting. It should be noted that except for the meeting immediately following this one, the dates still remain tentative for the more distantly planned meetings.

O7. The Coordinator of the meeting, Laura Bennett, requested that the members with creative

skills in this regard compose a logo for the society. She will arrange for their competitive display on the website.

Respectfully submitted Julian T. Parer, Scribe San Francisco

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2005 FETAL AND NEONATAL PHYSIOLOGICAL SOCIETY

33RD FNPS Conference Cambridge 2006

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34TH FNPS Conference Japan 2007

http:/

HOT2-20-1 HON-CHOU AOB

Telephone: 022-2August 26 (S

Cc/o MA C

5F, Dai 2 IzumChiyoda-k

TEL: +81-3-52E-

/www.macc.jp/34fnps/

Venue: EL SENDAI PLAZA

A-KU, SENDAI-CITY, MIYAGI, 980-0014 JAPAN

62-7111 Facsimile: 022-262-8169 unday) - 29 (Wednesday) 2007

ongress Secretariat: onvention Consulting Inc. i-shoji Bldg., 4-2-6 Kojimachi,

u, Tokyo 102-0083, JAPAN 75-1191 / FAX: +81-5275-1192 mail: [email protected]

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35TH FNPS Conference The Netherlands 2008 Hotel Van der Valk, Maastricht, the Netherlands

22 – 25 June 2008 Maastricht, the Netherlands, Awaits You in 2008!

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During this FNPS meeting we will celebrate the careers of Jelte de Haan and Carlos Blanco,

both of whom will have retired by 2008 Maastricht In June the weather will be mild and sunny in Maastricht. The oldest and most Burgundian city in the Netherlands offer many ways to relax (terraces, pubs and restaurants). Located in the hilliest part of the Netherlands, its hills and marlstone caves offer great views and numerous possibilities for nice walks, biking, horse riding, etc. Situated in the center of Europe, Maastricht is a one-hour drive from Brussels or Düsseldorf, a two-hour drive from Amsterdam. The city also has its own airport with connections to London, Amsterdam, Pisa, etc. Hotel Van der Valk Known for its spacious rooms and reasonable prices, the Hotel van der Valk in Maastricht is a luxury 5-star hotel and only a 10-minute walk from the centre of town. The 2005 price for a standard room is € 79 and a business suite is € 132,50. When sharing a room, this conference will certainly offer “value for money.” Society Sport Event The classic FNPS Sport Event will not be forgotten. With the Dutch organizing things, you can be sure that we will have a real competition! Organizing Committee The familiar faces from the Netherlands (i.e. Bilardo, Blanco, De Haan, Mulder, De Vries, Erwich, Hasaart, Nijhuis, Oei, Van den Berg, Visser, etc.) will participate in the Organizing Committee to create a meeting that is successful - both scientifically and socially. Contact for the Dutch 2008 FNPS Meeting Jan G. Nijhuis Conference Agency Maastricht e-mail: [email protected] [email protected] +31 43 387 47 68 T +31 43 361 91 92 P.O. Box 5800 P.O. Box 1402 NL-6202 AZ Maastricht NL-6201 BK Maastricht

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Sponsorship and Support

We thank the following for their sponsorship and support:

Major Sponsors

Department of Obstetrics & Gynaecology, University of Adelaide

The Florey Medical Research Foundation

The Ian Potter Foundation

Faculty Health Sciences, University of Adelaide

Discipline of Physiology, School of Molecular & Biomedical Sciences, University of Adelaide

Sponsors

Fetal & Neonatal Physiology Society - Geoffrey Dawes Lectureship

Diagnostic Systems Laboratories

ARC/NHMRC Network in Genes Environment and Development

Perinatal Research Centre - University of Queensland

Corbett Life Science

Wyeth

Supporters

Texas Peak Pty Ltd (Brooks)

Cambridge University Press

GroPep

Springer

Trade Exhibitor

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Student Prizes

Abstract Awards

Corbett Life Science Award for Best Student Abstract in the Oral Presentation Category:

Lina Gubhaju

Corbett Life Science Award for Best Student Abstract in the Poster Presentation Category:

Kathryn Franko

JSR Student Abstract Awards

The DSL Young Scientist Awardees: John Bromfield

Beverly Muhlhausler

The JSR Young Scientist Award: Amanda Sferruzzi-Perri

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Maps of Glenelg and Adelaide GLENELG:

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ADELAIDE CBD & SURROUNDS:

Maps courtesy of the RAA and The City of Holdfast Bay.

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Things to do while in Adelaide

Just some of the many suggestions (see the previous maps for locations):

Take a stroll along Adelaide’s cultural boulevard, North Terrace, and visit the Australian Aboriginal

Cultures Gallery and the Bradman Collection at the SA museum, State Library and Art Gallery of

South Australia, North Terrace

Shop at the Adelaide Central Markets, between Grote and Gouger Streets

Visit Haigh’s Chocolate Factory, Greenhill Road

Visit the Adelaide Zoo, Frome Road

Catch the famous Popeye boat for a cruise along the River Torrens, Elder Park

Take the historic tram from Glenelg to the city (weekend only), Victoria Square, Jetty Road Glenelg

Play a round of golf at the North Adelaide Golf course overlooking the city

Visit the Adelaide Botanic Gardens, North Terrace

Visit Old Government House on North Terrace

Do some shopping on Australia’s longest shopping mall, Rundle Mall

Have a meal or do some shopping in the many shopping and restaurant precincts in Adelaide:

Rundle Street, Gouger Street, O’Connell St North Adelaide, Norwood Parade Norwood, King

William Road Hyde Park, Burnside Village Burnside, Unley Road Unley

Visit the Bicentennial Conservatory in the Botanical Gardens, North Terrace

Visit the National Wine Centre of Australia on North terrace

Visit historic Port Adelaide and take a cruise on board a tall ship or take a dolphin sightseeing cruise

in the new dolphin heritage of Port Adelaide

Visit McLaren Vale (45 mins south of the city), or the Barossa Valley (1 hour north of the city) wine

regions for hundreds of cellar doors for tastings and purchases

Visit Hahndorf, the historical Lutheran settlement town in the Adelaide hills (30 mins from the city)

Visit Mount Lofty Summit, the highest point in the Adelaide hills; eat at the 5-star restaurant or café

at the summit (20 minutes from the city)

Visit Monarto Zoological Park, Belair National Park, Cleland Wildlife Park, or Warrawong Wildlife

Sanctuary

Do a day trip to Kangaroo Island on the Sealink Ferry, or visit Victor Harbor and Goolwa for some

whale watching

Ride the Steamranger Steam Train between Victor Harbor and Goolwa

Visit Murray Bridge and take a cruise on a Paddle Steamer along the famous Murray River

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Notes for presentations

ORAL presentation Keynote lectures and oral presentations will take place in Ballroom 2 and 3

All presentations will be given in Microsoft PowerPoint. Speakers are asked to submit their

presentation on an IBM compatible floppy disk, CD or USB memory stick to the technician the day

prior their presentation.

Oral abstract presentation will be 10 minutes in length followed by a 5-minute question session.

POSTER presentation Poster abstract presentations will be held in Ballroom 1.

Presenters are asked to put up their poster on the morning of Sunday September 25th (Including Data

Blitz Posters). All posters will be displayed throughout the meeting and be removed no later than

3pm 28th September.

As the presenting Author, you are expected to be present during the time of your assigned poster

session. Presenters will be asked to give a very short 2-minute summary of your results during the

poster session.

Data Blitz Posters: 1 minute talk with maximum 2 slides and 2 minutes question time.

FLOOR PLAN OF THE STAMFORD GRAND ADELAIDE (LEVEL 1):

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Program Outline 32nd ANNUAL FETAL AND NEONATAL PHYSIOLOGY SOCIETY JEFFREY ROBINSON SYMPOSIUM

(Begins with the Geoffrey Dawes lecture Sept 27th) SUNDAY

September 25th MONDAY

September 26th TUESDAY

September 27th WEDNESDAY September 28th

0730 Registration: FNPS or Combined FNPS/JSR 0815 WELCOME

Epidemiology 0825 WELCOME 0830-0910 Rajan Yajnick (India)

0910-0950 David Henderson-Smart (AUS)

0830-1030 8 Free Orals 8 Free Orals 7 Free Orals 0845-1030 0950-1020 Free oral communications 1 R Painter, M Schwab

1030-1100 MORNING TEA MORNING TEA MORNING TEA 1020-1100 MORNING TEA HPA / Programming / Placenta 1100-1140 Roger Smith (AUS)

1100-1300 8 Free Orals 8 Free Orals 8 Free Orals 1140-1220 Les Myatt (USA)

1220-1305 Free oral communications 2 J Bromfield, A Sferruzzi-Perri, M Tare

1300-1530 LUNCH & POSTERS LUNCH 1300-1345

LUNCH 1300-1430 AGM 1400-1430 1305-1500

LUNCH & POSTERS

Sponsored by DSL

Traditional Sporting Event Begins at 1400h 8 Free orals (1430-1630) Fetus / Development / Growth Factors

1530-1600 AFTERNOON TEA 1500-1540 Jane Harding (NZ)

Registration for attendees ofJSR Program only 1540-1620 David Hill (Canada)

1600-1745 7 Free Orals AFTERNOON TEA 1630-1700 1620-1705 Free oral communications 3

M Wallace, H Budge, B Muhlhausler

1800 Drinks & Nibbles Geoffrey Dawes Lecture

Jeffrey Robinson (1700-1800)

1705-1800 Round Table Discussion with drinks

1830-2000 DATA BLITZ (Selected Posters) Eugenie Lumbers (AUS) / Peter Gluckman (NZ) /

John Challis (Canada) 2000 DINNER at 2000h DINNER at 1900h

Combined FNPS/JSR GALA DINNER

1930h drinks for 2000h 1900 JSR SYMPOSIUM DINNEREND OF FNPS PROGRAM

END OF JSR PROGRAM

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Detailed Program Sunday September 25, 2005 0730 – 0820 Registration

(For FNPS or Combined FNPS / JSR Symposium)

0825 – 0830 Welcome by Cathie Coulter Ballroom 2 & 3

0830 – 1030 Free Oral Communications 1 – PERIRENAL STRUCTURE AND FUNCTION

Chairs: Mary Wlodek (Australia) and Jane Black (Australia)

Ballroom 2 & 3

0830 – 0845 1. Upregulation of adrenal expression of the type 1 VEGF receptor in a model of activated-adrenal growth in the late gestation fetal sheep K.E. Warnes, I.C. McMillen, J.S. Robinson, C. L. Coulter.

0845 – 0900 2. VEGF and eNos in the ovine placenta are upregulated with altered maternal adrenal steroids Jensen E, Wood CE, and Keller-Wood M

0900 – 0915 3. The effects of furosemide on lambs born to mothers with renal dysfunction AE O’Connell, AC Boyce and KJ Gibson.

0915 – 0930 4. The effects of maternal renal dysfunction on mother and fetus. Eugenie R Lumbers, Karen J Gibson, Clare L Thomson, Amanda C Boyce, Bilal M Karime.

0930 – 0945 5. The renal handling of sodium and the renal response to amino acid infusion is altered in fetuses whose mothers have renal dysfunction Karen J. Gibson, Amanda C. Boyce, Bilal M. Karime and Eugenie R. Lumbers

0945 – 1000 6. Renal function in adult sheep following prenatal exposure to glucocorticoids Karen Moritz, Andrew Jefferies, Miodrag Dodic, Marelyn Wintour and Robert DeMatteo

1000 – 1015 7. The tubuloglomerular feedback mechanism in the developing kidney Russell D Brown, Anita J Turner, A Erik G Persson, Karen J Gibson

1015 – 1030 8. The effect of pre-term birth on nephrogenesis Gubhaju L, Zulli, A, Black MJ

1030 – 1100 Morning Tea Ballroom 1

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1100 – 1300 Free Oral Communications 2 - CARDIOVASCULAR

Chairs: Eugenie Lumbers (Australia) and Janna Morrison (Australia)

Ballroom 2 & 3

1100 – 1115 9. Growth and gene expression of components of the renin angiotensin system in offspring of rabbits with chronic maternal hypertension Devaki Maduwegedera, Karen Moritz, Rebecca L Flower, E Marelyn Winour, Kate M Denton

1115 – 1130 10. Maternal Microvascular Response to Corticotrophin Releasing Hormone (CRH) is Altered By Gestation, Fetal Gender and Pre-eclampsia Stark MJ, Dierkx L, Clifton VL, Wright IMR

1130 – 1145 11. Vitamin D insufficiency during early life is associated with raised blood pressure and altered vascular reactivity M. Tare, S.J. Emmett, R. Morley, C. Skordilis, H.A. Coleman, H.C. Parkington

1145 – 1200 12. Fetal oxygenation and Doppler ultrasonography of cardiovascular hemodynamics in a chronic near term sheep model Kaarin Makikallio, Tiina Erkinaro, Niina Niemi, Tomi Kavasmaa, Ganesh Acharya, Mervi Pakkila, and Juha Rasanen

1200 – 1215 13. Armed stem cells delivered to injured myocardium restore function and assume elements of the gene program regulating fetal/neonatal cardiomyocyte proliferation Ting Zhao, Joan P Stabila, Beth G McGonnigal, Yi-Tang Tseng, Naohiro Yano, Lawrence Lum, James F Padbury.

1215 – 1230 14. Increased sympathetic nerve activity to the cerebral circulation during blood pressure surges in REM sleep Priscila A Cassaglia, Robert I Griffiths & Adrian M Walker

1230 – 1245 15. Sympathetic nerve activity in the preterm fetal sheep is regulated by baroreflexes; evidence from telemetry based recordings SC Malpas, L. Booth, CJ Barrett, A Gunn, L Bennet

1245 – 1300 16. Fetal cardiac natriuretic peptide expression and cardiovascular hemodynamics in endotoxin-induced acute cardiac dysfunction in mouse Kaarin Makikallio, Samuli Rounioja, Olli Vuolteenaho, Jenna Paakkari, Mikko Hallman, and Juha Rasanen

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1300 – 1530 Lunch and Poster Viewing (Posters DB1-15 are also part of the Data Blitz session: 1830 – 2000h) Ballroom 1

DB1. Maternal dexamethasone treatment increases hepatic glycogen content but not G6Pase activity in the pregnant rat and fetus UKL Franko U, AJ Forhead, AL Fowden

DB2. Differential effects of maternal cold exposure on mRNA abundance for the growth hormone secretagogue receptor (1α) and glycerol-3-phosphatase in adipose tissue of the newborn sheep. EA Butt, S Pearce, T Stephenson, ME Symonds and UH BudgeU

DB3. The Impact of Placental Restriction on the Expression of the Appetite Neuropeptide Regulators, NPY and POMC, in the Hypothalamus of the Fetal Sheep in Late Gestation. UBHA Chuang U, BS Muhlhausler, JL Morrison, SJ Williams, CL Adam, PA Findlay and IC McMillen

DB4. Timing of delivery of IUGR fetuses on the basis of hemodynamic changes UEV CosmiU, G. Mari and E. Cosmi

DB5. The vasoreactivity of small placental arteries and systemic vascular bed UNoriaki Imai U, Minori Saito, Junichi Sugawara, Yoshitaka Kimura, Kunihiro Okamura

DB6. Neonatal Gender Influences Microvascular Function in Normal Newborns Stark MJ, Clifton VL, UWright IMRU

DB7. Leptin infusion does not alter cardiomyocyte development in the sheep fetus in late gestation UK.BottingU, I.C. McMillen, J. Dorosz, J.L. Morrison

DB8. Role of Beta1-adrenergic receptor (β1AR) Pi3K/p70S6k signaling in the regulation of neonatal cardiomyocyte proliferation Yi-Tang Tseng, Naohiro Yano, Ting Zhao, Vlad Ianus, Joan P Stabila, Beth G McGonnigal, UJames F Padbury U

DB9. Effect of Suppression of Neurosteroid Synthesis on Asphyxia-Induced Fetal Brain Injury UYawno TU, Yan EB, Walker DW and Hirst JJ

DB10. Secondary post-asphyxial hypoperfusion is associated with suppression of cerebral metabolism and increased tissue oxygenation in near-term fetal sheep UJensen ECU, Bennet L, Hunter CJ, Power GC, and Gunn AJ.

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DB11. Altered fetal brain development and growth following repeated, acute exposure to alcohol: potential mechanisms. Penelope A Dalitz, Sandra M. Rees, Kathryn L. Gatford, Julie A. Owens, Megan L Cock, Philip N Henschke, Richard Harding

DB12. Regional specificity of detection of ischemic changes on magnetic resonance imaging in the preterm fetal sheep Mhoyra Fraser, Laura Bennet, Chris E Williams, Peter D Gluckman, Alistair J Gunn, and Terrie E Inder.

DB13. Suppression of post hypoxic-ischemic EEG transients with delayed dizocilpine is associated with moderate striatal protection in the preterm fetal sheep Justin M Dean, Alistair J Gunn, Guido Wassink, Sherly George, Laura Bennet

DB14. Dose-response study of lipopolysaccharide-induced fetal brain injury in the guinea pig E.L. Harnett, M.A. Dickinson, and G.N. Smith.

DB15. Pituitary and Adrenocortical Responses to Acute Hypoxemia in Highland and Lowland Newborn Llamas Riquelme RA, Herrera EA, Sanhueza EM, Reyes VR, Ebensperger G, Ebensperger R, Parer JT, Giussani DA, Blanco CE, Hanson MA, Llanos AJ.

P1. The Role of Calcitonin Gene Related Peptide (CGRP) in Fetal Life A.S. Thakor and D.A. Giussani

P2. Melatonin and Fetal Physiology A.S. Thakor and D.A. Giussani

P3. Effects of antenatal glucocorticoid treatment on fetal development in mice Cornelle W. Noorlander, Pierre N.E. de Graan, Gerard H.A. Visser

P4. Activation of H9C2 Cell Proliferation by b-Adrenergic Receptors (AR) is via Involvement of PI3K Signaling Pathway Naohiro Yano, Vlad Ianus, Ting C. Zhao, Yi-Tang Tseng, James F. Padbury

P5. Influence of Porcine Genotype on the Abundance of Thyroid Hormones and Leptin in Sow Milk and its Impact on Plasma Concentrations in the Resulting Offspring A. Mostyn, J. C. Litten, K. S. Perkins, J. Laws, M. E. Symonds & L. Clarke

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P6. Effect of maternal cold exposure on hepatic glycogen content, peroxisome proliferator-activated receptor alpha and insulin-like growth factor-II mRNA expression in the neonate. E.A.Butt, S. Pearce, H. Budge, T. Stephenson and M.E. Symonds

P7. Intrauterine growth restriction in the fetal lamb: effect on the size of the macula densa and the renal renin-angiotensin system Vladislava Zohdi, Karen M Moritz, Kristen Bubb, Megan L Cock, Richard Harding, M Jane Black

P8. N-acetyl cysteine, a potential neuroprotectant, induces severe fetal hypoxemia in the presence of endotoxin. Megan Probyn, Megan Cock, Jhodie Duncan, Sandra Rees, Richard Harding

P9. Human Maternal-Fetal Placental Transfer of Proinflammatory Cytokines L.K. Moore, G.N. Smith.

P10. Maternal parity determines the abundance of mitochondrial proteins in the lung and kidney of developing sheep D. Yakubu, A. Mostyn, T. Stephenson and M.E. Symonds

P11. Peripheral vascular response to local cooling in pregnancy as measured by ultrasound and laser doppler Hartgill TW, Bergersen TK, Wall E L, Pirhonen J

P12. Brain metabolic changes in the late gestation fetal sheep in response to umbilical cord occlusion Edwin B. Yan, Suzanne L. Miller, Margie Castillo-Melendez, Phuong N Nguyen, Alexis A Adamides, David W. Walker

P13. The sensitising effects of inflammation on hypoxic-ischaemic brain injury G Kendall, A Clements, H Varley, G Raivich, D Peebles

P14. Expression of leptin and the leptin receptor in the ovine placenta D. M. O’Connor, E. Brookes, F.B.P. Wooding, N. Hoggard, A.L. Fowden and A.J. Forhead.

P15. Antenatal Glucocorticoids Suppress Cytochrome C Oxidase Activity in the Fetal Ovine Brain M. Schwab, G. Wichmann, M. Loehle, P.W. Nathanielsz, I. Maurer, O.W. Witte

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P16. Spiny Mouse – A Model for Perinatal Research Hayley Dickinson, David W Walker

P17. Gonadotrophic hormones reduce pregnancy rates and perturb fetal development due to changes in the maternal uterine and hormonal environment in mice RL Kelley, KL Kind, M Lane, RL Robker, JG Thompson, LJ Edwards

P18. IGF-II and IGF2R Expression in Early Murine Implantation Sites: A Role in Both Placentation and Decidual Angiogenesis? Pringle KG & Roberts CT

P19. Transient NMDA receptor-mediated hypoperfusion following umbilical cord occlusion in preterm fetal sheep Justin M Dean, Alistair J Gunn, Guido Wassink, Sherly George, Laura Bennet

P20. Changes in force of contraction of the papillary muscle with increasing gestational age in the sheep fetus T. Spencer, J.L. Morrison, G. Posterino

P21. Classification of FHR Patterns according to risk of acidemia and evolution to higher risk patterns J.T. Parer and Tomoaki Ikeda

P22. Relation between ECG change and alteration of low frequency domain in the spectral analysis of R-R intervals in the progress of fetal acidemia. Kimura Y, Sugawara J, Imai N, Chisaka H, Okamura K

P23. Circadian Rhythm in Breathing Characteristics in Fetal Lambs Marta Wlodarska, John Challis, Stephen Lye, William Gibb, Nancy Gruber, Dan Rurak

P24. Temporal antenatal testing alterations in severe IUGR fetuses EV Cosmi, G Mari and E Cosmi

P25. The Pituitary-Adrenal Axis In The Lamb Is Re-set Following Placental Restriction In Utero S.L. Dunn, J.A.Duffield, I.C. McMillen and C.L. Coulter

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P26. Changes observed in Doppler studies of the fetal circulation in pregnancies complicated by pre-eclampsia or intrauterine growth restriction. Nayana Parange, Gustaaf Dekker, Chris Wilkinson.

P27. Ovine Placental Igf2 and Igf2r Gene Expression is Reduced by Repeated Antenatal Betamethasone Treatment Roberts CT, Moss TJ, Button JJ, Nitsos I, Harding R, Newnham JP

P28. Accelerated Apoptosis and Differentiation of Trophoblasts Isolated from Intrauterine Growth Restricted Pregnancies Sarah Newhouse, Gary Chan, Sandra Davidge, Larry Guilbert.

P29. Hypoxic/Ischemic models in newborn piglet: a constant versus variable approach S. Tracey Bjorkman, Kelley A. Foster, Stephanie M. O’Driscoll, Genevieve N. Healy, Barbara E. Lingwood and Paul B. Colditz

P30. Adenosine Receptor Modulation Effects on Behavioural State Activity and Cerebral Blood Flow in the Ovine Fetus Near Term Neesha Rao, Shannon Hemstreet, Brad Matushewski and Bryan Richardson

P31. Effect of sex, size at birth and metyrapone inhibition of cortisol production on body composition in young adult guinea pigs. Sanita Grover, Cathie L. Coulter, Melissa R. Walker, Karen L. Kind, Jeffrey S. Robinson and Julie A. Owens

P32. Uterine vessel ligation induces placental restriction and impairs mammary function but does not impact on perinatal growth following a second pregnancy Westcott, K.T. and Wlodek, M.E.

P33. In Vitro Embryo Culture Affects Ovine Placental Gene Expression Fletcher CJ, MacLaughlin SM, McMillen IC, Walker S, Sibbons J, Roberts CT

P34. Hypertension in adult male rats is prevented by cross-fostering a growth restricted pup onto a mother with normal lactation. Mibus, A.L., Westcott, K.T., O'Dowd, R., Owens, J.A. and Wlodek, M.E.

1530 – 1600 Afternoon Tea Ballroom 1

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1600 – 1745 Free Oral Communications 3 – LUNG DEVELOPMENT / PLACENTA

Chairs: Richard Harding (Australia) and Claire Roberts (Australia)

Ballroom 2 & 3

1600 – 1615 17. A Genomic Analysis of the Development of the Neuroendocrine Components of the Fetal Hypothalamus-Pituitary-Adrenal Axis. UCharles E. WoodU and Maureen Keller-Wood

1615 – 1630 18. In Utero Ventilation: A novel model for investigating neonatal lung disease. UAllison B.A.,U Crossley K.J., Flecknoe S.J., Morley C.J., Harding R., and Hooper S.B.

1630 – 1645 19. Prolonged Increases in Lung Expansion Alter Pulmonary Hemodynamics in Fetal sheep. UGraeme R. PolglaseU, Megan J. Wallace, David L. Morgan and Stuart B. Hooper.

1645 – 1700 20. Modulation of fetal pulmonary responses to intra-amniotic lipopolysaccharide by antagonism of the interleukin-1 receptor. UTJM Moss U, SG Kallapur, I Nitsos,M Ikegami, JP Newnham & AH Jobe.

1700 – 1715 21. Dynamic imaging of lung aeration at birth using X-ray phase contrast US.B. Hooper U, M.J. Wallace, K. Siu, M. Kitchen, N. Yagi, K. Uesugi, I. Williams, M. Morgan, C. Hall, S. Irvine, K. Pavlov, J. Whitley and R. Lewis

1715 – 1730 22. Effects of Sulfasalazine on Kynurenine – Potential Therapeutic Agent UP LigamU, U Manuelpillai, D Walker & EM Wallace.

1730 – 1745 23. 5α-reductase and human term parturition USheehan Penelope MU, Moses Eric K, Brennecke Shaun P

1800 Drinks and Nibbles Ballroom 1

1830 – 2000 Data Blitz Poster Session

(Selected Posters: maximum 2 slides, 1 minute talk, 2 minute questions, 1 minute break)

Chairs: Julia Pitcher (Australia) and Miles De Blasio (Australia)

Ballroom 2 & 3

1830 – 1833 DB1. Maternal dexamethasone treatment increases hepatic glycogen content but not G6Pase activity in the pregnant rat and fetus UKL Franko U, AJ Forhead, AL Fowden

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1834 – 1837 DB2. Differential effects of maternal cold exposure on mRNA abundance for the growth hormone secretagogue receptor (1α) and glycerol-3-phosphatase in adipose tissue of the newborn sheep. EA Butt, S Pearce, T Stephenson, ME Symonds and UH BudgeU

1838 – 1841 DB3. The Impact of Placental Restriction on the Expression of the Appetite Neuropeptide Regulators, NPY and POMC, in the Hypothalamus of the Fetal Sheep in Late Gestation. UBHA Chuang U, BS Muhlhausler, JL Morrison, SJ Williams, CL Adam, PA Findlay and IC McMillen

1842 - 1845 DB4. Timing of delivery of IUGR fetuses on the basis of hemodynamic changes UEV CosmiU, G. Mari and E. Cosmi

1846 – 1849 DB5. The vasoreactivity of small placental arteries and systemic vascular bed UNoriaki Imai U, Minori Saito, Junichi Sugawara, Yoshitaka Kimura, Kunihiro Okamura

1850 – 1853 DB6. Neonatal Gender Influences Microvascular Function in Normal Newborns Stark MJ, Clifton VL, UWright IMRU

1854 – 1857 DB7. Leptin infusion does not alter cardiomyocyte development in the sheep fetus in late gestation UK.BottingU, I.C. McMillen, J. Dorosz, J.L. Morrison

1858 – 1901 DB8. Role of Beta1-adrenergic receptor (β1AR) Pi3K/p70S6k signaling in the regulation of neonatal cardiomyocyte proliferation Yi-Tang Tseng, Naohiro Yano, Ting Zhao, Vlad Ianus, Joan P Stabila, Beth G McGonnigal, UJames F Padbury U

1902 – 1905 DB9. Effect of Suppression of Neurosteroid Synthesis on Asphyxia-Induced Fetal Brain Injury UYawno TU, Yan EB, Walker DW and Hirst JJ

1906 – 1909 DB10. Secondary post-asphyxial hypoperfusion is associated with suppression of cerebral metabolism and increased tissue oxygenation in near-term fetal sheep UJensen ECU, Bennet L, Hunter CJ, Power GC, and Gunn AJ.

1910 – 1913 DB11. Altered fetal brain development and growth following repeated, acute exposure to alcohol: potential mechanisms. UPenelope A DalitzU, Sandra M. Rees, Kathryn L. Gatford, Julie A. Owens, Megan L Cock, Philip N Henschke, Richard Harding

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1914 – 1917 DB12. Regional specificity of detection of ischemic changes on magnetic resonance imaging in the preterm fetal sheep Mhoyra Fraser, Laura Bennet, Chris E Williams, Peter D Gluckman, Alistair J Gunn, and Terrie E Inder.

1918 – 1921 DB13. Suppression of post hypoxic-ischemic EEG transients with delayed dizocilpine is associated with moderate striatal protection in the preterm fetal sheep Justin M Dean, Alistair J Gunn, Guido Wassink, Sherly George, Laura Bennet

1922 – 1925 DB14. Dose-response study of lipopolysaccharide-induced fetal brain injury in the guinea pig E.L. Harnett, M.A. Dickinson, and G.N. Smith.

1926 – 1929 DB15. Pituitary and Adrenocortical Responses to Acute Hypoxemia in Highland and Lowland Newborn Llamas Riquelme RA, Herrera EA, Sanhueza EM, Reyes VR, Ebensperger G, Ebensperger R, Parer JT, Giussani DA, Blanco CE, Hanson MA, Llanos AJ.

2000 – late Dinner

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Monday September 26, 2005 0830 – 1030 Free Oral Communications 4 - HYPOXIA

Chairs: David Walker (Australia) and Carina Mallard (Sweden)

Ballroom 2 & 3

0830 – 0845 24. Effect of maternal hypoxia during the preimplantation period on fetal and placental development in the mouse. Karen L Kind, Rebecca L Kelley, Jeremy G Thompson.

0845 – 0900 25. Abstract removed from this timeslot

0900 – 0915 26. Maintenance of Mean Arterial Blood Pressure in Chronically Hypoxic Fetal Sheep is not Dependent on Post Ganglionic Sympathetic Activation J.L. Morrison, I.C. McMillen

0915 – 0930 27. Physiological and neurodevelopmental effects of umbilical cord occlusion in fetal sheep exposed to chronic intrauterine inflammation I Nitsos, SM Rees, BW Kramer, R Harding, JP Newnham and TJM Moss

0930 – 0945 28. Regulation of Hypoxic Pulmonary Vasoconstriction in the Perinatal Period by Reactive Oxygen Species. Sewite Negash, Yuansheng Gao, J. Usha Raj.

0945 – 1000 29. The Effect of Acute and Chronic Induced Hypoxia on BDNF Protein Expression in the Ovine Fetal Brain Near Term H.Nishigori, D.Mazzuca, V.Han, R.Gagnon, and B Richardson.

1000 - 1015 30. Adensosine A2A receptors mediates respiratory roll-off response to hypoxia in awake lambs Brian J. Koos, Yoshikazu Kawasaki, Young-Han Kim, Fanor Bohorquez

1015 – 1030 31. A Framework for Standardized Management of Intrapartum FHR Patterns Tomoaki Ikdea, J.T. Parer, Junji Onishi, Shunichi Node, Hiroshi Sameshima and Tsuyomu Ikenoue

1030 – 1100 Morning Tea Ballroom 1

1100 – 1300 Free Oral Communications 5 – PERINATAL BRAIN INJURY

Chairs: Paul Colditz (Australia) and Brian Koos (USA)

Ballroom 2 & 3

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1100 – 1115 32. Melatonin Reduces Inflammation and Cell Death in Cerebral White Matter in the 0.65 Gestation Fetal Sheep Carina Mallard, Anna-Karin Welin, Pernilla Arvidsson, Bo Sultan, Henrik Hagberg, Pierre Gressens, Ingemar Kjellmer

1115 – 1130 33. Repeated Episodes of Umbilical Cord Occlusions Increase Vascular Endothelial Growth Factor (VEGF) Expression in the Cortical Gray Matter of the Near Term Ovine Fetus H.Hamrahi, A.Becks, K.Nygard, B.Richardson, V.Han, R.Harding and A.Bocking.

1130 – 1145 34. Response of fetal sheep to intra-amniotic E.coli in the presence or absence of umbilical cord occlusions. L Patrick, S Hemstreet, B Matushewski, J Homan, B Richardson and G Smith.

1145 – 1200 35. Time Course of Endotoxin Impairment of Cerebral Endothelial Function in Newborn Lambs Feng, YS, Zivkovic, S, McClure,L, Yu, VYH, Walker, AM

1200 – 1215 36. Acute Episodic Fetal Alcohol Exposure in Late Gestation Can Increase Apoptosis and Astrogliosis and Induce White Matter Injury in the Immature Brain Penelope A Dalitz, Richard Harding, Jhodie R Duncan, Megan L Cock, Philip N Henschke, Sandra M Rees.

1215 – 1230 37. The influence of birthweight for gestational age on neurodevelopmental outcome at 1 year of age in premature infants weighing <1000gms Lloyd J, Riley K, Wyatt J, Peebles D

1230 – 1245 38. Antenatal Glucocorticoids: Effects on Cerebral Substrate Metabolism and Blood Flow in the Preterm Ovine Fetus J McCallum, N Smith, M Schwab, P Nathanielsz, B Richardson

1245 – 1300 39. Birth weight and neurodevelopmental outcome of children at two years of age after planned vaginal delivery for breech presentation at term Molkenboer JFM, Roumen FJME, Smits LJM, Nijhuis JG.

1300 – 1345 Lunch Ballroom 1

1400 – 1700 FNPS Traditional Sporting Event

MEET IN THE STAMFORD GRAND HOTEL LOBBY AT 1345h

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1830 – late Dinner 1900h – Meet in Foyer at 1815h

BUSES DEPART FROM THE STAMFORD GRAND HOTEL FOR ADELAIDE SAILING CLUB AT 1830h

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Tuesday September 27, 2005 0845 – 1030 Free Oral Communications 6 – NUTRITION

Chairs: Karen Gibson (Australia) and Alison Forhead (United Kingdom)

Ballroom 2 & 3

0845 – 0900 40. Maternal Nutrient Restriction During Pregnancy Changes the Postnatal Pattern of Prostaglandin Regulation in the Rat Kidney K.A. Brennan, S.W. Reynolds, I. Christiaens, B. McCook, G. Kan, M.E. Symonds and D.M. Olson

0900 – 0915 41. Maternal Protein Restriction: Effects in the Heart of Offspring Later in Life Kyungjoon Lim, Monika A Zimanyi, M Jane Black

0915 – 0930 42. Periconceptional Nutrition and the Relationship Between Maternal Body Weight Changes in the Periconceptional Period and Fetal Kidney Growth in the Sheep SM MacLaughlin, SK Walker, DO Kleeman and IC McMillen

0930 – 0945 43. Periconceptional Undernutrition in the Ewe Down-Regulates Maternal, But Not Fetal, Adrenal ACTH Receptor, StAR and P450c17 mRNA in Early Gestation Frank H Bloomfield, Kristin L. Connor, Anne L. Jaquiery, Mark H. Oliver, John R.G. Challis and Jane E. Harding

0945 – 1000 44. Strain Differences in the Impact of Dietary Restriction on Fetal Growth and Pregnancy in Mice Brian Knight, Craig Pennell, Stephen Lye.

1000 – 1015 45. Reduced Physical Activity and ad libitum Nutrition in Early Postnatal Life Leads to Obesity, Glucose Intolerance and Insulin Resistance. Helen Budge, Terence Stephenson, Michael E Symonds, David Gardner.

1015 – 1030 46. Cross foster studies identify lactation and pup milk intake as critical to postnatal growth following placental restriction during pregnancy O’Dowd, R., Mibus, A.L. and Wlodek, M.E.

1030 – 1100 Morning Tea Ballroom 1

1100 – 1300 Free Oral Communications 7 - GLUCOCORTICOIDS

Chairs: John Newnham(Australia) and Karen Moritz (Australia)

Ballroom 2 & 3

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1100 – 1115 47. Developmental ontogeny of the glucocorticoid receptor and mitochondrial proteins in the ovine liver from mid-gestation through to early adulthood M. Hyatt, D. Yakubu, A. Mostyn, D. Walker, T. Stephenson and M.E. Symonds

1115 – 1130 48. Ontogeny of the spiny mouse placenta and the effects of dexamethasone treatment on its development Hayley Dickinson, David W Walker, Karen Moritz, Claire Roberts

1130 – 1145 49. Effects on lung structure of Intra-amniotic corticosteroids for preterm lung maturation in sheep. G.R. Polglase, I. Nitsos, A.H. Jobe, J.P. Newnham & T.J.M. Moss

1145 – 1200 50. Effects of late gestational betamethasone administration on fetal and postnatal weights and insulin-like growth factor (IGF) concentrations Deborah M Sloboda, Shaofu Li, John RG Challis, Timothy JM Moss, John P Newnham

1200 – 1215 51. Differential effects of maternal dexamethasone treatment on thyroid hormone activity in the pregnant ewe and fetus Alison J Forhead, Juanita K Jellyman, Dino A Giussani and Abigail L Fowden

1215 – 1230 52. Maternal Exposure to Glucocorticoids in Early Pregnancy Causes Fasting Hyperglycaemia and Hyperinsulinaemia in Adult Male Sheep Offspring De Blasio MJ, Dodic M, Wintour-Coghlan EM, Robinson JS, Owens JA.

1230 – 1245 53. Effects of Antenatal Glucocorticoids on Development of Fetal Brain Function M. Schwab, K. Schwab, T. Groh, M. Kott, P.W. Nathanielsz and O.W. Witte

1245 – 1300 54. Prenatal corticosterone exposure in the rat results in hypertension and low nephron number. Reetu R Singh, Luise A Cullen-McEwen, Michelle M Kett, Wee-Ming Boon, Karen M Moritz, John F Bertram

1300 – 1430 Lunch Ballroom 1

1400 - 1430 Annual General Meeting Ballroom 2 & 3

1430 – 1630 Free Oral Communications 8 – FETAL AND NEONATAL PROGRAMMING

Chairs: Charles Wood (USA) and Tim Moss (Australia)

Ballroom 2 & 3

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1430 – 1445 55. The Cytokine Response to Endotoxin is Altered in Early Life following a Prenatal Immune Challenge Nicolette A. Hodyl, Klara M. Krivanek and Deborah M. Hodgson

1445 – 1500 56. Neonatal Leptin Treatment Reverses Developmental Programming Vickers MH, Gluckman PD, Coveny AH, Hofman PL, Cutfield WS, Gertler A, Breier BH, Harris M.

1500 – 1515 57. Influence of Maternal Parity on mRNA Abundance for the Prolactin and Insulin-like Growth Factor Receptors and Adipose Tissue Deposition in the Resulting Offspring J. Bispham, T. Stephenson, H. Budge and M.E. Symonds

1515 – 1530 58. Insulin Resistance Following Fetal Growth Restriction: Is Intramyocellular Obesity to Blame? Melanie Tran, Irving Lee, Robyn Taylor, Miles De Blasio, Jeffrey S Robinson and Julie A Owens

1530 – 1545 59. Increased IGF Bioavailability and Altered IGF Binding Proteins During Catch-up Growth in the Young Lamb Following Placental Restriction. Kathryn L Gatford, Brenton D Bennett, Miles J De Blasio, Patricia A Grant, Robyn L Taylor, Jeffrey S Robinson and JA Owens

1545 - 1600 60. Dietary Zinc Supplementation Ameliorates LPS-Induced Teratogenicity In Mice JSC Chua, AM Rofe and P Coyle

1600 – 1615 61. Dietary Zinc Prevents Birth Abnormalities and Postnatal Death Caused By Prenatal Ethanol Exposure BL Summers, AM Rofe and P Coyle

1615 – 1630 62. Maternal IGF-II Treatment in Early Pregnancy Improves Placental Structural and Functional Development and Fetal Growth Near Term AN Sferruzzi-Perri, JS Robinson, CT Roberts

1630 – 1700 Afternoon Tea

Registration for attendees of Jeffrey Robinson Symposium only

Ballroom 1

1700 – 1800 Geoffrey S Dawes Lecture

Chair: Professor Bill Parer (USA)

“Fatal Fetal Growth” Professor Jeffrey Robinson (Australia)

Ballroom 2 & 3

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1930 – midnight Combined FNPS / JSR Gala Dinner

(1930h drinks for 2000h)

END OF FETAL AND NEONATAL PHYSIOLOGICAL SOCIETY PROGRAM

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2005 FETAL AND NEONATAL PHYSIOLOGICAL SOCIETY

JEFFREY ROBINSON SYMPOSIUM

A Satellite Meeting of the 2005 FNPS meeting, Adelaide, Australia A symposium in honour of the research contributions of Professor Jeffrey Robinson.

Tuesday September 27, 2005 1700 – 1800 Geoffrey S Dawes Lecture

Chair: Professor Bill Parer (USA)

The ‘Fatal Fetus’

Professor Jeffrey Robinson (Australia)

1930 – midnight Combined FNPS / JSR Gala Dinner

(1930h drinks for 2000h)

Stamford Grand

Ballroom

Wednesday September 28, 2005 0815 – 0830 Welcome

Dr Kathryn Gatford (Australia)

0830 – 1020 Session 1: Epidemiology

Chairs: Vivienne Moore (Australia) and Caroline Crowther (Australia)

Ballroom 2 & 3

0830 – 0910 Maternal Nutrition and Offspring Cardiovascular Risk

Ranjan Yajnik (India) Supported by Texas Peak Pty Ltd.

0910 – 0950 Clinical networks to enhance clinical care and research in neonatology

David Henderson-Smart (Australia)

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0950 – 1020 Free Oral Communications 1 Ballroom 2 & 3

0950 – 1005 1. Blood pressure during physiological stress in adults after prenatal exposure to the Dutch

famine

Rebecca Painter (Netherlands)

1005 – 1020 2. Thirty Percent Global nutrient Restriction Slows Development of the Cortical Neuronal

Network in Fetal Baboons at 0.5 of Gestation

Matthias Schwab (Germany)

1020 – 1100 Morning Tea Ballroom 1

1100 – 1250 Session 2: Hypothalamic Pituitary Adrenal Axis, Programming and Placenta

Chairs: Graham Jenkin (Australia) and Marelyn Wintour (Australia)

Ballroom 2 & 3

1100 – 1140 Turning on the Uterus

Roger Smith (Australia)

1140 – 1220 The Placenta: Active or Passive Participant in Programming?

Les Myatt (USA)

1220 – 1250 Free Oral Communications 2 Ballroom 2 & 3

1220 – 1235 1. Male seminal factors influence fetal and placental development, postnatal growth and adult

onset obesity in mice.

John Bromfield (Australia) – The DSL Young Scientist Awardee

1235 – 1250 2. The Effect of Hypoxia to Promote Human Cytotrophoblast Outgrowth is Mediated by Insulin-

like Growth Factor-II

Amanda Sferruzzi-Perri (Australia) – The JSR Young Scientist Awardee

1250 – 1305 3. Influence of Placental Restriction and the Lactational Environment on Vascular Function in

Adulthood

Marianne Tare (Australia)

1305 – 1500 Lunch and Poster Viewing.

Lunch is sponsored by DSL Australia

Ballroom 1

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2005 FETAL AND NEONATAL PHYSIOLOGICAL SOCIETY

1500 – 1650 Session 3: Fetus and Neonatal Growth and Growth Factors

Chairs: Caroline McMillen (Australia) and Richard Harding (Australia)

Ballroom 2 & 3

1500 – 1540 What should women eat during pregnancy, and does it matter?

Jane Harding (New Zealand)

1540 – 1620 The plastic pancreas: Early developmental decisions with life-long consequences

David Hill (Canada) - Faculty of Health Sciences Senior Visiting Research Fellowship

1620 – 1650 Free Oral Communications 3 Ballroom 2 & 3

1620 – 1635 1. Molecular markers of early-stage lung injury in ventilated, very preterm lambs.

Megan Wallace (Australia)

1635 – 1650 2. Maternal nutrient restriction in late gestation up-regulates mRNA abundance for the insulin

receptor and peroxisome proliferator-activated receptor gamma in adipose tissue of newborn

sheep.

Helen Budge (United Kingdom)

1650 – 1705 3. Maternal Overnutrition Programmes Postnatal Appetite Regulation: Increased Nutrition

Before Birth Increases Hypothalamic Expression of the Appetite-Inhibiting Neuropeptide

POMC in the Neonatal Lamb

Beverly Muhlhausler (Australia) – The DSL Young Scientist Awardee

1705 – 1800 Round Table Discussion and Drinks

Chairs: Julie Owens (Australia) and Laura Bennet (New Zealand)

Ballroom 2 & 3

1700 – 1720 Eugenie Lumbers (Australia) Summation

1720 – 1740 Peter Gluckman (New Zealand) Perspective

1740 – 1800

John Challis (Canada) Discussion - Florey Foundation Visiting Scientist.

1900 – midnight Jeffrey Robinson Symposium Dinner

END OF JEFFREY ROBINSON SYMPOSIUM PROGRAM

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Plenary and Keynote Speakers

Geoffrey Dawes Lecture:

Professor Jeffrey Robinson

Keynote Speakers

Chittaranjan Yajnik (India)

David Henderson-Smart (Australia)

Roger Smith (Australia)

Les Myatt (USA)

Jane Harding (NZ)

David Hill (Canada)

Eugenie Lumbers (Australia)

Peter Gluckman (NZ)

John Challis (Canada)

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32ND FNPS 2005 ORAL ABSTRACTS

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Perirenal Structure and Function

Free Oral Communications 1 Chairs: Mary Wlodek (Australia) and Jane Black (Australia)

Sunday September 25, 2005 0830 - 1030

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O1. Upregulation of adrenal expression of the type 1 VEGF receptor in a model of activated-adrenal growth in the late gestation fetal sheep K.E. Warnes, I.C. McMillen, J.S. Robinson, C. L. Coulter. The development of a vascular network is a fundamental requirement for adrenal growth and functional differentiation and yet our understanding of the intra-cellular mechanisms which underlie the essential process of angiogenesis, remain unknown. Vascular endothelial growth factor (VEGF) and angiopoietins (ang) are key members of a family of angiogenic factors, which regulate a number of endothelial cell effects including proliferation, differentiation, migration, survival and vessel tube formation. VEGF mediates these biological effects through two high affinity tyrosine kinase receptors VEGF-R1 and VEGF-R2. Previously, VEGF expression has been detected in the human fetal adrenal gland and VEGF expression increases in human fetal adrenal cells in vitro following administration of ACTH. However, it remains to be determined whether VEGF plays a key role in angiogenesis during fetal adrenal development in vivo. To elucidate the potential molecular mechanisms involved in adrenal angiogenesis in a model of activated adrenal growth, we have measured the level of expression of VEGF, VEGF-R1 & R2 and angiopoietin-2 mRNA in adrenals from fetuses infused with a competitive inhibitor of cortisol biosynthesis (metyrapone) (n=8) or vehicle (n=6) for 15 days in late gestation. Recently, we have demonstrated that intrafetal infusion of metyrapone suppresses cortisol synthesis, increases circulating ACTH and results in a 2 fold increase in adrenocortical growth in the late gestation sheep fetus. In the present study, VEGF mRNA expression was significantly lower (P<0.05) in adrenals from metyrapone-infused (mean +/- SE: 8245.0 +/- 459.2) compared to vehicle-infused fetuses (10445.0 +/- 650.7). In contrast, adrenal expression of VEGF-R1 mRNA was significantly higher (P<0.05) in the metyrapone- (14342.5 +/- 974.2) than in the vehicle-infused (6916.0 +/- 926.6) fetuses. There was no difference, however, in adrenal expression of VEGF-R2 mRNA between metyrapone- (57.0 +/- 6.1) and vehicle-infused (45.8 +/- 3.4) fetuses or angiogenic inhibitory protein ang-2 (Vehicle: 1029.8 +/- 254.1; Metyrapone 949.6 +/- 127.6). In summary, this model of metyrapone-activated adrenal growth results in a suppression of VEGF mRNA, and a concomitant increase in VEGF-R1 mRNA. Thus, the increase in the growth of the vascular network which is required to support adrenal growth and functional differentiation in late gestation maybe mediated via a specific upregulation of the type 1 VEGF receptor. O2. VEGF and eNos in the ovine placenta are upregulated with altered maternal adrenal steroids Jensen E, Wood CE, and Keller-Wood M Late gestation fetal sheep studies have shown that reduced maternal cortisol or aldosterone levels alter placental morphology, with an increase in the ratio of fetal to maternal vascular tissue in the placentome, but overall reduced placental blood flow. We have now tested the hypothesis that vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (eNOS) mediate fetal blood vessel overgrowth in the placentome in response to relative adrenal hypoadrenalism. Pregnant ewes with singleton fetuses at 112 to 115 days gestation were allocated into 4 groups; controls (n=7, intact adrenals), normal cortisol (n= 4, ewe was adrenalectomized and replaced with normal cortisol and aldosterone levels), low cortisol (n=6, ewe was adrenalectomized and replaced with low cortisol levels), and low aldosterone (n=6, ewe was adrenalectomized and replaced with low aldosterone levels). At 130 + 0.2 days gestation the ewe was killed. The placenta was dissected and categorized into A, B or C type placentomes. There were significantly more B and C type placentomes in the adrenalectomized groups than in controls (p<0.001). Overall, B types had more VEGF mRNA than A types (p<0.05), with C types having an intermediate amount. VEGF protein levels corresponding to a 23 kDa band in A type placentomes were significantly higher in low aldosterone animals than in the other groups (p<0.05). In addition, VEGF protein levels corresponding to a 23 kDa band in C type placentomes were higher in low aldosterone than normal cortisol animals. VEGF protein levels corresponding to a 47 kDa band were higher in C type placentomes than A types, but were not significantly different from B types; protein levels were also higher overall in low cortisol animals compared to controls. Fetal eNOS protein levels were lower in the adrenalectomized groups than in controls (p<0.05). In conclusion, our results indicate that VEGF induction was associated with fetal tissue overgrowth in the placenta when the pregnancy-induced surge in adrenal steroids was prevented in the ewe. Despite the increase in VEGF and fetal vascular tissue, eNOS was suppressed with reduced maternal adrenal steroids, which is consistent with the reduced placental perfusion previously observed in this model.

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O3. The effects of furosemide on lambs born to mothers with renal dysfunction AE O’Connell, AC Boyce and KJ Gibson. We have developed a model of maternal renal disease in which non-pregnant ewes underwent subtotal nephrectomy (STNx) prior to mating. One kidney was removed and the remaining kidney was partially infarcted. We have shown previously that fetuses of STNx mothers appear to be volume expanded (1), however this did not persist after birth (2). This study aimed to determine if there were any differences in renal function in newborn offspring of STNx ewes. Lambs from STNx mothers (STNxL) and intact mothers (ConL) underwent surgery 3-7 days after birth. Catheters were placed in the femoral artery and vein, and bladder. Experiments were carried out 4-9 days later. After a 90min baseline period, furosemide (2mg/kg) was given i.v. and renal function was monitored for another 90min. Experiments were carried out in 9 ConL and 8 STNxL. There was no difference in age between the two groups, however, because there were more singletons in the STNxL, their body weights were higher (p=0.053). Most variables measured were not different between the two groups in the control period, however there was a suppression of fractional sodium reabsorption by the proximal tubule (FRNaP) which was compensated for by the distal tubule (FRNaD, p<0.05). STNxL and ConL responded similarly to furosemide with a naturesis (p<0.05) and diuresis (p<0.005), and glomerular filtration rate (p<0.001) was increased in both groups at 30min. There were some differences between the two groups in relation to potassium handling (less was excreted in the STNxL group, p<0.05) and tubular sodium handling (FRNaP was suppressed further and FRNaD increased more in the STNxL, p<0.05). It appears that after birth lambs from STNx mothers are able to rid themselves of the fluid they accumulated in utero and have relatively normal renal function, although the suppression of proximal sodium reabsorption that occurred in utero persists. When renal function was challenged by furosemide, subtle differences in sodium and potassium handling were brought to light. What consequences this may have later in life are under investigation. 1. Gibson et al. (2003). PSANZ 7th Annual Conference, A16. 2. O’Connell et al. (2005). 19th National Workshop on Fetal and Neonatal Physiology, A21 O4. The effects of maternal renal dysfunction on mother and fetus. Eugenie R Lumbers, Karen J Gibson, Clare L Thomson, Amanda C Boyce, Bilal M Karime. Moderate to severe maternal renal disease is associated with maternal hypertension, prematurity and growth retardation. In indigenous Australians in particular, there is a high incidence of maternal renal dysfunction during pregnancy, of low birth weight infants and of hypertension and renal disease in the adult population. To study the effects of maternal renal dysfunction on mother and fetus, a maternal kidney was removed and a branch of the renal artery to the remaining kidney ligated in non-pregnant cross-bred Merino ewes 2.5-17 months before these ewes (STNx) and a control intact group of ewes were time-mated. At 119-120 days gestation, ewes and their fetuses were cannulated under general anesthesia (1-2% halothane in oxygen) after induction with i.v. 0.8-1g of sodium thiopentone. Compared with intact pregnant ewes, STNx ewes had hypertrophy of the remaining renal mass; their effective renal plasma flows and GFRs relative to body weight were about 55%. They excreted more protein. They had higher arterial pressures and left ventricular hypertrophy. STNx ewes had higher plasma creatinines but were hypochloremic compared to intact ewes. Fetuses carried by STNx ewes had lower haematocrits, plasma chloride (P=0.01) and creatinine levels (P<0.01). Their plasma sodium levels also tended to be lower but not significantly so (P<0.1). The fetal: maternal plasma creatinine ratio was significantly lower in STNx fetuses compared with intact fetuses (1.8 + 1.5, n=8 compared with 2.5 + 0.29, n=7, P<0.05) but there were no differences in the fetal: maternal ratios of plasma sodium, potassium, chloride, haematocrits between the two groups of fetuses. Thus maternal renal dysfunction, affected the composition of maternal blood supplying the uterus, adversely affected maternal blood pressure and affected the composition of fetal blood. These changes in fetal and maternal electrolytes and in maternal blood pressure may affect fetal development and program the fetus for renal and cardiovascular disease in adult life.

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O5. The renal handling of sodium and the renal response to amino acid infusion is altered in fetuses whose mothers have renal dysfunction Karen J. Gibson, Amanda C. Boyce, Bilal M. Karime and Eugenie R. Lumbers To determine whether maternal renal dysfunction and its associated changes in fetal blood composition altered fetal renal function, studies were carried out in 9 chronically catheterized fetal sheep whose mothers had undergone subtotal nephrectomy at least 2 months prior to mating (STNxF) and in 7 control fetuses (ConF). Fetal renal function was studied at 126-128 days gestation (term = 150 days) under baseline conditions and then during fetal i.v. infusion of amino acids (alanine, glycine, proline and serine) at 0.32 mmol/l for 1 h and then at 0.64 mmol/min for 2 h. Amino acids were used to challenge the kidney since they stimulate fetal glomerular filtration rate (GFR) (1). Under baseline conditions, STNxF had higher urine flow rates (1.240.15 ml/min) and sodium excretion rates (686 umol/min) than ConF (0.630.16, P=0.02 and 299, P=0.002 respectively). GFR, although tending to be higher in STNxF, was not different between the groups (3.60.6 vs 2.90.5 ml/min, n.s.). STNx fetuses had lower fractional reabsorption of sodium (83+2.5% vs 93.52.0%, P=0.01) and a lower fraction of sodium reabsorbed by the proximal tubule (41.14.6 vs 63.48.2%, P=0.03). Fractional reabsorption of sodium by the distal tubule was higher (P=0.04). In response to amino acid infusion, although both groups had a natriuresis (P<0.005), the rise in sodium excretion was 10 fold in ConF and only 3 fold in STNxF. In ConF, arterial blood pressure rose (P<0.005) and GFR rose (P=0.01) reaching 5.30.9 ml/min by the 3rd hour. Fractional reabsorption of sodium fell to 75.03.5% (P=0.003) due to a decrease in the fraction reabsorbed by the proximal tubule (P=0.04). By contrast, in the STNx fetuses there was no change in arterial pressure or GFR but fractional reabsorption of sodium fell to 61.84.2% (P<0.005). It is concluded that STNxF excrete more sodium than ConF, possibly because they are volume expanded. The absence of a change in GFR in response to amino acid infusions suggests that these fetuses might already be operating at maximum GFR. This finding has implications for renal function after birth when GFR normally rises. (1) Marsh et al. (2002). J Physiol. 540, 717-728. O6. Renal function in adult sheep following prenatal exposure to glucocorticoids Karen Moritz, Andrew Jefferies, Miodrag Dodic, Marelyn Wintour and Robert DeMatteo Sheep exposed prenatally to glucocorticoids (dexamethasone or cortisol) develop high blood pressure in adulthood. We have previously shown that this is associated with a significant decrease in the number of glomeruli (and thus nephrons) within the kidney. In addition, we have found an upregulation of the renal renin-angiotensin system (RAS) in the late gestation fetus along with increases in the renal sodium transporters. It is unknown if these changes result in alterations in basal renal function in the adult animal. Single female offspring at 4-5 years of age were used in this study. Animals had been exposed in utero to saline (n=5), dexamethasone (0.48mg/h, n=5) or cortisol (5mg/day, n=5) for 2 days between 26-28 of gestation. On the day prior to experimentation, animals were instrumented with catheters in the carotid artery, jugular vein and bladder. After a one hour equilibration period, glomerular filtration was measured using 51-Cr-EDTA and renal plasma flow was measured using PAH. Three twenty minute collections were made and averages over the hour obtained for each animal. Results are shown in the table below: Parameter Saline Dexamethasone Cortisol GFR (ml/kg/h) 113+/-4 117+/-5 110+/-5 Urine Flow (ml/min) 3.8+/-0.3 3.0+/-0.3 2.9+/-0.3 Renal plasma flow (ml/min) 570+/-54 805+/-82 630+/-66 Urinary Na (mmol/l) 64+/-8 70+/-8 54+/-7 Urinary K (mmol/l) 54+/-9 67+/-10 137+/-34 Urinary Cl (mmol/l) 46+/-4 45+/-9 74+/-9 Urinary Na/K ratio 1.6+/-0.3 1.2+/-0.2 0.7+/-0.2 Urinary Na excretion rate 267+/-40 243+/-34 167+/-36 Urinary K excretion rate 189+/-19 209+/-23 251+/-18 Overall, there were no statistical differences between any parameter in the 3 groups. However, the cortisol group tended to excrete less sodium and more potassium (P=0.1) suggesting subtle differences in renal function may be present in these animals. In conclusion, a reduction in nephron endowment of approximately 30% induced by prenatal glucocorticoid exposure does not result in changes in basal renal function in the adult sheep. It is speculated that under altered conditions such as in response to increased sodium intake, alterations in renal function may become apparent.

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O7. The tubuloglomerular feedback mechanism in the developing kidney URussell D Brown U, Anita J Turner, A Erik G Persson, Karen J Gibson The tubuloglomerular feedback (TGF) mechanism plays an important role in regulating single nephron GFR by coupling distal tubular flow to afferent arteriolar tone. The prenatal kidney is known to have high vascular tone and low GFR. After birth, vascular tone decreases causing increased GFR. It is not known, however, if the TGF mechanism is active during fetal and neonatal life and whether it plays a role in maintaining vascular tone and GFR during this period. Therefore, in this study we have investigated TGF characteristics before and after birth. Renal micropuncture studies were carried out in ovine fetuses (133-140 days of gestation) and lambs (12-17 days after birth) during euvolemia and acute volume expansion. Lambs and pregnant ewes were anaesthetized and the fetus was delivered through a cesarean incision into a heated water bath (39P

oPC), with the umbilical cord

intact. The left kidney of the fetus/lamb was exposed through a subcostal flank incision and prepared for micropuncture. TGF activity was evaluated by measuring changes in proximal tubular stop-flow pressure (PSF) in response to various perfusion rates of the loop of Henle. Mean arterial pressures in the fetuses and lambs were 540.6 and 751.2 mmHg, respectively. In the fetus, free-flow pressure (PFF) was found to be 6.5 U+U 0.6 and PSF 29.3 U+U 0.73 mmHg. Maximal change in PSF (∆PSF) was 4.9 U+U 0.9 with the turning point (TP) at 16.3 U+U 1.3 nl/min. Volume expansion increased the TP by ~5 nl/min but did not affect PFF, PSF or ∆PSF. In lambs, PFF was 9.0 U+U 0.3 and PSF 34.0 U+U 0.6 mmHg. ∆PSF was ~2-fold higher (10.6 U+U 0.6 mmHg) than in the fetuses and the TP was 19.4 U+U 1.1 nl/min. Volume expansion decreased ∆PSF to 4.3 U+U 0.4 nl/min and increased the TP to 28.3 U+U 1.7 mmHg. Our results show that the TGF mechanism is active in the prenatal kidney. The lower TP indicates a more sensitive TGF to increased tubular flow than that found in the lamb. The sensitive TGF could in part be responsible for the increased renal vascular resistance and decreased GFR seen in the fetal kidney.

O8. The effect of pre-term birth on nephrogenesis UGubhaju LU, Zulli, A, Black MJ Nephrogenesis, the formation of nephrons in the kidney, occurs mainly during late gestation of pregnancy with no new nephrons formed after term in humans. Hence, it is imperative to investigate the effects of pre-term birth on kidney development. We have recently shown that the baboon is a suitable experimental model of human nephrogenesis. The aims of the present study were to morphologically assess whether nephrogenesis continues after premature delivery in pre-term baboons and if so, to determine whether nephrogenesis is affected by premature delivery. Delivery and care of all fetal baboons were undertaken at the Southwest Foundation (San Antonio, Texas, USA). Fixed kidneys from premature fetal baboons (n = 8) delivered at 125 days gestation and ventilated for 6, 14 and 21 days were morphologically assessed and compared to kidneys of fetal baboons from normal gestation (n = 8) delivered at 125, 140, 175 and 185 days gestation. Immunohistochemistry was also undertaken using the endothelial cell marker, CD31. Nephron number was estimated using unbiased stereological techniques. Morphological assessment confirmed that nephrogenesis in the baboon was on-going at 125 and 140 days during normal gestation and was complete by 175 days gestation. Importantly, morphological assessment confirmed that nephrogenesis continues after premature delivery until 21 days. Nephron endowment in pre-term fetal baboons did not appear to be compromised. There was also a highly significant correlation between kidney weight and nephron number in the kidneys from fetal baboons in normal gestation (R2=0.918, P=0.0002) and in the kidneys from prematurely delivered baboons (R2=0.750; P=0.001). However, an interesting observation from the current study was the appearance of apparently abnormal glomeruli in kidney sections from the premature baboons. These glomeruli in the outer renal cortex exhibited a dilated Bowman’s space and a shrunken glomerular tuft. Immunohistochemical studies with CD31 demonstrated that these glomeruli were not well vascularised, although they appeared to be the same size as mature glomeruli. In conclusion, these findings demonstrate that although nephron endowment may not be compromised in premature neonates it appears that there may be some pathological changes in the glomeruli, possibly due to abnormal development and maturation of the glomerular capillaries.

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CARDIOVASCULAR

Free Oral Communications 2 Chairs: Eugenie Lumbers (Australia) and Janna Morrison (Australia)

Sunday September 25, 2005 1100 - 1300

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O9. Growth and gene expression of components of the renin angiotensin system in offspring of rabbits with chronic maternal hypertension Devaki Maduwegedera, Karen Moritz, Rebecca L Flower, E Marelyn Winour, Kate M Denton Chronic hypertension complicates up to 5% of all pregnancies, an incidence expected to rise following the demographic trend for women to have children in later life. It has been suggested that permanent alterations in fetal development due to adverse conditions such as chronic hypertension ‘program’ the fetus. We have previously shown using a two kidney, one wrap (2K-1W; renin dependent) model of hypertension that adult female offspring of hypertensive mothers have increased blood pressure. These offspring of hypertensive mothers are also proportionately larger than offspring of normotensive mothers. To better understand the developmental process that may pre-dispose offspring of chronically hypertensive mothers to develop hypertension in later life, the aims of this study were to characterise growth and gene expression of the renal renin angiotensin system (RAS) during development. Three groups of rabbits were used, 2K-1W hypertensive (n=18); two kidney, two wrap (2K-2W; renin independent, n=21) hypertensive, and sham operated (n=20) normotensive mothers. Four weeks following surgery rabbits were mated with normotensive males. Biometric measurements of offspring were made and kidneys collected for determination of gene expression levels at gestational age (GA) 21, 28, birth, and 5 weeks of age. Metanephric development begins at GA11.5 in the rabbit and is completed by postnatal day 21. Expression of AT1, AT2 and renin was studied using real-time PCR. Body and kidney weights were similar between the groups at GA21 and 28. However, 2K-1W offspring demonstrated 20% greater birth weights and 30% greater kidney weights compared to sham and 2K-2W offspring. At 5 weeks there was no significant difference in body or organ weights between groups. Gene expression results from GA 21 and birth (Table) indicate differences in the ratio of AT1 to AT2 receptors in the kidneys of 2K-1W offspring at birth, and 2K-2W offspring in early gestation. GA21 Birth

Sham (13) 2K-1W (12) 2K-2W (12) Sham (18) 2K-1W (10) 2K-2W (11) AT1 1.1+/-0.1 1.1+/-0.1 1.9+/-0.3# 1.4+/-0.3 1.0+/-0.1 1.4+/-0.3 AT2 1.1+/-0.1 1.0+/-0.1 1.2+/-0.1 1.0+/-0.2 1.5+/-0.2# 1.4+/-0.3 AT1:AT2 1.1+/-0.1 1.3+/-0.1 1.5+/-0.1# 1.5+/-0.2 0.8+/-0.1# 1.1+/-0.1 Renin 1.2+/-0.2 0.9+/-0.1 1.6+/-0.2 1.2+/-0.2 1.3+/-0.2 1.5+/-0.3 # compared to sham at same age, P<0.05 Offspring of 2K-1W and 2K-2W hypertensive mothers show alterations in the key receptors of the RAS. Given the significant role of the RAS in kidney development and salt and fluid homeostasis, these changes may contribute to the development of adult hypertension in offspring of hypertensive mothers.

O10. Maternal Microvascular Response to Corticotrophin Releasing Hormone (CRH) is Altered By Gestation, Fetal Gender and Pre-eclampsia Stark MJ, Dierkx L, Clifton VL, Wright IMR AIMS: Normal pregnancy is characterized by hemodynamic modifications that peak in the second trimester, then plateau until delivery [1]. In pre-eclampsia (PE) the adaptation of vascular reactivity and tone in early pregnancy that precedes and may trigger these changes is altered [2]. Previous studies by our group have demonstrated significant maternal physiological differences dependant upon fetal sex [3]. We hypothesized that the influence of gestation and fetal gender on maternal peripheral microvascular function would be altered in PE. METHOD: Peripheral microvascular responses were examined using Laser Doppler flowmetry in normotensive pregnant women at 0-20 weeks (n=12), 21-30 weeks (n=31), and 31-40 weeks gestation (n=30) and in women diagnosed with PE at 31-40 weeks (n=23). Fetal sex was not known prior to delivery. Baseline perfusion, response to thermal hyperemia, and post-occlusive reperfusion were measured. Corticotropin-releasing hormone (CRH), a potent vasodilator, was administered transcutaneously by iontophoresis to a small area of skin in the forearm and blood flow measured simultaneously by laser Doppler. RESULTS: Gestation did not significantly influence maternal microvascular response within the normotensive women. At 31-40 weeks gestation those women with a male fetus exhibited increased vasodilation in response to CRH (p=<0.05) and greater baseline perfusion (p=0.04) than those with a female. The pre-eclamptic group exhibited significantly lower baseline perfusion (p=0.037), post-occlusive reperfusion (p=0.008), response to thermal hyperemia (p=0.001) and CRH induced dilation (p=<0.05) compared with the normotensive group. Pre-eclamptic women with a male fetus demonstrated a significantly reduced vasodilation in response to CRH (p=<0.05), reduced baseline perfusion (p=<0.05), and response to thermal hyperemia (p=<0.05) compared to normotensive women with a male fetus. Microvascular function was not significantly different between pre-clamptic and normotensive women with a female fetus. CONCLUSION: Fetal sex has a significant effect on the maternal microvascular response in both in normotensive and pre-eclamptic pregnancies. The observed baseline constriction and decreased vasodilatory response of the peripheral microvasculature in pre-eclamptic women is predominantly due to those pregnant with a male fetus. This supports a global endothelial dysfunction that may be a factor associated with the significantly greater neonatal morbidity and mortality observed in males born following pre-eclampsia. [1] M.P. Bosio PM, Conroy R, O'Herlihy C, Obstet Gynecol. 1999, 94, 978-984. [2] Brown MA, Clin Exp Pharmacol Physiol. 1995, 22, 781-791. [3] M.V. Clifton VL, Placenta. 2003, 168(Supplement A), S45-52.

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O11. Vitamin D insufficiency during early life is associated with raised blood pressure and altered vascular reactivity M. Tare, S.J. Emmett, R. Morley, C. Skordilis, H.A. Coleman, H.C. Parkington The incidence vitamin D (VitD) deficiency is increasing in western societies including Australia. Of increasing public heath concern is the prevalence of low VitD levels in women of reproductive age, especially those that are dark skinned or who dress modestly (1). Maternal VitD insufficiency has been linked to increased risk in the offspring of disease including autoimmune disease, some cancers, heart disease, and elements of Syndrome X. In this study we investigated the influence of VitD deficiency in utero and in early life on blood pressure and vascular function in young rats. Female Sprague Dawley rats were fed either VitD deficient (no added VitD) or control (1000U/Kg cholecalciferol) chow for 6 weeks prior to mating and maintained on the same diet during pregnancy and lactation. Pups were fed the same diet as their mothers until experimentation at 7 weeks of age. Conscious blood pressure was measured via a tail artery catheter. Rats were killed by decapitation and the mesentery isolated. To identify the stage of the estrous cycle in females, vaginal smears were taken, uterine weight recorded and ovaries examined. Leak-free segments of small mesenteric arteries were mounted onto a pressure myograph and pressurized to around 57mmHg without flow. Arteries were continuously superfused with warmed, oxygenated physiological saline and outside diameter was continuously monitored. Serum VitD levels were lower in rats on VitD deficient chow compared with controls (8+1, n=26 vs 126+10ng/ml, n=21). Serum Ca2+ was not different (control, 2.3 + 0.2 vs VitD deficient, 2.3 + 0.3mmol/L). Mean arterial blood pressure and heart rate were higher in the VitD group (by 93mmHg and 4013 beats/min, n=11, males; by 164mmHg and 299 beats/min, n=11, females). In mesenteric arteries from VitD deficient rats, the level of basal myogenic tone was doubled, endothelial nitric oxide mediated dilation was halved in males and diestrous females and endothelium-derived hyperpolarizing factor mediated dilation was all but abolished in estrous females. This study demonstrates that VitD insufficiency in early life is associated with cardiovascular dysfunction consisting of elevated blood pressure and heart rate and impaired vascular function. (1) Grover S & Morley R (2001). Medical Journal of Australia, 175:251-252.

O12. Fetal oxygenation and Doppler ultrasonography of cardiovascular hemodynamics in a chronic near term sheep model Kaarin Makikallio, Tiina Erkinaro, Niina Niemi, Tomi Kavasmaa, Ganesh Acharya, Mervi Pakkila, and Juha Rasanen Objective: We hypothesized that a decrease in fetal oxygenation without acidemia in a near term fetal sheep leads to cardiovascular hemodynamic changes detectable by Doppler ultrasonography. Study Design: Twelve ewes and fetuses were instrumented at 112-127 days of gestation. After a five-day recovery period, experiments were performed. Uterine and placental volume blood flows and fetal arterial and venous blood pressures were measured. Fetal cardiovascular hemodynamics were assessed by Doppler ultrasonography. All the measurements were performed at baseline, during fetal hypo-oxygenation and at recovery phase. Results: A drop in fetal Po2 was related to increased (p<0.05) weight-indexed right ventricular and combined cardiac outputs, and proximal branch pulmonary artery (PPA) pulsatility index (PI) values. The increase in PPA PIs correlated (R=0.59, p<0.05) with the decrease in fetal oxygen saturation. In the aortic isthmus, the time-velocity integral ratio between antegrade and retrograde blood flow components decreased (p<0.05) when fetal Po2 dropped. Conclusion: Decreased fetal oxygenation is associated with increased fetal cardiac output and pulmonary arterial vascular impedance, and with a relative increase in the retrograde blood flow component in the aortic isthmus.

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O13. Armed stem cells delivered to injured myocardium restore function and assume elements of the gene program regulating fetal/neonatal cardiomyocyte proliferation Ting Zhao, Joan P Stabila, Beth G McGonnigal, Yi-Tang Tseng, Naohiro Yano, Lawrence Lum, James F Padbury. Stem cell therapy for myocardial repair is limited by the number of stem cells that home to the injured myocardium and their ability to proliferate. To bypass this limitation, we have adapted a novel strategy of using a bivalent antibody to increase targeting of human stem cells to injured myocardium and facilitate repair. We induced injury in adult 20-30 gm ICR mice by ligation of the left anterior descending artery (LAD). Anti-CD3 x anti-myosin light chain bispecific antibody ‘armed’ adult human bone marrow derived CD34+ stem cells (5x105) were administered IV 2 days later (n=6). Control animals included sham thoracotomy (n=5) or animals with coronary ligation but no stem cells (n=6). 2 weeks following stem cells, animals were sacrificed, hearts removed and evaluated with the Langendorff isolated heart model. We showed improved myocardial performance in the animals receiving armed stem cells compared to the animals subjected to LAD ligation alone. This was reflected by enhanced left ventricular systolic pressure, enhanced developed pressure and better peak dP/dt at similar heart rates (p=0.005). Rate Pressure Product (RPP) is an indirect measure of myocardial work and also myocardial oxygen consumption. RPP was significantly higher in the mice that received armed stem cells than those that received no cells (p = 0.004). The greater myocardial performance (e.g. developed pressure) at comparable heart rate, suggests the increased RPP reflects enhanced myocardial oxygen utilization. Immunohistochemistry studies have shown that armed CD34+ cells specifically localized to the infarct region of the heart, co-localized with troponin T stained cells and co-localized with vascular structures. Most remarkably, we observed higher levels of expression of p70S6 Kinase in the peri-infarct zone in animals which received armed stem cells than those who were ligated alone. We suggest arming cells with bivalent antibody improves delivery of stem cells and improves functional outcome following myocardial injury. Additionally, the expression of p70S6Kinase in the ‘regenerating zone’ suggests recapitulation of the ‘fetal gene program’ regulating cardiomyocyte proliferation during development underlies the ability of adult cells to re-enter the cell cycle. These studies also provide evidence that arming human cells with bivalent antibodies is a non-invasive strategy that significantly enriches homing of cells to injured myocardium. O14. Increased sympathetic nerve activity to the cerebral circulation during blood pressure surges in REM sleep Priscila A Cassaglia, Robert I Griffiths & Adrian M Walker Introduction: cerebral sympathetic activity is believed to protect the brain against large increases in blood pressure, however, its role in sleep has never been studied. We have previously shown that drug-induced increases in arterial blood pressure (ABP) result in increased sympathetic nerve activity (SNA) within the superior cervical ganglion (SCG) to the cerebral circulation (1). In addition, removal of the SCG lowers baseline cerebrovascular resistance (CVR) and increases cerebral blood flow (CBF) during sleep, particularly in REM sleep (2). We hypothesize that increases in SNA may, through vasoconstriction, protect the cerebral microcirculation from natural increases in ABP, particularly the large increases which commonly occur in REM sleep. Methods: newborn lambs (n=2) were instrumented under halothane (2%) oxygen (50%) balance NO2 general anesthesia for recording ABP (femoral arterial catheter), CBF and determination of sleep-wake states (ECoG, EMG and EOG electrodes). Tungsten microelectrodes (25 m diameter with a 1 mm bared tip) were implanted in the SCG for measurement of SNA. After a minimum of 2 days recovery, physiological signals were recorded continuously while animals were undergoing spontaneous sleep-wake cycles. The SNA signal was differentially amplified (25000 times), filtered (100-2000 Hz) and stored (sampling rate 10 kHz) for off-line spectral analysis. Results: substantially increased SNA (average 30 + 7%) from baseline was observed as ABP (n = 7) rose acutely (average 41 + 6 %) during REM sleep, with the onset of the SNA increase preceding the natural ABP surge by 5 seconds. Conclusions: these studies provide preliminary evidence for increased activity of the cerebral sympathetic innervation in REM sleep. As increased activity precedes the onset of blood pressure surges, sympathetic nerves may play an important protective role in the cerebro-vascular microcirculation. (1) Cassaglia P. et al. (2004) 8th PSANZ (P12) Sydney Australia. (2) Loos N. et al. (2005) J Sleep Res 14(3)

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O15. Sympathetic nerve activity in the preterm fetal sheep is regulated by baroreflexes; evidence from telemetry based recordings USC MalpasU, L. Booth, CJ Barrett, A Gunn, L Bennet It is well established that in adult animals the sympathetic nervous system plays a critical role in the control of blood pressure and is linked to the pathogenesis of a number of cardiovascular diseases such as hypertension and heart failure. In the fetus growing evidence indicates an important role for sympathetic nerve activity (SNA) in regulating the long term consequences of perinatal events such as asphyxia. However to date there have only been limited attempts to directly record SNA in the fetus close to term. These have all involved either paralysing the fetus and or mother to prevent movement related artefacts. In this study we have utilised a novel telemetry based system (HTUwww.telemetryresearch.comUTH) to record renal SNA in the unparalysed

unanesthetized preterm fetal sheep at ~99 days gestation. Recordings obtained subsequent to surgery revealed distinct bursts of SNA indicative of synchronisation (Figure), where many individual nerve fibres fire at approximately the same time. Furthermore bursts occurred at a certain phase with the cardiac cycle (140 ms between diastolic pressure and the maximum SNA level). This phase relationship to the cardiac cycle is found in all adult animals and reflects phasic input from arterial baroreceptors regulating when a sympathetic discharge can occur. These phenomena are indicative of two key aspects; firstly that the central nervous system circuits generating and regulating SNA are well developed at this stage of gestation and secondly that arterial baroreflexes are capable of modulating the ongoing level of SNA. We propose that telemetry based recordings of SNA offers a new approach for examining the role of the sympathetic nervous system in the regulation of fetal pathophysiology. UFigureU Direct telemetry based recordings of SNA obtained in the fetal sheep 2 days after surgery.

O16. Fetal cardiac natriuretic peptide expression and cardiovascular hemodynamics in endotoxin-induced acute cardiac dysfunction in mouse Kaarin Makikallio, Samuli Rounioja, Olli Vuolteenaho, Jenna Paakkari, Mikko Hallman, and UJuha RasanenU

OBJECTIVE: We hypothesized that, in acute endotoxin-induced fetal cardiac dysfunction, atrial (ANP) and B-type (BNP) natriuretic peptide mRNA expressions are increased in proportion to the severity of fetal cardiovascular compromise in mouse. STUDY DESIGN: To investigate in vitro the effect of endotoxin-induced inflammation on cardiac natriuretic peptide expression, fetal hearts were harvested at 15-16 days of gestation and incubated for 6 hours with lipopolysaccharide (LPS). To examine the relationship between fetal cardiovascular compromise and cardiac natriuretic peptide expression in endotoxin-induced cardiac dysfunction in the in vivo model, fetuses received intra-amniotically 25 µl LPS (10 mg/ml) or 25µl of 0.9% saline. Fetal Doppler ultrasonography was performed before and six hours after the injections. RESULTS: In in vitro cultured fetal hearts, LPS induced the production of proinflammatory cytokines without affecting the basal expressions of natriuretic peptides. In the in vivo model, Doppler ultrasonography revealed severe cardiac dysfunction after LPS injection. No significant changes in ANP or atrial BNP mRNAs were found. The fetal ventricular BNP mRNA levels were about 2.6-fold (p<0.05) in the LPS group compared to the control group. Decreased fetal cardiac outflow mean velocity, increased proportion of isovolumetric contraction time of the cardiac cycle, and increased pulsatility indices of the descending aorta and inferior vena cava were related to elevated ventricular BNP mRNA levels. CONCLUSIONS: Our results show that LPS did not increase the mRNA expression of natriuretic peptides in cultured fetal hearts. In contrast, fetal ventricular BNP gene expression was increased in proportion to the severity of the hemodynamic compromise in vivo.

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LUNG DEVELOPMENT / PLACENTA

Free Oral Communications 3 Chairs: Richard Harding (Australia) and Claire Roberts (Australia)

Sunday September 25, 2005 1600 - 1745

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O17. A Genomic Analysis of the Development of the Neuroendocrine Components of the Fetal Hypothalamus-Pituitary-Adrenal Axis. Charles E. Wood and Maureen Keller-Wood The present study was performed to identify the genomic changes in expression of critical components of the hypothalamus-pituitary-adrenal axis in the second half of gestation in fetal sheep. We isolated mRNA from pituitary, hypothalamus, lung, skeletal muscle, and GI tract in fetal sheep that were 80, 100, 120, 130, 145 days gestation, and 1 and 7 days postnatal (n=4-5/group). Using real-time RT-PCR, we measured mRNA expression levels of proopiomelanocortin (POMC), corticotropin releasing hormone (CRH), arginine vasopressin (AVP), and urocortin-1 (URO). Pituitary POMC was increased at 100 days gestation and later compared to 80 days. In hypothalamus, AVP was more highly expressed than CRH, while URO was nearly undetectable. Hypothalamic AVP and CRH expression were increased at the end of gestation compared to younger ages, although in both cases this appeared to be a part of a progressive increase that continued postnatally. Hypothalamic POMC was increased at 120 days, reduced at later fetal gestational ages, then increased again postnatally. Lung expressed significant quantities of POMC, CRH, and URO mRNA in 80 day fetuses: expression in this tissue decreased throughout gestation, reaching a minimum expression in postnatal life. Expression of POMC in skeletal muscle and GI tract was low and unchanging throughout gestation. Overall, the results demonstrate that the expression of CRH, AVP, and POMC are developmentally-regulated and tissue-specific. While there are developmental changes in the patterns of expression of all these genes, there is no time at which a sudden increase in expression heralds parturition. Such changes are more likely to be acute, within hours of normal parturition.

O18. In Utero Ventilation: A novel model for investigating neonatal lung disease. Allison B.A., Crossley K.J., Flecknoe S.J., Morley C.J., Harding R., and Hooper S.B. Background: Very premature infants usually require ventilation from birth and many develop acute RDS and then chronic lung disease (CLD). These diseases are caused by ventilation-induced lung injury (VILI). CLD is characterized by changes in lung structure; including simplified alveoli, hypercellularity and alveolar fibrosis. The causes of VILI and the factors mediating the lung injury remain unknown. Animal models for investigating CLD use prolonged postnatal ventilation, making it difficult to delineate the causative factors in this disease, particularly if injury occurred straight after ventilation started. However, if the ventilation is performed in utero, the fetus can continue to be supported by the placenta thus allowing time for inflammation to manifest, but removing the need for further ventilation. Our aim was to develop a model of CLD in fetal sheep using in utero ventilation. Methods: A tracheotomy was preformed at surgery in fetal sheep at 105d GA (n=8; term ~147d) to insert an endotracheal tube which was connected to a neonatal ventilation circuit. At 110d GA, lung liquid was drained and fetuses were ventilated for 12 hrs using room air, at a PEEP of 4 cmH2O and a PIP of 40 cmH2O. The fetuses were then either euthanased or allowed to survive in utero for 7 days in which case lung liquid was replaced and normal liquid flow restored. At autopsy, lung tissue was examined for lung injury and arrest of alveolar development; unventilated age-matched controls were used for comparison. Results: Following ventilation for 12 hrs fetal lungs displayed non-uniform inflation, variable regions of interstitial cell hyperplasia and delayed development of secondary septa in all fetuses. Immunohistochemical analysis demonstrated marked interstitial cell proliferation, premature and markedly enhanced myofibroblast differentiation and abnormal deposition of collagen and elastin. Conclusions: Ventilation of fetal sheep in utero causes morphological changes that are similar to those seen in VILI and CLD. This model has the potential to delineate the individual ventilatory factors that cause lung injury as well as specific mechanisms leading to the arrest of lung development that characterizes CLD.

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O19. Prolonged Increases in Lung Expansion Alter Pulmonary Hemodynamics in Fetal sheep. UGraeme R. PolglaseU, Megan J. Wallace, David L. Morgan and Stuart B. Hooper. Although the degree of fetal lung expansion determines its growth and development, the effect of prolonged increases in lung expansion on pulmonary hemodynamics is unknown. Our aim was to determine the effect of prolonged increases in fetal lung expansion, induced by tracheal obstruction (TO), on pulmonary blood flow (PBF) and vascular resistance (PVR). Chronically catheterized fetal sheep (n=6) underwent TO from 120-127d gestational age (term ~147d); tracheas were not obstructed in control fetuses (n=6). PBF, PVR and changes to the PBF waveform were determined. TO significantly increased lung wet weight compared to control (166.3 +/- 20.2g vs 102.0 +/- 18.8g; p<0.05). Despite the increase in intraluminal pressure caused by TO (5.0+/-0.9 vs 2.4+/-1.0 mmHg; p<0.001), PBF and PVR were similar between groups after 7 days (TO; 28.1+/-3.2 vs Control; 34.1+/-10.0 mL/min/100g lung weight). However, TO markedly altered pulmonary haemodynamics associated with fetal breathing movements, causing a reduction rather than an increase in PBF after 7 days of TO. To account for the increase in intra-luminal pressure caused by TO, the intra-luminal pressure was equalized by draining the lungs of liquid on day 7 of TO. Pressure equalization increased PBF from 36.8+/-5.2 to 112.4+/-22.8 mL/min (p=0.01) and markedly altered the PBF waveform. These studies provide further evidence to indicate that intra-luminal pressure is an important determinant of PBF and PVR in the fetus. We suggest that the increase in PBF associated with pressure equalisation following TO, reflects an increase in growth of the pulmonary vascular bed leading to an increase in its cross-sectional area.

O20. Modulation of fetal pulmonary responses to intra-amniotic lipopolysaccharide by antagonism of the interleukin-1 receptor. UTJM MossU, SG Kallapur, I Nitsos,M Ikegami, JP Newnham & AH Jobe. Intra-amniotic (IA) injection of lipopolysaccharide (LPS) from E. coli causes fetal pulmonary inflammation and alters lung development in sheep, resulting in functional improvement after preterm birth due to structural changes and increases in surfactant. We hypothesised that the pro-inflammatory cytokine interleukin-1 (IL-1) is a critical contributor to the response to IA LPS; IL-1 increases in the fetal lungs after IA LPS and IA IL-1 has effects on the fetal lungs similar to those caused by IA LPS. We aimed to determine the effects of inhibition of IL-1 receptor signalling on the inflammatory and developmental responses of the fetal lungs to IA LPS or IA IL-1β. We studied singleton preterm lambs aged 125 days’ gestation; we assessed pulmonary inflammatory and developmental responses, 2d or 7d (respectively), after IA LPS (10mg), IA IL-1β (100g) or IA saline, with or without concomitant treatment with the IL-1 receptor antagonist, r-met-hu-IL1ra (Amgen; 20mg IA and 20mg fetal IM) or (IA and IM) saline (n=4-11/group). Fetal body weights and umbilical arterial pH and blood gases at delivery were not different between groups. The IL-1 antagonist itself did not affect the fetal lungs. The antagonist completely blocked IA IL-1 β-induced lung inflammation and developmental effects. Pulmonary neutrophil recruitment, 2d after IA LPS (39.6x10P

6P U+U 17.0x10P

6P cells/kg), was reduced by r-met-hu-IL1ra (9.0x10P

6P U+U 3.8x10P

6P cells/kg) but was

greater than control (no cells). Similarly, IA LPS-induced increases in IL-1 β, IL-6 and IL-8MrNA, and IL-8 protein levels were attenuated, but not completely blocked, by r-met-hu-IL1ra. Lung compliance 7d after IA LPS (lung volume at 40 cmH2O pressure; 20.6 U+U 3.4 ml/kg) was greater than control (7.9 U+U 0.6 ml/kg) and pulmonary surfactant levels were increased (control, 0.09 U+U 0.02 mol saturated phosphatidylcholine/kg; IA LPS, 2.1 U+U 0.9 mol/kg); these effects were attenuated, but not abolished (lung volume, 15.7 U+U 1.8 ml/kg; saturated phosphatidylcholine 0.3 U+U 0.03 mol/kg), by r-met-hu-IL1ra. IL-1 signalling contributes to, but is not the sole mediator of, the fetal pulmonary response to IA LPS. IL-1 may act as an amplifier of specific components of the fetal pulmonary response to IA LPS.

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O21. Dynamic imaging of lung aeration at birth using X-ray phase contrast US.B. HooperU, M.J. Wallace, K. Siu, M. Kitchen, N. Yagi, K. Uesugi, I. Williams, M. Morgan, C. Hall, S. Irvine, K. Pavlov, J. Whitley and R. Lewis Background: Before birth, the lungs are liquid-filled but this liquid must be cleared to allow the entry of air at birth. Infants that are born preterm, commonly suffer from airway liquid retention which restricts gas exchange and promotes non-uniform pulmonary ventilation, which likely increases the risk of lung injury. Little is known of the dynamics of lung aeration or the factors that regulate it because of the inability to measure the distribution of air within the lung. Aim: To study the rate and pattern of lung aeration at birth using phase contrast X-ray imaging Methods: Phase contrast X-ray imaging utilizes differences in refractive index between air and water to image low absorbing structures. After birth, the lung is predominantly comprised of air (~80% by volume) which contrasts strongly with surrounding tissue structures (mainly water), allowing them to be imaged with high contrast and spatial resolution. A Synchrotron (SPring-8, Japan) was used as the radiation source because of its brightness and high beam intensity. At 29-31d GA (term is 32d), pregnant rabbits (n=15) were anaesthetized and pups delivered by C-section and either imaged live from birth (n=10) or killed then imaged. Images of live pups were collected at 1-2 sec intervals within the first few breaths of birth. Pups imaged after death were killed after a few breaths, at 3-5mins, 10mins, 15mins, 30mins, 1hr or 2hrs after ventilation onset (n=6 per group); fetuses were also imaged. Results: Phase contrast X-ray imaging can clearly resolve both large (trachea) and small (<100m) airways of the air-filled lung; liquid-filled fetal lungs are not visible using this technique. Lung aeration at birth spreads rapidly from the large, central airways to the peripheral regions, depending on respiratory activity. The aeration pattern is not uniform as the basal regions aerate last and there is a marked effect of body position; dependent regions aerate at significantly reduced rates. Conclusions: This technique is ideal for determining the rates and patterns of lung aeration after birth. We are currently developing mathematical models to calculate air volumes and size distribution of respiratory units within specific lung regions.

O22. Effects of Sulfasalazine on Kynurenine – Potential Therapeutic Agent UP LigamU, U Manuelpillai, D Walker & EM Wallace. Introduction: The kynurenine pathway is involved in the degradation of tryptophan and is up-regulated in the human placenta by intrauterine infection, resulting in the increased secretion of neuroactive kynurenine metabolites into the fetal circulation; a possible cause of fetal cerebral injury. How infection induces activity in the pathway is unknown. Objectives: To determine if NF-κB is involved in the inflammation-induced increase of kynurenine pathway activity in the human placenta. Methods: Explants were prepared from placentas obtained after elective caesarian section at 37-40 weeks gestation (n=8), and incubated under standard conditions in the presence of 10g/ml LPS for 24 or 48 hours; duplicates of each explant were incubated either with or without 5 mM sulfasalazine added to the medium. Expression of mRNAs for key kynurenine forming enzymes, IDO and TDO and the inflammatory cytokines TNF and IL-6 were studied by RT-PCR. Kynurenine output by explants was measured in samples if the incubation medium by absorbance at 363 nm after separation from other metabolites using a HPLC technique. Results: mRNA expression of IDO, TDO, TNFα and IL-6 increased with LPS treatment; a response mitigated in the presence of sulfasalazine (p<0.01, p<0.01, p=0.03 & p=0.04 respectively). Kynurenine output in explant media (mean U+U SEM) increased with LPS but this was also prevented by sulfasalazine treatment at 24 hours (412.1 U+U 40nM/mg vs. 147.7 U+U 48.9nM/mg, p=0.01, respectively) and also 48 hours (636 U+U 39.1nM/mg vs. 135.5 U+U 29.8nM/mg, p=0.001, respectively) Conclusions: Sulfasalazine inhibits LPS induction of both the kynurenine pathway and proinflammatory cytokines in the placenta, implicating NF-κB in the LPS effect. These observations offer potential new therapeutic strategies for the management of pregnancies with in utero infection.

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O23. 5α-reductase and human term parturition USheehan Penelope MU, Moses Eric K, Brennecke Shaun P Recent data from 5α-reductase 1 knockout mice have suggested that local progesterone metabolism of progesterone by 5 α-reductase is essential for successful murine parturition. Activity of this enzyme has previously been demonstrated in human reproductive tract tissues but its role is still unclear. The aim of this study was to investigate changes in mRNA expression of both isoforms of 5 α-reductase in human reproductive tract tissues in association with parturition. Relative quantification was performed using real time RT-PCR on samples of amnion, chorion, placenta and myometrium from two groups of term gestation patients, in labour and not in labour. Results in amnion showed a significant decrease in 5 α-reductase type 1 mRNA between the ‘not in labour’ and ‘in labour’ groups. Chorion expressed very low levels of 5 α-reductase mRNA which did not change significantly with spontaneous labour. Higher levels were found in placenta and in myometrium but there were no significant changes in association with labour. There was no evidence of 5 α-reductase type 2 mRNA expression in any of the tissues examined. Significant changes in amnion raise the possibility of a feto-placental compartment role for 5 α-reduced steroids such as 5α-dihydroprogesterone. Known to be a potent steroid anaesthetic it may act on the fetus to decrease arousal. A similar action of progesterone has been demonstrated in fetal sheep. The lack of other significant differences suggest that the 5 α-reductase pathway is probably not a significant contributor to the endocrinology of human parturition.

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HYPOXIA

Free Oral Communications 4 Chairs: David Walker (Australia) and Carina Mallard (Sweden)

Monday September 26, 2005 0830 - 1030

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O24. Effect of maternal hypoxia during the preimplantation period on fetal and placental development in the mouse. Karen L Kind, Rebecca L Kelley, Jeremy G Thompson. Perturbation of the preimplantation embryonic environment can alter fetal and placental development. Oxygen is an important component of the embryonic environment. Expression of oxygen-regulated genes, including glucose transporters, is increased in mouse blastocysts cultured under 2% oxygen post-compaction, compared to embryos cultured under 7% oxygen or developed in vivo (1). Furthermore, weight of fetal mice developed from embryos cultured at 2% oxygen is reduced. Thus, culture of mouse embryos under low oxygen from morula to blastocyst perturbs embryonic gene expression and alters fetal development. This study investigated the effect of exposing mouse embryos to maternal hypoxia in vivo during the post-compaction period on fetal and placental development. CBAxC57Bl6 F1 female mice were mated with males of the same strain. Mice were placed in a clear box infused with 10% oxygen in nitrogen for 6 hours, from 0600 on day 4 of pregnancy (~77 hour post-mating). Embryos at this time were at the compact morula to blastocyst stage. Control mice were exposed to infused air. At day 18 of pregnancy (term=19-20 days) fetal and placental weights were measured. Further mice were allowed to deliver and pup weight was measured 24 hours following birth. Maternal hypoxia did not alter litter size (control: 8.4 + 0.4, n=11; hypoxia: 7.9 + 0.5, n=10). Day 18 fetal weight was reduced (-6%) by preimplantation hypoxia (control: 910.7 + 11.2 mg; hypoxia: 858.3 + 7.3 mg, p<0.05). Maternal hypoxia did not alter placental weight (control: 74.2 + 1.1; hypoxia: 73.5 + 1.0 mg) or fetal:placental weight ratio (control: 12.6 + 0.3; hypoxia: 11.8 + 0.2). No differences were detected in pup weight 24 hour after birth (control; 1.44 + 0.01 g; hypoxia: 1.46 + 0.02 g, n=7 litters/group). Six hours maternal hypoxia during the preimplantation period reduced fetal weight in late gestation, but did not alter birth weight. Further analyses are required to investigate the effect of maternal hypoxia on embryonic gene expression and metabolism. This study supports the suggestion that oxygen is an important component of the embryonic environment, that if varied may have consequences for fetal development. 1. Kind et al. (2005) Mol. Reprod. Dev. 70:37-44. O25. Abstract removed from this timeslot

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O26. Maintenance of Mean Arterial Blood Pressure in Chronically Hypoxic Fetal Sheep is not Dependent on Post Ganglionic Sympathetic Activation UJ.L. MorrisonU, I.C. McMillen Epidemiological studies demonstrate an association between intrauterine growth restriction and increased risk of cardiovascular disease in adult life. The mechanisms underlying this association are less well understood. We have shown that placental restriction (PR) in the sheep, resulting from removal of the majority of the endometrial caruncles prior to conception, results in chronically hypoxic, growth restricted fetuses. PR fetuses have higher circulating plasma noradrenaline concentrations than control fetuses. Pharmacologic studies show that the PR fetus exhibits a greater fall in mean arterial pressure (MAP) during α-adrenergic blockade. This study aimed to determine the role of post ganglionic sympathetic activation in regulating MAP in growth restricted fetal sheep. Four ewes underwent carunclectomy surgery 10 wk prior to mating. Fetal catheterisation was performed at 104-118 d gestation (term, 150 d) in 6 control and 4 PR fetuses. At 117.6±0.4 d gestation, fetuses received a bolus infusion of saline followed 1 h later by a bolus i.v. injection of 16.5±2.2 mg/kg hexamethonium bromide, a nicotinic antagonist which blocks post ganglionic sympathetic activation. MAP was monitored using Chart 4 (PowerLab, ADInstruments, Sydney, AUS) for 2 h before and 24 h after drug treatment. PR fetuses had a lower POB2 B (17.5±1.3 mmHg) than control fetuses. There was no difference in the effect of hexamethonium on MAP in PR and control fetal sheep (PR; -5.3±1.3 mmHg: Control: -4.3±1.0 mmHg). Thus these data suggest that the increased dependence of maintenance of MAP on α adrenergic activation in the PR fetus is not a result of increased post ganglionic sympathetic activation. O27. Physiological and neurodevelopmental effects of umbilical cord occlusion in fetal sheep exposed to chronic intrauterine inflammation UI NitsosU, SM Rees, BW Kramer, R Harding, JP Newnham and TJM Moss The ability of the fetus to respond appropriately to acute episodes of asphyxia may be impaired in the presence of intrauterine inflammation, resulting in adverse neurodevelopmental effects. We aimed to compare fetal effects of umbilical cord occlusion (UCO) in the presence and absence of chronic intrauterine inflammation. Pregnant ewes bearing singletons underwent surgery at 80 days of pregnancy (d; term is ~150d) to implant an osmotic pump for intra-amniotic infusion of lipopolysaccharide (LPS, from E coli 055:B5; 1.1mg/day, n=17) or saline (n=8) for 28 days. At 110d, an umbilical cord occluder was implanted, and fetal and maternal vascular and amniotic catheters inserted. At 117 and 118d, surviving fetuses underwent UCO for 2min every half hour over a 2h period (LPS+UCO, n=6; saline+UCO, n=8) and arterial pressure, blood gas, pH and lactate responses were measured. Fetal blood was then sampled daily until 125d. At 125d the fetal brain was perfusion fixed and sections of cerebral cortex were stained with H&E and immunostained with Lycopersicon Esculentum, glial fibrillary acidic protein (GFAP) and CNPase to identify activated microglia/macrophages, reactive astrocytes and myelinated fibres, respectively. Intra-amniotic LPS caused fetal death before 110d in 11 of 17 sheep, in contrast to our previous experiences of complete survival with this level of LPS exposure. Fetal pH, blood gas, lactate and mean arterial pressure responses to UCO were not different between LPS+UCO and saline+UCO groups. After UCO, from 120d, fetal lactate concentrations were higher in the LPS+UCO group than in the saline+UCO group (p=0.038). Intra-amniotic LPS caused histological chorioamnionitis and elevated amniotic fluid IL-6 (LPS+UCO; 1.20.5ng/ml; saline+UCO, 00ng/ml: p=0.017) and IL-8 concentrations (LPS+UCO; 97.28.0ng/ml; saline+UCO, 2.50.4ng/ml: p=0.017) at 125d. Fetal and maternal white blood cell counts, and fetal body and organ weights, were not different between groups. Reduced myelination was observed in white matter regions with mild-to-moderate levels of activated microglia/macrophages and hypertrophic astrocytes in the brains of 3/6 LPS+UCO fetuses. These effects were observed in 3/8 saline+UCO fetuses, which also exhibited white matter lesions. Fetal physiological responses and neuropathology as a result of intermittent UCO do not appear to be exacerbated by mild chronic intrauterine inflammation.

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O28. Regulation of Hypoxic Pulmonary Vasoconstriction in the Perinatal Period by Reactive Oxygen Species. Sewite Negash, Yuansheng Gao, J. Usha Raj. We have previously reported that nitric oxide-cGMP-mediated vasodilation in the fetal pulmonary circulation is primarily mediated through cGMP-dependent protein kinase (PKG) (J. Appl Physiol, 88:2000). In hypoxia, PKG protein expression and activity is downregulated thus promoting vasoconstriction (Am J Physiol, 285:2003), but the mechanisms by which this occurs is not known. In hypoxia, we have found an increased production of reactive oxygen (ROS) and nitrogen (RNS) species and therefore we have investigated their role in regulation of PKG. Fetal ovine pulmonary veins (PV) and vein smooth muscle cells (PVSMC) were incubated in hypoxia (pO2 30-40 torr) or normoxia (pO2 100 torr), and assayed for ROS production, PKG expression and presence of nitrotyrosine residues within PKG. Relaxation responses of PV to cGMP were also obtained. All studies were done in the presence and absence of N-acetyl cysteine (NAC), a scavenger of ROS. We find that ROS generated in hypoxia in both PV and PVSMC leads to significant reduction in total PKG protein and also an increase in nitrotyrosine content in PKG. Nitration of PKG results in decreased PKG activity. These effects in hypoxia are reversed by scavenging ROS by NAC. These effects are also reversed by re-exposure to normoxia within 30 min, indicating that PKG nitration in hypoxia is rapidly reversible by oxygenation. We conclude that hypoxic generation of ROS in pulmonary SMC leads to nitration of PKG and vasoconstriction and that this mechanism is rapidly reversible with oxygenation. Supported by NHLBI grants HL59435 and HL75187. O29. The Effect of Acute and Chronic Induced Hypoxia on BDNF Protein Expression in the Ovine Fetal Brain Near Term H.Nishigori, D.Mazzuca, V.Han, R.Gagnon, and B Richardson. Introduction: Brain derived neurotrophic factor (BDNF) plays an important role in the regulation of neuronal proliferation, differentiation, and connective plasticity during development and is dependant in part on active stimulus input to the brain. We therefore sought to determine the effect of acute hypoxia with umbilical cord occlusion (UCO) and chronic hypoxia with placental embolization (EMB) which are known to alter electrocortical activity and thereby stimulatory input within the brain, on BDNF levels in the near term ovine fetal brain. Method: Chronically instrumented fetal sheep were studied over two days near term; UCO group, n=7, complete cord occlusion for 2 min every hr over 6 hr each day; EMB group, n=7, latex microspheres injected into the common umbilical artery until arterial O2 content (CaO2) was reduced by 50%; and control group, n=6. Animals were then euthanized, the fetal brains rapidly extracted and fast frozen in liquid nitrogen, and then subsequently analyzed for BDNF using the Emax Immuno Assay system from Promega. Results: UCO caused brief severe 80-90% reductions in fetal CaO2 (p<0.001) which returned to baseline levels by 5 min post UCO while EMB caused a sustained 50% reduction in fetal CaO2 (p<0.001). In control group animals, BDNF levels measured 5.7 + 0.2 (SEM) pg/mg in the cortex, which was significantly lower and with less variance, than that measured in the subcortex at 10.6 + 1.4 pg/mg. UCO and EMB resulted in a marginal fall in BDNF levels in the cortex, to 5.0 + 0.6 pg/mg (NS) and 4.8 + 0.4 pg/mg (p=0.08), respectively, but with no change in the subcortex. Conclusion: We conclude that induced hypoxia decreases BDNF levels in the cerebral cortex of the ovine fetus near term, and more so with chronic hypoxia, which may relate to decreased stimulus input and contribute to altered neuronal connectivity over time.

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O30. Adensosine A2A receptors mediates respiratory roll-off response to hypoxia in awake lambs Brian J. Koos, Yoshikazu Kawasaki, Young-Han Kim, Fanor Bohorquez Adenosine (ADO) receptor antagonists blunt the respiratory roll-off response to hypoxia in the newborn. This study was designed to determine the ADO receptor subtype involved in the respiratory depression. Chronically catheterized lambs of 7-16 days of age breathed via facemask a gas mixture with a fraction of inspired O2 of 0.21 (normoxia) or 0.07 (hypoxia) while being infused intravascularly with ZM-241385 (ADO A2A receptor antagonist, n = 7), DPCPX (ADO A1 receptor antagonist, n = 8), or vehicle. Ventilation was measured at 20o C by a turbine transducer flow meter. In normoxia (PaO2 ~ 83 Torr), ZM-241385 or vehicle did not alter respiration, mean arterial pressure (MAP), or heart rate (HR). In vehicle studies, hypoxia (PaO2 ~ 31 Torr, 15 min) increased minute ventilation to a peak value of ~2.5 times control within the first 3 min, which was followed by a significant (P < 0.05) decline to ~50% of the maximum increment over the subsequent 7 min. ZM-241385 abolished the hypoxic ventilatory roll-off. Rectal temperature responses to infusion during normoxia and hypoxia were similar in both groups. DPCPX increased ventilation in normoxia but did not blunt the roll-off in hypoxia. It is concluded that ADO A2A (rather than A1) receptors are critically involved in the ventilatory roll-off response to hypoxia. Supported by NIHCHD Grant HD-18478

O31. A Framework for Standardized Management of Intrapartum FHR Patterns Tomoaki Ikdea, J.T. Parer, Junji Onishi, Shunichi Node, Hiroshi Sameshima and Tsuyomu Ikenoue Objectives: 1) To classify FHR patterns according to risk of fetal acidemia, and 2) to determine evolution of the patterns to those signifying increased risk of acidemia. Methods: We have identified 134 FHR patterns based on baseline rate, baseline variability, and type of deceleration. Based on the best available evidence (Parer et al, WCPM, Osaka, Japan, 2003) we have assigned a risk of newborn acidemia or low 5 min Apgar score to these patterns and coded them in five levels from I (no intervention required) to V (immediate intervention). A hierarchy of more conservative interventions has been specified and related to management of intermediate levels of risk of acidemia. In order to determine changes in patterns over time a consecutive series of 1,000 FHR tracings from low risk pregnancies was used from one Japanese institution. Tracings in the last 60 min before delivery were classified in 10 min windows. The data were depicted as incidence of each pattern and its severity at each 10-minute interval, and evolution of each pattern to either milder or more severe patterns. Results: As an example, the incidence of one pattern (variable decelerations with normal baseline rate and variability) in this series at 60 min before delivery was 27% mild, 0.3% moderate, and 1% severe. In the 10 min before delivery, the incidence was 31% mild, 11% moderate, and 4% severe. Similar trends were seen in late and prolonged decelerations. As a further example, we compared mild variable decelerations (VDs) in the period 60 min before birth to those in the 10 min before birth. Twenty-nine % remained mild, 4% reverted to normal, and 23% became late decelerations (LDs). Only 16% evolved to moderate or severe VDs, and 27% to prolonged decelerations. None evolved to VDs with decreased FHR variability. In the case of mild LDs with normal baseline variability, none remained mild, 3% reverted to normal, 23% evolved to VDs, and 23% evolved to moderate or severe LDs. None evolved directly to LDs with decreased variability. Discussion and Conclusion: Several principles are evident from these data. In the last 60 min before birth there is considerable lability in decelerative patterns. Furthermore fetuses do not decompensate without evolving to the more severe decelerative patterns. We speculate that these observations can be used to relate FHR patterns to the risk of umbilical arterial acidemia, and evolution of the patterns can be used to aid the practitioner in determining timing of intervention to avoid such acidemia. This may provide a framework for developing algorithms for standardized management of FHR patterns during labour, which can be tested for validity.

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PERINATAL BRAIN INJURY

Free Oral Communications 5 Chairs: Paul Colditz (Australia) and Brian Koos (USA)

Monday September 26, 2005 1100 - 1300

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O32. Melatonin Reduces Inflammation and Cell Death in Cerebral White Matter in the 0.65 Gestation Fetal Sheep Carina Mallard, Anna-Karin Welin, Pernilla Arvidsson, Bo Sultan, Henrik Hagberg, Pierre Gressens, Ingemar Kjellmer Background and aim: The premature infant is at increased risk of cerebral white matter injury. Melatonin is neuroprotective in adult models of cerebral ischemia and attenuates ibotenate-induced white matter cysts in neonatal mice. The aim of this study was to investigate the possible protective and anti-inflammatory effects of melatonin in the immature brain following intrauterine asphyxia. Method: Fetal sheep at 90 days of gestation were subjected to 23.5 minutes of umbilical cord occlusion. Melatonin (20mg/kg, n=9) or vehicle (NaCl + 10% ethanol, n=10) was administered intravenously starting 10 min after the start of reperfusion and continued for 6 hours. Four days after the insult, fetal brains were examined for morphological injury; ameboid microglia cells and TUNEL-positive cells were counted. Results: There were no deaths in any of the groups. In response to asphyxia, fetal mean arterial blood pressure (FMAP) dropped to a similar level in vehicle (16.32.2 mmHg) and melatonin treated fetuses (14.81.5 mmHg). In vehicle treated animals, FMAP was normal within 3h after asphyxia, while FMAP was normalized at 8h after the occlusion in melatonin treated animals. Melatonin treated animals demonstrated a reduced number of activated microglia in the periventricular white matter (Veh: 359117 cells/mm2 vs Mel: 8617 cells/mm2, p<0.01) and in sub-cortical white matter (Veh: 32865 cells/mm2 vs Mel: 7427 cells/mm2, p<0.05) and TUNEL-positive cells in white matter (Veh: 28744 cells/mm2 vs 17926 cells/mm2, p<0.05) and thalamus (Veh: 22270 cells/mm2 vs Mel: 7532 cells/mm2, p<0.05). Conclusion: This study suggests that melatonin attenuates white matter injury, possibly through anti-inflammatory mechanism.

O33. Repeated Episodes of Umbilical Cord Occlusions Increase Vascular Endothelial Growth Factor (VEGF) Expression in the Cortical Gray Matter of the Near Term Ovine Fetus H.Hamrahi, A.Becks, K.Nygard, B.Richardson, V.Han, R.Harding and A.Bocking. Umbilical cord occlusions (UCO) compromise oxygen delivery to the fetus and may result in fetal brain injury. Vascular endothelial growth factor (VEGF) is a neuroprotective agent released in response to hypoxia. Objective: To determine the effect of repeated UCO on VEGF expression in the brain of chronically catheterized near term fetal sheep. Methods: Fetal sheep underwent surgical placement of vascular catheters and an umbilical cord occluder at 122d gestation. Animals were divided into two studies, each with two experimental groups. Study 1: 7dUCO group (n=3) received repeated UCO of 2min (d1-6) and 4min duration (d7) every hour for 4hr, Nave group (n=4) received repeated UCO of 4min duration (d7) every hour for 4hr. The fetuses were sacrificed 2hr after the final UCO. Study 2 animals received the same treatments (7dUCO n=7, Nave n=7) and were sacrificed 72hr after the final UCO. A separate group (n=6) received no UCO and served as controls for both studies. VEGF expression in layers 1-3 of the parasaggital gray matter of fetal brains was studied by immunohistochemistry followed by quantification using an image analysis system (OpenLab v3.1.5). Results: In both studies, 4minUCO on d7 resulted in transient severe asphyxia; a decrease in fetal PaO2 (22.64.2mmHg to 5.62.2mmHg), and an increase in PaCO2 (47.01.1 to 88.02.5mmHg) with progressive acidemia (pHa decreased from 7.3 to 7.0). Fetal blood gases returned to baseline values by the following day in all animals. VEGF immunoreactivity appeared to be clustered in the neurons and neurites in layers 1-2 of the parasaggital gray matter. Quantification expressed as the fractional area of positive stain showed VEGF immunoreactivity was significantly higher in Study 1 7dUCO and Nave animals compared to Control animals (p< 0.005), while in Study 2 this immunoreactivity had returned to Control levels. Conclusions: Repeated episodes of mild fetal asphyxia over a six-day period followed by a severe insult lead to an increase in VEGF levels similar to that of the severe insult. This increase in VEGF expression returns to control values within 72 hours following the final UCO. Our findings support a role for VEGF in neurogenesis in response to asphyxia in the developing sheep brain.

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O34. Response of fetal sheep to intra-amniotic E.coli in the presence or absence of umbilical cord occlusions. UL PatrickU, S Hemstreet, B Matushewski, J Homan, B Richardson and G Smith. It is unknown at present to what extent fetal hypoxia complements the processes by which chorioamnionitis increases risk of fetal brain injury, and whether damage is increased in the presence of both insults. We utilized the chronically instrumented fetal sheep model in order to establish three study groups: (1) repeated umbilical cord occlusions (n=4), (2) exposure to 500CFU intraamniotic E.coli (n=6), and (3) exposure to 500CFU intraamniotic E.coli followed by repeated umbilical cord occlusions (n=5). The fetus was instrumented such that samples of fetal arterial and sagittal vein blood, amniotic fluid and maternal vein and artery could be collected for analysis. Cord occlusions were performed using an inflatable cuff. In addition, the fetal brain was instrumented with a Doppler crystal to measure brain blood flow velocity in each experiment. Initial analysis indicates that amniotic fluid proinflammatory cytokines IL-1 and IL-6 levels are increased after both exposure to bacterial insult and after umbilical cord occlusions. In experiments where the fetus was exposed to cord occlusions in the absence of bacterial insult, fetal sagittal Doppler readings showed a sharp increase in the period during and after the occlusions, and full recovery to baseline levels was not achieved after one hour, such that Doppler flow gradually increased in magnitude after each occlusion throughout the day. In the experiments where E.coli was injected 24 hours before occlusions were started, Doppler flow again increased during and after the occlusion, but the levels tended to decrease to baseline levels after one hour. In experiments where the fetus was exposed to intraamniotic E.coli but was not subject to umbilical cord occlusions, the fetal Doppler readings decreased over time, with a decrease of approximately 10% of baseline values after one day, and decreases of 25-30% from baseline after 2-4 days. Currently, data is being analyzed in order to determine fetal and maternal arterial and amniotic fluid cytokine concentrations, and the degree to which chorioamnionitis may increase the levels of fetal brain injury in the presence of hypoxic episodes. O35. Time Course of Endotoxin Impairment of Cerebral Endothelial Function in Newborn Lambs UFeng, YSU, Zivkovic, S, McClure,L, Yu, VYH, Walker, AM Introduction: Infection is an established risk-factor for perinatal brain injury, including cerebral palsy, but the exact mechanisms remain unclear. Recent interest has focused on endothelial injury and the loss of important endothelium-derived vasodilators such as nitric oxide (NO) as a critical mechanism underpinning the damaging effects of infection. In this study we used bradykinin (BK), an endothelium-dependant NO precursor to assess the progress of endothelial impairment in the cerebral circulation followed a lipopolysaccharide (LPS) induced inflammatory insult. Aim: To assess the pattern of cerebral vasodilatation and its response to BK, before and after LPS administration. Methods: Lambs (n = 10) aged 1-4 weeks were instrumented for recording cerebral blood flow (CBF, flow probe on the superior saggital sinus), carotid arterial pressure (Pca), intra-cranial pressure (Pic). Cerebral vascular resistance (CVR) was calculated as (Pca + Pic)/CBF. Studies began after 48 hour recovery with an endothelial function test using BK (intra-carotid injections of 0.001 to 0.5 µg/kg, 0.5ml/dose). This was followed by LPS (2µg/kg IV) infusions on 3 consecutive days. A repeat BK study was performed 24 hr post LPS. Results: Acute responses to LPS (0-4hr): 3hr post-LPS, CBF was reduced to a minimum of -31 U+U 4%, (mean U+ U SEM) due to an increment of CVR (117 U+U 32%, P<0.05, ANOVA). Delayed responses to LPS (5-10hr): CBF remained low despite a return of CVR to baseline because blood pressure declined by -21U+U6% (P<0.05). Chronic responses to LPS (24hr): CBF had returned to baseline levels, but the CVR reductions in response to BK that had existed pre LPS (-18U+U5%) were abolished (P<0.05). Summary: LPS begins to disrupt the cerebral circulation in newborn lambs within an hour of its administration. An acute cerebral vasoconstriction which peaks at 3hr is followed by a period of hypotension extending through to 10hr, as a result, CBF is depressed throughout the period 1-10hr post LPS. Conclusion: Endotoxin impairs cerebral perfusion in newborn lambs via two mechanisms: an acute vasoconstriction (hours) followed by a prolonged period of hypotension. Endothelial dysfunction which persists 24hr after LPS suggests that endothelial injury is a critical component of the circulatory disruption that follows LPS.

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O36. Acute Episodic Fetal Alcohol Exposure in Late Gestation Can Increase Apoptosis and Astrogliosis and Induce White Matter Injury in the Immature Brain Penelope A Dalitz, Richard Harding, Jhodie R Duncan, Megan L Cock, Philip N Henschke, Sandra M Rees. BACKGROUND: New evidence suggests acute patterns of alcohol exposure in late gestation can be particularly harmful to the developing brain. In the fetus, characteristics of neuropathology following acute alcohol exposure are largely unknown. OBJECTIVE: Using an ovine model, we have examined the effects of repeated, acute fetal alcohol exposure on the developing cerebral and cerebellar white matter (WM) and hippocampus. METHODS: EtOH (1g/kg of maternal weight) was administered i.v. to 8 twin-bearing ewes for 1 hour on 3 consecutive days from 1161 days gestation (DGA; term ~147); controls (n=5) received saline. At autopsy (1201 DGA), brains were processed for structural analysis. Fetal brains were assessed for alterations in gross morphology (H&E), astrocytes (GFAP), microglia/macrophages (lectin), axons (Bielschowsky and amyloid precursor protein (APP)), oligodendrocytes (CNPase) and apoptosis (TUNEL). The incidence of astrogliosis, and the aerial density of apoptotic cells and oligodendrocytes were examined by quantifying GFAP-, TUNEL- and CNPase- positive cells (cells/mm2) respectively within the peri-ventricular-, deep- and sub-cortical- WM in the frontal, parietal, temporal and occipital lobes. TUNEL positive cells were also quantified in the entire hippocampus and cerebellar deep WM (cells/mm2). RESULTS: Maternal and fetal BAC peaked at 0.11 + 0.01 g/dL 1h after EtOH infusions on each experimental day. Fetuses did not become hypoxemic or acidemic. In 4/8 EtOH fetuses, areas of subcortical WM injury were observed in sections from all lobes: damage was substantial in 2 fetuses and modest in 2 fetuses. Injury was characterised by axonal disruption and infiltrating microglia/macrophages. TUNEL- and GFAP-positive cells were significantly increased throughout the cerebral WM of EtOH fetuses compared to controls (p=0.005 and p=0.05 respectively). Oligodendrocyte numbers did not differ between treatment groups. In 2/8 EtOH fetuses, cerebellar WM gliosis was observed. TUNEL positive cells tended to be increased in the deep cerebellar WM of EtOH fetuses in comparison to controls (p=0.063). TUNEL-positive cell numbers in the hippocampus did not differ between treatment groups. No brain injury was observed in controls. CONCLUSION: We conclude that episodic exposures of the immature fetus to modest levels of EtOH can result in overt brain injury and an increase in WM apoptosis and astrogliosis. O37. The influence of birthweight for gestational age on neurodevelopmental outcome at 1 year of age in premature infants weighing <1000gms Lloyd J, Riley K, Wyatt J, Peebles D Background: Premature infants are at increased risk of developing cerebral palsy and other neurological problems, such as cognitive and behavioural problems. Materno-fetal infection/inflammation has been implicated, both as a cause of preterm labour, and perinatal brain injury. By contrast, the precise role of fetal growth restriction in impairing brain development remains unclear. Aims: To investigate the association between intrauterine growth restriction and neurodevelopmental outcome in a cohort of premature infants at a single tertiary centre. Methods: 220 premature infants with a birth weight 1000g cared for at University College Hospital London from 1999 to 2002, were prospectively followed up at a corrected age of 1 year using a validated neurodevelopmental assessment tool, the Griffith’s Mental Development (GQ) Scales. Birth weight was classified as <2nd, 2nd to 9th and 10th centile and z-scores calculated (the number of standard deviations from the mean birth weight for gestational age). Histological evidence of placental infarction or chorioamnionitis and ultrasound evidence of brain lesions (cystic periventricular leucomalacia [PVL] or ventricular dilatation >2SD) were also recorded. A multivariate analysis of risk factors was performed, adjusting for z-score and gestational age. Results: 220/236 survivors had a GQ recorded (93.2% follow up). On univariate analysis, babies with a birth weight <2nd centile had a lower mean GQ score than those with a birth weight 10th centile (82.78 +18.85 versus 95.31+16.54, p<0.001). The presence of cystic PVL or ventricular dilatation was associated with a lower mean GQ score of 72.50+23.13 versus 97.20+14.98 (p<0.001) and 86.54+20.44 versus 98.09+14.84 (p<0.001) respectively. There was no association between histological chorioamnionitis and GQ score. These findings were confirmed by multivariate analysis with a significant association between lower Z-score, gestational age, male gender, the presence of cystic PVL or ventricular dilatation and lower GQ score. In the multivariate model an increase in z-score of 1.0 was associated with an increase in GQ score of 5.6+1.48 points; similarly, an increase of 1 week in gestational age increased GQ by 2.1+0.71 points. Conclusions: Growth restriction, extreme prematurity and major cranial ultrasound abnormalities have a significant negative association with neurodevelopmental outcome in premature infants at 1 year of corrected age.

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O38. Antenatal Glucocorticoids: Effects on Cerebral Substrate Metabolism and Blood Flow in the Preterm Ovine Fetus J McCallum, N Smith, M Schwab, P Nathanielsz, B Richardson OBJECTIVE: While glucocorticoids (GCs) have proven to be of clinical benefit in pregnant women at risk for preterm delivery with the enhancement of fetal lung maturation, there is increasing evidence of adverse effects on brain development, which may relate to altered growth processes and metabolic activity. We therefore sought to determine the effect of maternal GC administration on cerebral substrate metabolism and blood flow in the preterm ovine fetus in amounts comparable to that used clinically in human pregnancy. METHODS: Chronically instrumented pregnant sheep at 0.85 gestation received two intramuscular injections of betamethasone at 170 g/kg maternal weight 24 hours apart (experimental group, n=13) or saline (control group, n=10). Fetal cerebral measures of substrate arteriovenous difference (O2, glucose, leucine) and blood flow (fluorescent microspheres) were obtained pre 1st GC injection or baseline, 24 hours post 1st GC injection or late GC 1, and 4 hours post 2nd GC injection or early GC 2. Results are presented as group means + SE with statistical significance determined using repeated measures ANOVA. RESULTS: Fetal blood gases, pH, and blood metabolites were little changed in either animal group over the course of study except glucose values in GC animals, which increased 1.4 and 1.9 fold, late GC 1 and early GC 2, respectively (both p<0.01). Cerebral blood flow while little changed in the control animals, or in GC animals when measured late GC 1, was decreased in GC animals when measured in early GC 2 by 28% (p<0.05). As such, GC animals showed a decrease in cerebral O2 delivery of ~ 21% early GC 2 (p<0.05) and conversely an increase in cerebral glucose delivery of 1.4 and 1.3 fold, late GC 1 and early GC 2, respectively (both p<0.05). Fractional extraction values for these substrates were not significantly changed resulting in corresponding decreases in O2 uptake and increases in glucose uptake, such that the estimated glucose/oxygen quotient as an index of glucose oxidative metabolism measured ~1.45 early GC 2, which was considerably greater than control group/baseline values at 1.0 to 1.1. Fractional extraction values for leucine were also higher in the GC animals by ~1.6 fold when measured both late GC 1 and early GC 2. CONCLUSIONS: Fetal cerebral metabolism in the preterm ovine fetus is altered by GC administration comparable to that used in human pregnancy, including an acute decrease in cerebral blood flow with vasoconstriction, an acute increase in anaerobic glucose metabolism, and an overall increase in leucine flux into the brain. These metabolic alterations may in turn reflect and/or give rise to altered biosynthetic processes, and limit the brain’s adaptive response for superimposed hypoxic injury.

O39. Birth weight and neurodevelopmental outcome of children at two years of age after planned vaginal delivery for breech presentation at term Molkenboer JFM, Roumen FJME, Smits LJM, Nijhuis JG. Objective: The purpose of this prospective cohort study was to determine whether planned vaginal delivery for the term singleton baby in breech position increases the risk of abnormal neurodevelopment at two years of age, and to assess whether the effect is modified by birth weight. Study design: At two years of age, all non-randomized children born in breech position during our participation in the Term Breech Trial (TBT), were screened for abnormal neurodevelopment with the Ages and Stages Questionnaire (ASQ). Results: An ASQ at two years of age was obtained in 183 of 203 children (90.1%). Twenty-eight percent of these children showed one or more abnormal ASQ domains. There were no differences in the risk of having abnormal ASQ domains between planned vaginal delivery and planned cesarean section (p=0.99). There was, however, evidence of interaction between mode of delivery and birth weight, with significantly higher risk in neurodevelopmental delay in children over 3500 grams with planned vaginal birth (adjusted OR for interaction term, 3.37; 95% CI, 1.14-9.95). Conclusion: Based on the ASQ results at two years of age, planned vaginal delivery is associated with an increased risk of neurodevelopmental delay at two years of age in term breech children with a birth weight above 3500 grams.

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NUTRITION

Free Oral Communications 6 Chairs: Karen Gibson (Australia) and Alison Forhead (United Kingdom)

Tuesday September 27, 2005 0830 - 1030

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O40. Maternal Nutrient Restriction During Pregnancy Changes the Postnatal Pattern of Prostaglandin Regulation in the Rat Kidney K.A. Brennan, S.W. Reynolds, I. Christiaens, B. McCook, G. Kan, M.E. Symonds and D.M. Olson Introduction. Epidemiological and animal studies have shown that maternal nutrient restriction (NR) during pregnancy can adversely affect both the development and later functioning of the offspring’s kidneys. Prostaglandins (PGs) are known to play an important role in these aspects but so far little research has been done into the effects of maternal NR on prostaglandin synthesis or action. Therefore, the present study examined the influence of NR on the expression of the key enzymes in prostaglandin synthesis (Cyclooxygenase-1 and -2) and degradation (Prostaglandin Dehydrogenase, PGDH) as well as the expression of the PG receptors EP1-4 and FP. This study also investigated whether NR leads to a persistent change in the tissue content of PGE2 itself. Since glucocorticoids are known to be affected by NR and can regulate PG expression this study also examined plasma corticosterone levels and glucocorticoid receptor (GCR) expression. Methods. At conception rats were randomly allocated to either a control group that was fed ad libitum or a NR group that was fed half the amount eaten by the control animals. After birth all animals were fed ad libitum. Pups from each litter were euthanased at postnatal days 0, 7, 14, 21, 28 and at 12 weeks old. Their kidneys were sampled and used to measure the mRNA and protein abundance of the enzymes and receptors mentioned above. Commercially available assay kits were used to measure PGE2 content in the adult kidneys and plasma corticosterone levels at postnatal days 0, 14, 28 and 12 weeks. Results. It was found that NR affects the mRNA expression of all genes examined either by accelerating or delaying developmental changes. The only exceptions to this being cyclooxygenase-1 and EP1. The mRNA differences were not always matched by protein expression suggesting regulation at the post-transcriptional level. Further, maternal NR decreased the kidney levels of PGE2 in the adult offspring and plasma corticosterone levels were raised in NR animals at postnatal days 14, 21, 28 and adult animals. Conclusion. This study is the first to report the developmental expression pattern of the EP receptors, FP receptor and PGDH in the rat kidney. The NR-induced changes in the PG system may be linked to changes in organ development and function that have been previously reported in NR rats and may be mediated by glucocorticoids.

O41. Maternal protein restriction: effects in the heart of offspring later in life Kyungjoon Lim, Monika A Zimanyi, M Jane Black Background: To date, the long-term effects of intrauterine growth restriction on the structural architecture of the heart, including the vasculature, has not been well described. Aims: The aim of this study was to examine the effect of maternal protein restriction in rats on the levels of cardiac fibrosis, myocardial capillarisation and media to lumen ratio of intramyocardial arteries in the heart of the offspring in adulthood. Methods: Female WKY rats were fed either a normal protein diet (NPD; 20% casein) or a low protein diet (LPD; 8.7% casein) during pregnancy and lactation. Female offspring (n=7/group) were weaned at 4 weeks of age and grown to adulthood. At 24 weeks of age, the offspring were perfusion-fixed, the hearts excised, weighed and sliced into 1mm slices. Every second heart slice was used in a comprehensive evaluation of the levels of cardiac fibrosis using image analysis. The media:lumen ratio of the intramyocardial arterioles was also assessed. The remaining heart slices were used to stereologically investigate capillarisation of the heart. Results: Body weights at both 2 weeks and 24 weeks of age were significantly reduced (31% and 8%, respectively) in the LPD offspring however, heart size was not different at 24 weeks. Importantly, by adulthood, there was a significant 15% increase in fibrosis in the interstitium of the left ventricle in the LPD offspring. There were no differences in levels of perivascular fibrosis or in the media:lumen ratio of intramyocardial arteries in the LPD and NPD offspring. Similarly, there was no significant difference in surface area and length density of myocardial capillaries or in total surface area or total length of myocardial capillaries in the LPD and NPD hearts. In Conclusion: Since cardiac fibrosis is associated with impaired cardiac contractility and arrhythmia, our results suggest that induction of interstitial fibrosis may be a likely contributor to the increased cardiac disease in adult subjects that were growth-restricted at birth.

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O42. Periconceptional Nutrition and the Relationship Between Maternal Body Weight Changes in the Periconceptional Period and Fetal Kidney Growth in the Sheep SM MacLaughlin, SK Walker, DO Kleeman and IC McMillen Recent studies in the sheep have shown that maternal undernutrition during the periconceptional period can alter the subsequent development of the metabolic, endocrine and cardiovascular systems and that these effects may, in part, depend on embryo number. We have tested the hypotheses that there are relationships between fetal kidney growth at ~ d 55 pregnancy and maternal weight change during the periconceptional period and early pregnancy, and that periconceptional undernutrition has a differential effect on these relationships in singleton and twin pregnancies. We have investigated the effect of periconceptional undernutrition in the ewe (control n = 24, restricted at 70 % of control feed allowance, PCUN n = 21) from 45 days prior to mating until 7 days after mating on fetal kidney weight at d~ d 55 pregnancy. In control singleton fetuses there were no relationships between relative fetal kidney weight at ~ d 55 pregnancy and maternal weight change during the periconceptional period or between d 10 of pregnancy and PM, however, there was a significant inverse relationship between relative fetal kidney weight at d ~ 55 pregnancy and maternal weight change before PM. In control twin fetuses there was an emergence of an inverse relationship between relative kidney weight at ~ d 55 pregnancy and maternal weight change during the periconceptional period and before PM, however, no relationship with maternal weight change between d 10 of pregnancy and PM. Interestingly, in fetuses exposed to PCUN there were inverse relationships between relative fetal kidney weight at ~ d 55 pregnancy and maternal weight change during the periconceptional period, and positive relationships between relative kidney weight at ~ d 55 pregnancy and maternal weight change between d 10 of pregnancy and PM but no relationships with maternal weight change before PM in both singleton and twin fetuses. Fetal kidney development at ~ d 55 pregnancy is affected by maternal weight change during the periconceptional period and early pregnancy, and this renal development is differentially affected by fetal number and PCUN. These changes highlight the importance of the periconceptional environment in setting the fetal kidney growth trajectory.

O43. Periconceptional Undernutrition in the Ewe Down-Regulates Maternal, But Not Fetal, Adrenal ACTH Receptor, StAR and P450c17 mRNA in Early Gestation Frank H Bloomfield, Kristin L. Connor, Anne L. Jaquiery, Mark H. Oliver, John R.G. Challis and Jane E. Harding Maternal undernutrition, either around the time of conception or throughout pregnancy, has been demonstrated to result in altered fetal development. The mechanism has been hypothesised to be exposure of the fetus to excess glucocorticoids of maternal origin, with the nutritional insult acting as a stressful insult to the mother. However, there are few data confirming this, particularly in animals with a longer gestation. Indeed, we have previously reported that undernutrition leading to a 15% weight loss in the singleton-bearing ewe from 60 days prior to until 30 days after mating results in a profound suppression of maternal circulating ACTH and cortisol concentrations (1). The aim of this study was to determine the effects of this nutritional insult on the mRNA levels of the ACTH receptor, steroidogenic acute regulatory protein (StAR) and P450c17 in maternal and fetal adrenal glands 20 and 100 days after relief of undernutrition (i.e. at 50 (N = 13 ewes and 18 fetuses, UN = 11 ewes and 15 fetuses) and 131 d (N = 7, UN = 10) gestation)). Ewes were randomly assigned to undernutrition (UN) or ad libitum (N) groups and killed at 50 or 131 d gestation. Maternal and fetal tissues were rapidly dissected, frozen and stored at –80oC until analysis. Gene expression was analysed by real-time RT-PCR. Maternal ACTH-R, StAR and P450c17 were reduced by 17 to 45% at 50 d gestation in UN animals (P<0.05). Fetal levels of mRNA for these genes were not different between groups at 50 d gestation. However, at 131 d gestation, fetal mRNA and protein levels of P450c17 were increased in the UN group, consistent with the advanced maturation of the fetal HPA axis that we have reported previously (1). We conclude that, as there is profound down-regulation of maternal adrenal steroidogenesis that persists for at least 20 days after undernutrition has ceased with no effect on the fetal adrenal gland until late gestation, it is most unlikely that exposure of the fetus to excess maternal glucocorticoids is the mechanism by which maternal undernutrition results in altered fetal development. 1. Bloomfield FH et al. Endocrinology 2004;145(9):4278-4285.

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O44. Strain Differences in the Impact of Dietary Restriction on Fetal Growth and Pregnancy in Mice UBrian KnightU, Craig Pennell, Stephen Lye. Objective: The association between sub-optimal intrauterine environment and development of adult disease (DoHAD) is variable suggesting that genotype may contribute to eventual outcome. Chromosome substitution stains of mice allow the genetic dissection of complex traits; however, to date, they have not been utilized to evaluate the genetic contribution to DoHAD. The objective of this study was to characterize maternal and fetal responses to dietary restriction (DR) during pregnancy in two genetically different strains of mice. Methods: Fourteen A/J and 14 C57BL/6J (B6) pregnant mice underwent 30% Dr from 6.5 days (d) until 18.5d of gestation: a further 14 mice in each strain served as controls on an ad libitum diet. Half of the animals were dissected on 18.5d to assess fetal and organ weights and maternal endocrine status. The remaining mice were allowed to deliver spontaneously. Growth, blood pressure, and glucose tolerance was assessed at 3 months of age. Results: Dr resulted in reduced fetal body weight (p=0.01) in both strains. Fetal kidney weight was reduced in both strains (p=0.001); however, the magnitude was three times greater in B6vsAJ (p=0.039). Dr-B6 when compared to Dr-AJ had: significantly overgrown placentas, increased fetal brain-liver ratio and decreased fetal-placental weight ratio (all p<0.05). Maternal cortisol at day 18.5 was increased 2-fold in the Dr-B6 (p=0.02) whilst no change was seen in Dr-AJ mice. Dr resulted in a reduction in placental expression of 11B-HSD2 in both strains (p=0.02); however an increase in 11β-HSD1 protein expression was only seen in Dr-B6 mice. Dr was associated with pre-term delivery in both strains with Dr-B6 delivering earlier than Dr-AJ (p=0.012). At 3 months of age the Dr-AJ mice were still smaller than controls whereas the Dr-B6 mice had catch up growth to controls. Systolic and diastolic BP was increased in Dr-B6 at 3 months whereas it was decreased in Dr-AJ compared to strain specific control (P<0.05). Conclusion: The observed strain variations in response to dietary restriction may offer a unique opportunity to investigate gene:environment interactions associated with DOHAD. O45. Reduced Physical Activity and ad libitum Nutrition in Early Postnatal Life Leads to Obesity, Glucose Intolerance and Insulin Resistance. UHelen BudgeU, Terence Stephenson, Michael E Symonds, David Gardner. The prevalence of obesity in early childhood is accelerating and tracks to adverse metabolic and cardiovascular sequelae in adulthood. The contributions of excess food intake and reduced physical activity in early life and whether similar changes may be present in sheep are unknown. Twelve lambs were weaned at 3 months, randomly assigned to intervention group and either housed with restricted space for physical activity and provided with unlimited access to food or provided with unrestricted space for physical activity at pasture. At one year postnatal age, blood sampling was performed before and during standard intravenous glucose and insulin tolerance tests (Bonora et al. 1989; Hirst et al. 1993) conducted on separate occasions and after hay and concentrate were withheld for 24 hr. Plasma concentrations of glucose were measured enzymatically. Reduced environmental space not only reduced physical activity to 25% of controls but, when combined with excess food provision (AL-R) from weaning at 3 months postnatal age, produced sedentary hyperphagic behaviour, marked adiposity (subcutaneous fat depth; AL-R, 153; C-C, 2-3 1 cm (p<0.05)) and early obesity (body weight; AL-R, 62 U+U 2 kg; C-C, 42 U+U 3 kg: abdominal circumference; AL-R, 79 U+U 2 cms; C-C, 48 U+U 3 cm (p<0.05)) not seen in those with normal food intake and physical activity. Fasting plasma glucose concentrations (C-C, 3.5 U+U 0.2; AL-R, 4.3 U+U 0.3 mmol/L (p<0.05)) were higher in AL-R sheep at one year postnatal age than controls. Standard intravenous glucose infusion significantly increased plasma glucose concentration in both groups with a higher peak achieved in AL-R juveniles (C-C, 7.8 U+U 0.6; AL-R, 10.8 U+U 0.4 mmol/L (p<0.01)) whereas insulin infusion resulted in a slower decline in glucose towards baseline in AL-R sheep (C-C, 2.8 U+U 0.2; AL-R, 1.7 U+U 0.2 % decline/min (p<0.05)). Therefore, early postnatal exposure to ad libitum nutrition and reduced physical activity results in obesity, glucose and insulin intolerance in an ovine model. Bonora et al. 1989 J Clin Endocrinol Metab 68: 374-378, Hirst et al. 1993 Diabet Med 10: 839-842

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O46. Cross foster studies identify lactation and pup milk intake as critical to postnatal growth following placental restriction during pregnancy O’Dowd, R., Mibus, A.L. and Wlodek, M.E. Placental restriction is characteristic of human intrauterine growth restriction and these small for gestational age babies often undergo accelerated growth postnatally. This postnatal accelerated growth and hence the lactation environment are implicated as critical in the programming of many adult diseases. We have shown that placental restriction in the rat reduces birth weight, impairs maternal mammary gland function and consequently lactation, which further restricts postnatal growth. Our aim was to determine the relative importance of the in utero placental environment and the postnatal lactation environment on milk intake for programming the slow postnatal growth of placentally restricted offspring. Bilateral uterine vessel ligation (Restriction, R) or Sham (Control, C) surgery was performed on day 18 of gestation in Wistar Kyoto rats. Pups were cross-fostered on day one following birth. Pup body weight and dimensions were measured throughout lactation. Pup milk intake was also measured using the weigh-suckle-weigh protocol. Data were analysed by one-way ANOVA (n=8 mothers per group). Restricted pups were born smaller than Control pups (p<0.05). Male R and C pups, when cross fostered onto placentally restricted mothers, had a lower milk intake on day 10 than when cross-fostered onto C mothers (p<0.05). This demonstrates that mothers that have placental restriction during their pregnancy produce less milk regardless of the size of the pup suckling. R-on-R pups were smaller than all other groups throughout lactation (p<0.05). Despite the reduced milk intake of C-on-R, there was no detectable reduction in pup weight during lactation and it remains unknown whether this impacts on later growth or disease development. Male pups born small and cross-fostered onto a mother with normal lactation (R-on-C) increased their body weight in the first week after birth in response to the increase in milk delivery (p<0.05). Female pups showed a similar pattern with R-on-C increasing body weight above R-on-R by day 14 (p<0.05). Correcting a poor postnatal lactation environment by cross-fostering pups born small onto a mother with normal lactation, enables increased delivery of milk and nutrition with profound consequences for enhanced postnatal growth. This study has identified the lactation environment as a critical programming period for growth and disease development.

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GLUCOCORTICOIDS

Free Oral Communications 7 Chairs: John Newnham (Australia) and Karen Moritz (Australia)

Tuesday September 27, 2005 1100 - 1300

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O47. Developmental ontogeny of the glucocorticoid receptor and mitochondrial proteins in the ovine liver from mid-gestation through to early adulthood M. Hyatt, D. Yakubu, A. Mostyn, D. Walker, T. Stephenson and M.E. Symonds In order to thrive after birth the newborn must be able to control its own growth and metabolism independent of maternal systems. Fetal liver maturation is initiated by cortisol and is dependent upon an intact adrenal gland although the normal developmental ontogeny of the glucocorticoid receptor (GR) from fetal to postnatal life has yet to be determined. Furthermore, the involvement of energy regulating mitochondrial proteins such as voltage-dependent anion channel (VDAC) and cytochrome c to the commencement of liver function is unknown. Therefore, this study determined the normal developmental ontogeny of GR, VDAC and cytochrome c in liver tissue from mid-gestation up to early adulthood. Livers were sampled from fetuses at 80 and 140 days gestation (term = 147 days), and 1, 30, 180 and 1098 (~3 years) days after birth (n = 4-8 per sampling age). Representative samples of liver was excised from the right hepatic lobe and stored at -80oC until analyzed. Total RNA was extracted and the relative abundance of GR to 18S rRNA was measured by RT-PCR. Mitochondrial fractions were prepared and VDAC and cytochrome c abundance determined by immunoblotting. Results are given as arbitrary units and are expressed as a percentage of a reference sample. GR mRNA, VDAC and cytochrome c were expressed in liver tissue at all sampling ages, the abundance of which was developmentally regulated. Prenatally, GR and cytochrome c steadily increased throughout gestation whilst VDAC on the other hand remained low. The abundance of all three proteins was maximal one day postpartum and steadily decreased thereafter reaching basal levels by 3 years of age. In conclusion, the abundance of GR and cytochrome c steadily increases during late gestation coincident with the prepartum cortisol surge responsible for liver cell maturation and onset of adult liver function prior to birth. Up-regulation of GR and initiation of gluconeogenesis during late gestation precedes the ontogenic peak in VDAC, the expression of which may play a role in glucose homeostasis during the neonatal period through the transport of ATP and glucose-6-phosphate. O48. Ontogeny of the spiny mouse placenta and the effects of dexamethasone treatment on its development Hayley Dickinson, David W Walker, Karen Moritz, Claire Roberts In several species exposure of the pregnant mother to synthetic glucocorticoids alters placental gene expression and function, and programs hypertension in the adult offspring. We have established that the spiny mouse placenta expresses abundant IGF-II which plays an important role in placental morphogenesis in most mammalian species. IGF-II is particularly sensitive to the inhibitory effects of glucocorticoids. The spiny mouse (Acomys cahirinus) is a new animal model that is especially suited to perinatal research as it has a relatively long gestation (40 days) and is developmentally more similar to humans than other rodents. We aim to describe the ontogeny of the spiny mouse placenta throughout gestation and examine the effects of maternal dexamethasone treatment on placental development. Placentas were collected from days 14-39 of gestation. Fetal and placental weights were recorded and each placenta bisected, with one half frozen and the other fixed and processed into paraffin. Mid-sagittal placental sections were stained with Masson’s trichrome and the proportions of labyrinth (exchange region) and junctional zone (germinative region) quantified using point counting. Dexamethasone (150g/kg for 60hrs from days 20-23 of gestation via mini osmotic pump) and saline treated females were killed at 23 and 37 days of gestation and placentas processed. The spiny mouse placenta appears to develop in a similar fashion to that of the laboratory mouse, although the time scale is significantly lengthened. Placental growth is linear from day 14 of gestation and is predominantly junctional zone until mid gestation when the labyrinth differentiates and its growth escalates such that it occupies approximately 55% of the placenta thereafter. Fetal weight increases rapidly after differentiation of the labyrinth, increasing from 330mg at day 22 to 5.5g at day 39. Fetal and placental weights and the labyrinth to junctional zone ratio at day 23 are not altered by dex treatment (Table 1). We have established that the timing of dex treatment is prior to the differentiation of the labyrinth and our ongoing research will determine its impact on placental IGF-II expression and structural correlates of placental function later in gestation when fetal demand is high. Table 1. Fetal & placental (23d GA) data (mean + SEM): Fetal Wt (mg) Placental Wt (mg) Fetal:Placental Wt Ratio Lab: JZ Dex (n=9, 3 litters) 621 + 24 219 + 9 2.86 + 0.12 0.81 + 0.07 Sal (n=8, 3 litters) 676 + 22 214 + 4 3.17 + 0.11 0.75 + 0.04

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O49. Effects on lung structure of Intra-amniotic corticosteroids for preterm lung maturation in sheep. G.R. Polglase, I. Nitsos, A.H. Jobe, J.P. Newnham & T.J.M. Moss Objective: Intra-amniotic corticosteroids can improve lung function in preterm lambs without significantly altering pulmonary surfactant levels1, suggesting that changes in lung structure, which have potential long-term adverse consequences, underlie the functional effects. Methods: Pregnant ewes received intra-amniotic betamethasone (0.5 mg/kg or 2 mg/kg fetal weight), intra-amniotic budesonide (0.5 mg/kg or 2 mg/kg fetal weight), or maternal intra-muscular betamethasone (0.5mg/kg maternal weight); controls received equivalent saline injections. Lambs were delivered 2 days later, at 124 days’ gestation, for measurement of pulmonary maturational changes (presented previously) and lung morphometric analysis. Results: All corticosteroid treatments resulted in a 10-20% increase in the volume fraction of alveolar ducts, an increase in the fraction of airspace within the lungs (with the exception of the low dose budesonide group), and a significant reduction in alveolar wall volume fraction. Alveolar numerical density was reduced by approximately 20-40% by intra-amniotic corticosteroids, average alveolar volume tended to increase and total alveolar number tended to decrease. These statistically significant fractional changes in alveolar airspace and alveolar wall volumes translated into a tendency for increased airspace volume and reduced alveolar wall thickness. Conclusions: Structural alterations caused by intra-amniotic corticosteroid administration, including increased airspace and reduced alveolar wall volume, explain the improved preterm lung function observed by us previously1. These structural effects, while beneficial for function of the preterm lung, may reflect impaired alveolarisation, which is a characteristic of infants dying of bronchopulmonary dysplasia. Intra-amniotic corticosteroids lead to improved preterm lung function by causing structural changes in the developing lung. 1. Moss et al. Am J Obstet Gynecol. 2003 Nov;189(5):1389-95.

O50. Effects of late gestational betamethasone administration on fetal and postnatal weights and insulin-like growth factor (IGF) concentrations Deborah M Sloboda, Shaofu Li, John RG Challis, Timothy JM Moss, John P Newnham Prenatal exposure to synthetic glucocorticoids reduces fetal weight in sheep. To help understand this relationship we evaluated the effects of maternal betamethasone administration on circulating levels of IGF peptides in the fetus and in newborn animals. Pregnant ewes carrying male fetuses were injected with saline or 1 (104 days of gestation; dG), 2 (104, 111 dG) or 3 (104, 111, 118 dG) doses of betamethasone (0.5mg/kg). Maternal and fetal plasma was collected prior to (75, 84, 101), during (109, 116), and after betamethasone at 121-122, 132-133 and 146 dG and in offspring at 6 and 12 weeks of age. Plasma IGF1 and 2 levels were measured using RIA. Fetal tissues were also collected at these times. Fetal liver weight was reduced significantly by 109 dG and remained lower than controls until 133 dG but not thereafter. Fetal birth weight was reduced after betamethasone treatment. Lamb weights at 6 and 12 weeks postnatal age were not significantly different in treated animals. In controls, fetal IGF1 levels increased with advancing gestation and then decreased postnatally (P<0.05). After betamethasone, fetal IGF1 levels were reduced significantly at 133 days but were higher in offspring of treated animals at 12 weeks postnatal age (P<0.05). In controls, fetal IGF1 levels correlated positively with fetal weight at 122 and 133 dG and in newborn lambs at 6 weeks (P<0.05). This relationship was lost after maternal betamethasone treatment. Maternal IGF1 levels were higher in betamethasone treated animals at day 109 dG compared to controls (P<0.05). Fetal IGF2 levels were significantly decreased in fetuses at 133 dG (P<0.05) after betamethasone, but maternal IGF2 levels were not different. Neither fetal nor maternal IGF2 levels were significantly related to fetal weight in either group of animals. Prenatal exposure to betamethasone significantly reduced fetal liver weight and IGF1 and 2 levels in later gestation. These changes may underlie the significant reductions in birth weight. The observation that lamb weights at 6 and 12 weeks postnatal age were not different between the two groups might imply different growth trajectories in the immediate postnatal period of offspring from mothers treated with glucocorticoid during pregnancy.

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O51. Differential effects of maternal dexamethasone treatment on thyroid hormone activity in the pregnant ewe and fetus Alison J Forhead, Juanita K Jellyman, Dino A Giussani and Abigail L Fowden Type I 5’-monodeiodinase (5’-MDI) converts thyroxine (T4) to triiodothyronine (T3). In fetal sheep, hepatic 5’-MDI activity increases near term in association with the prepartum cortisol surge. This study investigated the effect of maternal dexamethasone treatment, in clinically relevant doses, on hepatic and renal type I 5’-MDI activity and plasma thyroid hormone concentrations in ewes and their fetuses. From 125d of gestation (term 145 + 2d), 21 ewes, carrying single fetuses, were injected twice i.m. with either saline (2ml 0.9% NaCl, n=11) or dexamethasone (2 x 12mg in 2ml 0.9% NaCl, n=10) at 24h intervals. Eleven of the ewes and fetuses were chronically catheterised between 116-119d of gestation, and arterial blood samples were taken before, at 24h after each injection and at 48h after the last injection. The 10 remaining non-instrumented ewes and fetuses were euthanased 10h after the second injection, and tissues and umbilical arterial blood were collected. Plasma T4 and T3 were measured by radioimmunoassay, and tissue type I 5’-MDI activity by outer ring deiodination of iodinated reverse-T3. Data (mean + SEM) were analysed by unpaired t-test and linear regression. In the fetuses, plasma T3, but not T4, concentrations were increased by maternal dexamethasone treatment (p<0.001). Fetal hepatic type I 5’-MDI activity in the dexamethasone-treated group (2308 + 112 pmoles I- (mg protein)-1 h-1) was greater than in the saline group (1221 + 45, p<0.001). Using data from all fetuses, plasma T3 correlated with hepatic type I 5’-MDI activity (r=0.94, n=10, p<0.001). There was no difference in renal type I 5’-MDI activity between the saline and dexamethasone groups of fetuses (28.8 + 3.8 vs 35.3 + 4.1 pmoles I- (mg protein)-1 h-1). In contrast, in the ewes, plasma T3 and T4 concentrations were reduced by dexamethasone treatment (p<0.001), without any change in hepatic or renal type I 5’-MDI activity. Therefore, in pregnant sheep, maternal dexamethasone treatment has different effects on the thyroid hormone axis of the mother and fetus. Changes in T3 bioavailability may mediate some of the maturational effects of dexamethasone in the preterm fetus. This work was supported by the BBSRC and Tommy’s, the baby charity. O52. Maternal Exposure to Glucocorticoids in Early Pregnancy Causes Fasting Hyperglycaemia and Hyperinsulinaemia in Adult Male Sheep Offspring De Blasio MJ, Dodic M, Wintour-Coghlan EM, Robinson JS, Owens JA. Fetal exposure to adverse intra-uterine environments at critical periods during development increases the risk of adult-onset cardiovascular and/or metabolic disease. Exposure to maternal undernutrition during early gestation (Dutch Winter Hunger Famine) resulted in obesity and increased plasma cholesterol and the incidence of coronary heart disease in adults, whereas exposure in mid-late gestation impaired glucose tolerance. It has been proposed that programming effects seen in offspring exposed to famine are due in part to endocrine responses to altered nutrient availability. Severe morning sickness leading to maternal undernutrition can increase plasma cortisol. In sheep, dexamethasone treatment of pregnant ewes (0.24mg/kg/day maternal body weight), in early pregnancy (26–28 days of gestation), impairs adult cardiovascular function but not metabolic homeostasis in adult female offspring. It is unknown what effect maternal glucocorticoid exposure in early gestation has on male adult offspring, who are often more susceptible to programming. Pregnant sheep were infused with saline, dexamethasone (0.48mg/hour) or cortisol (5mg/hour), from 26-28 days gestation (term ~150 days). Size at birth, glucose tolerance (intravenous glucose tolerance test, IVGTT), insulin secretion (during IVGTT), and whole body insulin sensitivity (hyperinsulinaemic euglycaemic clamp) were measured in 4-year-old adult male offspring. Plasma metabolites were measured by enzymatic assay, and plasma insulin by radioimmunoassay. Glucocorticoids caused fasting hyperglycaemia, while dexamethasone improved glucose tolerance. Glucocorticoids did not alter insulin sensitivity, but induced fasting or stimulated hyperinsulinaemia. Dexamethasone induced first phase hyperinsulinaemia and increased the stimulated insulin disposition index for glucose (insulin action), whereas cortisol induced second phase hyperinsulinaemia. Maternal glucocorticoid exposure impairs fasting glycaemia, but does not impair glucose tolerance in adult male offspring. The latter is maintained or even improved by hyperinsulinaemia following glucocorticoids, but nevertheless may lead to pancreatic exhaustion and impaired glucose tolerance with ageing. Glucocorticoid specific effects on first and second phase insulin secretion were evident and may reflect targeting of different aspects of endocrine pancreatic function or may indirectly reflect impairment of insulin sensitivity in different tissues. The mechanisms involved and specific tissues targeted require further investigation.

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O53. Effects of Antenatal Glucocorticoids on Development of Fetal Brain Function M. Schwab, K. Schwab, T. Groh, M. Kott, P.W. Nathanielsz and O.W. Witte Objective: To determine the effects of antenatal glucocorticoids (GC) administered to accelerate fetal lung maturation on development of fetal electrocortical activity. Methods: Chronically instrumented pregnant sheep were treated with single (112 days gestation, dGA, term 150 dGA) or repetitive courses of betamethasone phosphate (Celestan solubile) at 106, 112 and 118 dGA (n=8) or an equal amount of saline (n=7). Each course consisted of 2x110g/kg betamethasone 24h apart equivalent to 2x8 mg betamethasone administered to a 70 kg pregnant woman. Fetal electrocorticogram (ECoG) was recorded continuously between 105-130 dGA. Artifact-free one hour ECoG-epochs were analyzed using spectral analysis. Results: Betamethasone administered at 106 dGA when the sleep states are not yet matured leads to maturation of REM sleep but not NREM sleep ECoG within four days (p<0.05). The power spectrum of the ECoG was similar to that in controls at 130 dGA. Repetitive treatment did not have any further effect on ECoG maturation. Early maturation of REM sleep was accompanied by fragmentation of REM and NREM sleep states that remained fragmented until the end of the examination period. In addition to the maturational effects, betamethasone suppressed acute and transiently electrocortical activity from 112 dGA onwards, i.e. when the sleep states matured (p<0.05). Betamethasone treatment also disrupted diurnal variations of REM sleep state length in agreement with acute behavioral state changes and the delay of the latency of auditory evoked potentials during betamethasone administration in human pregnancy (p<0.05). The acute effects on the ECoG was abolished by preceding betamethasone treatment. Conclusions: A single course of betamethasone induces maturation of cortical neuronal activity but not of thalamic pacemaker circuits at the expense of sleep state fragmentation. Sleep state fragmentation has been shown to continue until adulthood after prenatal stress in rats. The absence of acute suppression of the ECoG during repeated courses of betamethasone indicates programming of glucocorticoid resistance. The extent to which these changes affect post-natal brain function remain to be determined. O54. Prenatal corticosterone exposure in the rat results in hypertension and low nephron number. Reetu R Singh, Luise A Cullen-McEwen, Michelle M Kett, Wee-Ming Boon, Karen M Moritz, John F Bertram Development of hypertension in adult life may result from the fetus being exposed to a sub-optimal intrauterine environment. Using the synthetic and natural (in sheep only) glucocorticoid, a low nephron number and hypertension have been reported in the rat and sheep. As yet, no study has been undertaken to examine the direct effect of elevated maternal concentrations of the natural glucocorticoid, corticosterone (CORT), on blood pressure and nephron endowment in the rat. In the present study, the aims were: 1) to determine blood pressure and nephron number in male and female offspring of rat dams exposed to high but physiological concentrations of corticosterone for 2 days 2) to investigate if components of the renal renin-angiotensin system (RAS) were altered by CORT treatment. Time-mated Sprague Dawley rats were injected intraperitoneally twice daily on days (embryonic day 14) E14 and E15 of pregnancy with either saline (0.2 ml) or corticosterone (0.8mg/kg/day). Kidneys (embryos/offspring) were collected at E16, E20 and (postnatal day 30) PN30 for gene studies and at PN30 for nephron number determination. Nephron number was determined using unbiased stereology while gene studies were performed using Real-Time PCR (using 18S as the housekeeping gene). Mean Arterial Blood pressure (MAP) and Heart rate measurements were obtained in PN120, conscious, unrestrained animals, using the tail artery catheter method. Results: Body weights were not affected following prenatal CORT exposure. CORT increased MAP in both sexes (male: SAL=1142mmHg, CORT=1221mmHg); (female: SAL=1121mmHg, CORT= 1221mmHg). Heart rates were not different between the treatment groups or the sexes.

Age/Sex *P<0.05

E16 SAL

E16 CORT

E20 SAL

E20 CORT

PN30 SAL (M) PN30 CORT (M)

PN30 SAL (F)

PN30 CORT (F)

Nephronnumber - - 28514 ±932 *22522±522 24867±566 *20036±546 L.Kid.vol (mm3) - - - - 419±17 411±22 444±32 360±21 AT1aR/18S 1.0±0.1 *1.3±0.1 1.3±0.5 1.5±0.4 1.1±0.2 *1.7±0.2 1.0±0.1 1.0±0.1 AT1bR/18S 1.2±0.6 *0.7±0.2 1.0±0.3 *0.5±0.4 1.0±0.1 *3.7±0.4 1.6±0.7 *4.3±0.9 AT2R/18S 1.0±0.1 *2.0±0.3 0.9±0.3 0.8±0.2 1.2±0.4 *2.3±0.6 1.3±0.4 *9.6±1.2

Conclusion: This is the first report demonstrating that prenatal exposure to corticosterone results in a nephron deficit and adult hypertension in both male and female rat offspring. More importantly, alterations in the renal renin-angiotensin system have also been observed in this study, at both embryonic and adult life stages and may elucidate the mechanism contributing to the nephron deficit and hypertension observed in this study.

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FETAL AND NEONATAL PROGRAMMING

Free Oral Communications 8 Chairs: Charles Wood (USA) and Tim Moss (Australia)

Tuesday September 27, 2005 1430 - 1630

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O55. The Cytokine Response to Endotoxin is Altered in Early Life following a Prenatal Immune Challenge UNicolette A.U Hodyl, Klara M. Krivanek and Deborah M. Hodgson Exposure to infection during pregnancy is known to be a major factor in predicting future health outcomes for the offspring. In previous studies, we have demonstrated that exposure to an immune stimulus during pregnancy results in altered immune and HPA function in the adult offspring. The current study addresses the short-term effects of such exposure on both HPA axis function and on two, primary, immune-mediating cytokines - tumour necrosis factor alpha (TNFα) and interleukin-1beta (IL-1β). A sustained bacterial infection was mimicked in pregnant rats with exposure to endotoxin (200ug/kg, s.c.) on gestational days 16, 18 and 20. Both the glucocorticoid and the immune (TNFα and IL-1β) responses to an immune challenge (Salmonella Enteritidis, 50ug/kg, i.p.) were tested in the neonate offspring (postnatal day (PND) 5) and during the pre-weaning period (PND 19). Results demonstrate that the offspring of mothers exposed to endotoxin during gestation exhibited a significantly lower corticosterone response to the endotoxin challenge than the control offspring (p<0.05) at both 5 and 19 days. Similarly, the TNFα and IL-1β response to the endotoxin challenge was also significantly reduced in the offspring of endotoxin treated mothers compared to that of control offspring (p<0.05). These results suggest that the ability of offspring to respond appropriately to an immune challenge in early life is reduced following prenatal endotoxin exposure. As the early life period is one characterized by significant natural bacterial invasion, the current evidence suggests that the normal physiological response to such invasion can be altered by prenatal factors, such as maternal immune activation. This has implications on the developmental trajectory of both the neuroendocrine and immune systems, and ultimately on both short- and long-term health outcomes. O56. Neonatal Leptin Treatment Reverses Developmental Programming UVickers MHU, Gluckman PD, Coveny AH, Hofman PL, Cutfield WS, Gertler A, Breier BH, Harris M. An adverse prenatal environment may induce long-term metabolic consequences, in particular obesity and insulin resistance. Although the mechanisms are unclear, this ‘programming’ has generally been considered an irreversible change in developmental trajectory. Adult offspring of rats subjected to undernutrition during pregnancy develop obesity, hyperinsulinaemia and hyperleptinemia, especially in the presence of a high fat diet. Reduced locomotor activity and hyperphagia contribute to the increased fat mass. Using this model of maternal undernutrition, we investigated the effects of neonatal leptin treatment on the metabolic phenotype of adult female offspring. Virgin Wistar rats were time mated and randomly assigned to receive food either ad-libitum (control group) or at 30% of ad-libitum intake (small for gestational age (SGA) group). Offspring from SGA mothers were significantly lighter and shorter at birth than control offspring (p<0.0001). Leptin treatment (rec-rat leptin, 2.5 ug/g/day, sc) from postnatal d3 to d13 resulted in a transient slowing of neonatal weight gain, particularly in programmed offspring. At weaning, offspring were weight matched to be fed a standard chow diet (C) or a high fat (HF) diet ad-libitum until the end of the trial. SGA HF animals treated neonatally with saline showed a marked amplification in diet-induced weight gain compared to control HF saline treated animals (programming x diet interaction p<0.0001) and displayed obesity, hyperinsulinaemia, hyperleptinemia, elevated c-peptide levels, increased food intake and reduced locomotor activity in adult life. Neonatal leptin treatment completely normalised caloric intake, locomotor activity, body weight, fat mass, and fasting plasma insulin, c-peptide and leptin concentrations in programmed SGA offspring in adult life. Moreover, neonatal leptin had no demonstrable effects on the adult offspring of normally fed mothers. This study suggests that developmental metabolic programming is potentially reversible by an intervention late in the phase of developmental plasticity. The complete normalisation of the ‘programmed’ phenotype by neonatal leptin treatment implies that leptin has effects that reverse the prenatal adaptations resulting from relative fetal undernutrition. This work was supported by the Health Research Council of New Zealand and the National Research Centre for Growth and Development.

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O57. Influence of Maternal Parity on mRNA Abundance for the Prolactin and Insulin-like Growth Factor Receptors and Adipose Tissue Deposition in the Resulting Offspring J. Bispham, T. Stephenson, H. Budge and M.E. Symonds Weight at birth increases with maternal parity but the extent to which this is accompanied by changes in fat mass in the resulting offspring is unknown. Both the prolactin (PRLR) and insulin-like growth factor (IGFR) receptors are highly abundant in newborn adipose tissue and they are nutritionally regulated1, 2. The following study determined whether differences in fat growth over the first month of life between primiparous (P) and multiparous (M) offspring are related to differences in mRNA abundance for these receptors. Fourteen twin-bearing ewes of similar body weight were entered into the study of which 8 were P and 6 M. Within 24 h of birth one randomly selected twin from each mother was humanely euthanased and perirenal adipose tissue sampled. The remaining twin was reared with its mother until 30 days of age when it was similarly sampled. The relative abundances of the long and short forms of PRLR and IGF-I and –IIR mRNA to 18S rRNA were determined using RT-PCR and expressed in arbitrary units (a.u.). Despite there being no effect of parity on total body weight perirenal fat mass was greater in P offspring at one month of age (P 15023; M 10114 g (P<0.01)) and was accompanied by a significant increase in abundance of mRNA for both the long and short forms of PRLR at one and 30 days of age (e.g. at 30 days’ long form PRLR - P 13721; M 4510 a.u. (P<0.01)). In contrast, mRNA abundance for IGF-I and -IIR was only increased in P offspring at one day of age (e.g. IGF-II R - P 12110; M 737 a.u. (P<0.01)). By 30 days postnatal age IGFR expression had dramatically reduced and differences in IGFR with parity were abolished. In conclusion, maternal parity has a pronounced impact on fat deposition in the resulting progeny, with P offspring depositing more fat after birth which may be partly mediated by increased sensitivity to prolactin. 1. Bispham J. et al. Endocrinology 144, 3575-3585 (2003) 2. Budge H. et al. Pediatric Research 47, 781-786 (2000).

O58. Insulin Resistance Following Fetal Growth Restriction: Is Intramyocellular Obesity to Blame? Melanie Tran, Irving Lee, Robyn Taylor, Miles De Blasio, Jeffrey S Robinson and Julie A Owens Restricted fetal growth is now recognised as a novel and significant risk factor for diabetes and insulin resistance in later life, but the underlying defects induced to cause these conditions are poorly understood. Recent evidence has shown that restricted fetal growth is characterised by insulin resistance of glucose uptake and utilisation in skeletal muscle in the adult. As in insulin resistance caused by other factors, this may be due to impaired 5’ AMP activated protein kinase (AMPK)/ malonyl CoA fuel sensing and activity, dysregulated lipid metabolism and accumulation of intramyocellular lipid and related metabolites, which impair insulin signalling and its targets. We have shown that fetal growth restriction induces insulin resistance and increases intramyocellular lipid in skeletal muscle in the adult guinea pig. AMPK in skeletal muscle and other tissues can be activated pharmacologically by AICAR (5-aminoimidazole 4-carboxamine-riboside), lowering blood glucose and lipids and intracellular lipids and increasing insulin sensitivity in the rat. We hypothesized that young adult guinea pigs would respond similarly to AICAR by acutely lowering plasma glucose, free fatty acids and triglycerides and that restriction of fetal growth and size at birth would impair this metabolic responsiveness. Vehicle or AICAR (60mg/kg iv) was administered to 25 young adult guinea pigs (10 males, 15 females) of varying birth weight, blood sampled before and after and plasma glucose, triglyceride and free fatty acid concentrations determined. AICAR transiently reduced plasma triglycerides and free fatty acids within ten minutes and reduced plasma glucose from 15 to 45 minutes, consistent with AMPK activation and downstream responsiveness in this species. Basal plasma glucose was unchanged and plasma triglyceride and free fatty acids were lower in low compared to high birth weight guinea pigs. Plasma glucose fell less in response to AICAR in low compared to high birth weight guinea pigs. This suggests that reduced AMPK activation, deficiency or down stream resistance to its actions may contribute to increased intramyocellular lipid and insulin resistance following fetal growth restriction.

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O59. Increased IGF Bioavailability and Altered IGF Binding Proteins During Catch-up Growth in the Young Lamb Following Placental Restriction. Kathryn L Gatford, Brenton D Bennett, Miles J De Blasio, Patricia A Grant, Robyn L Taylor, Jeffrey S Robinson and JA Owens Accelerated or catch-up growth occurs in most human neonates who were small for gestational age at birth. Although catch-up growth positively predicts adult height, it is a risk factor for later obesity, insulin resistance and cardiovascular disease. Neonatal plasma IGF levels are reduced at birth in SGA infants, remain low in those who fail to catch-up, but increase into the normal range in those who do, suggesting that the insulin-like growth factor (IGF) axis may play a role in catch-up growth. We have therefore investigated the effects of experimental placental restriction (PR) on circulating IGFs and IGF-binding proteins in the neonatal lamb. Placental growth was restricted by surgically removing the majority of endometrial implantation sites before pregnancy. Plasma was collected from catheterised fed lambs (12 control, 12 PR) at 5, 10, 15, 20, 25, 35 and 43 days of age, and subjected to size-exclusion high pressure liquid chromatography at pH 2.5. IGFs and IGF-binding capacity were measured by radioimmunoassay in fractions containing free IGFs and IGFBPs respectively. Weight and other size parameters of lambs were measured at birth and at 5-day intervals until post-mortem at 43 days of age. PR reduced placental weight by 20%, and reduced the birth weight of singleton lambs by 19.5% (P=0.049). PR also increased fractional growth rate (control, 6.5 + 0.4 %.d-1, PR, 7.7 + 0.3 %.d-1, P=0.042), and visceral fat at 43 d (P=0.038). PR did not alter plasma IGF-I or IGF-II, but did reduce plasma IGF-binding capacity (an index of total IGFBP), and increased the ratios of plasma IGF-I and IGF-II to IGF-binding capacity (indices of IGF bioavailability). Preliminary Western blot analysis suggests that PR lambs have less IGFBP-3, but more smaller IGFBPs than control lambs. Ratios of plasma IGF-I to IGF-binding capacity correlated positively with fractional growth rates for weight and the ratios of plasma IGF-I or IGF-II to IGF-binding capacity correlated positively with visceral fatness. Therefore placental restriction alters the abundance of multiple IGFBPs and increases IGF bioavailability postnatally, which in turn predicts and may contribute to neonatal catch-up growth and increased adiposity following fetal growth restriction.

O60. Dietary Zinc Supplementation Ameliorates LPS-Induced Teratogenicity In Mice JSC Chua, AM Rofe and P Coyle Maternal infection during the first trimester of pregnancy has been associated with pre-term birth, spontaneous abortion, growth retardation and congenital anomalies. Previously, our group have shown that subcutaneous injection of zinc prevents endotoxin (LPS)-induced teratogenicity (1). The purpose of this study was to investigate whether increasing or decreasing dietary zinc alters the teratogenic effects of LPS. Female C57BL6 mice were mated and placed on diets containing 5, 35 or 100 mg/kg zinc. On gestational day (GD) 8, pregnant dams were injected with either LPS (0.5 mg/kg sc) or saline and killed on GD18. LPS-treated fetuses from dams fed 5 & 35 mg/kg zinc diet had a significantly higher number of abnormalities per litter (2- & 1- fold saline controls, respectively) compared to those from LPS + zinc supplemented dams, which were not significantly different from the saline control groups. The beneficial effect and importance of zinc was also reflected in the larger size of fetuses (weight and crown-rump length) from the LPS + zinc supplemented treatment group. We have demonstrated that low dietary zinc during exposure to infection (i.e. LPS) in pregnancy augments the negative impact of LPS alone, and that dietary zinc supplementation throughout pregnancy ameliorates LPS-induced teratogenicity. 1. Carey, L.C., Berbee, P.L., Coyle, P., Philcox, J.C., and Rofe, A.M. 2003. Zinc treatment prevents lipopolysaccharide-induced teratogenicity in mice. Birth Defects Res Part A Clin Mol Teratol 67:240-245.

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O61. Dietary Zinc Prevents Birth Abnormalities and Postnatal Death Caused By Prenatal Ethanol Exposure BL Summers, AM Rofe and P Coyle Subcutaneous Zinc (Zn) treatment given together with ethanol in early pregnancy has previously been shown to protect against alcohol-related physical birth abnormalities in mice. The current study examined whether dietary Zn supplementation throughout pregnancy can also prevent physical abnormalities, as well as postnatal death caused by prenatal ethanol exposure. Pregnant dams were injected with saline (0.85% w/v NaCl) or 25% ethanol (0.015ml/g mouse) intraperitoneally at 0 and 4hr on gestational day (GD) 8. Dams were fed either a control diet (35mg Zn/g), or a Zn supplemented diet (200 mg Zn/g) throughout pregnancy. At GD18, dams supplemented with dietary Zn throughout pregnancy had increased plasma Zn concentrations (>1-fold) compared to unsupplemented saline and ethanol dams (p<0.05). Fetuses prenatally exposed to ethanol alone had a higher incidence of physical abnormalities compared to saline treatment alone, as well as Zn supplemented saline and ethanol-exposed fetuses (P<0.05). In a separate group of mice, postnatal death was analysed by counting the number of surviving pups from birth until postnatal day (PD) 60. The risk of dying from birth until PD60 in the ethanol-treated litters (65% survival) was significantly greater than in the other three treatment groups (95% survival) (P<0.05). These findings demonstrate that dietary Zn supplementation throughout gestation can limit the physical abnormalities and the number of postnatal deaths caused by ethanol exposure in early pregnancy. O62. Maternal IGF-II Treatment in Early Pregnancy Improves Placental Structural and Functional Development and Fetal Growth Near Term AN Sferruzzi-Perri, JA Owens, KG Pringle, JS Robinson and CT Roberts The placenta transports substrates and wastes between the maternal and fetal circulations and is essential for the health of both the mother and fetus. Poor placental development impairs fetal growth and is associated with pregnancy complications which are potentially life threatening for both the mother and child and for which there are no effective therapeutics. Maternal circulating insulin-like growth factors (IGFs) increase in early pregnancy and treatment of the pregnant guinea pig with either IGF-I or IGF-II increases placental and fetal weights by mid-gestation. This study examined whether such maternal IGF treatment in early pregnancy could sustainedly promote placental structural development and glucose transport and thereby, growth of the fetus. Pregnant guinea pigs received IGF-I, IGF-II (both 1mg/kg/day) or vehicle subcutaneously from days 20 to 38 of pregnancy (term ~70 days). Transfer of [3H]-methyl-D-glucose (3MG) in vivo was measured on day 62, just prior to maternal death, when fetal and placental weights and maternal body composition were also measured. Placental structure was analysed morphometrically in mid-sagittal sections. IGF-II increased the mid-sagittal area and volume of placenta devoted to exchange (both +30%), but did not alter placental weight. The total surface area for placental exchange (+39%), the volume of trophoblast and that of maternal blood spaces within the exchange region of the placenta (+29% and +46%) were also increased by IGF-II. In contrast, IGF-I did not alter placental structure, but reduced maternal adipose tissue stores by ~30%. Both IGFs increased total placental 3MG uptake by 40-60% and fetal plasma 3MG concentrations by ~40%. IGF-II increased the number of viable fetuses by 26% and both IGFs increased fetal growth by 11-17%. In conclusion, maternal IGF-II treatment in early pregnancy promotes fetal growth and viability by substantially increasing placental structural and functional capacity, while IGF-I may act by diverting nutrients from the mother to the conceptus, near term. As poor placental development impairs fetal growth and survival, maternal IGF treatment may have therapeutic potential in pregnancies at risk.

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32ND FNPS 2005 DATA BLITZ ABSTRACTS

DATA BLITZ POSTER SESSION

Chairs: Julia Pitcher (Australia) and Miles De Blasio (Australia)

Sunday September 25, 2005 1630 - 2000

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DB1. Maternal dexamethasone treatment increases hepatic glycogen content but not G6Pase activity in the pregnant rat and fetus UKL Franko U, AJ Forhead, AL Fowden Synthetic glucocorticoids such as dexamethasone (dex) are administered clinically to mimic the maturational effects of the natural fetal prepartum glucocorticoid surge (1). In fetal sheep, these maturational changes include an increased glucogenic capacity mediated partially by increased hepatic glycogen deposition and increased activity of glucose-6-phosphatase (G6Pase) (2). G6Pase is the final enzyme in both the glycogenolytic and gluconeogenic pathways, while glycogen is a limiting factor in glycogenolysis. This study measured G6Pase activity and hepatic glycogen content in the pregnant rat and fetus after maternal dex administration. Pregnant Wistar rats were injected subcutaneously with 200ug/kg Dexamethasone Phosphate (dex, n=7) from d15-19 of a 21-day gestation. Controls were injected with an equal-volume of saline (0.9% NaCl) for the same period (n=7). On d20, fetal and maternal tissues were collected after terminal anaesthesia and frozen for later analysis of G6Pase activity (2) and glycogen content (3). Sex of the pups was determined by measuring ano-genital distance. Tissue from two males and two females from each litter was analysed. Litter means are reported below. Maternal dex administration had no effect on hepatic G6Pase activity in either mothers or fetuses (Table 1). Both fetal and maternal hepatic glycogen contents were higher in the dex than saline treated animals, but this only reached statistical significance in the dex-treated mothers (Table 1). Data were analysed by unpaired t-test. Table 1: G6Pase activity and glycogen content of maternal and fetal livers after maternal dex administration. Maternal Liver Fetal Liver Saline Dex Saline Dex G6Pase Activity (U/g) 11.3 U+U 1.6 8.1 U+U 0.7 1.9 U+U 0.1 2.0 U+U 0.3 Glycogen Content (mg/g) 12.2 U+U 3.4 41.4 U+U 8.4* 49.0 U+U 6.7 62.7 U+U 3.9 *Significant difference from controls (p<0.01) Dexamethasone treatment therefore increases the maternal but not the fetal glucogenic capacity by increasing glycogen availability. This has implications for glucose homeostasis in women treated antenatally with glucocorticoids. 1. Fowden AL et al., (1998). Proc. Nutr. Soc. 57 113-122. 2. Fowden AL et al., (1993). J. Endo. 137 213-222 3. Roehrig KL and JB Allred, (1974). Ana. Biochem. 58 414-421 DB2. Differential effects of maternal cold exposure on mRNA abundance for the growth hormone secretagogue receptor (1α) and glycerol-3-phosphatase in adipose tissue of the newborn sheep. EA Butt, S Pearce, T Stephenson, ME Symonds and UH BudgeU Fetal adipose tissue growth is a coordinated process involving the growth hormone-insulin-like growth factor (GH-IGF) axis (Symonds et al. 2003). Maternal cold exposure (CE) induced by winter shearing can promote fetal fat growth depending on maternal food intake (Pearce et al. 2005). The following study examined the interaction between maternal CE and dietary intake on the mRNA abundance for the endogenous ligand of the GH secretagogue receptor (GHSR-1α), the IGF-I and II receptors and the enzyme glycerol-3-phosphatase (G3P) which has a role in regulating lipolysis. Twenty-six multiparous ewes of similar body weight were entered into the study, thirteen were shorn (S) at 70 days gestation and thirteen remained unshorn (US), with 7 ewes per group being nutrient restricted (NR) over the final month of gestation, consuming 50% of the intake of controls. All offspring were humanely euthanased at one day of age to enable perirenal adipose tissue sampling. The relative abundances of GHSR-1a and G3PMrNA to 18S rRNA were determined using RT-PCR and are expressed in arbitrary units (a.u.). Maternal cold exposure resulted in a significant up regulation of mRNA for the GHSR-1aMrNA (S 9.9 U+U 3; US 3.2 U+U 1 (p<0.01)). Whereas mRNA for the IGF-I and II receptors were unaffected, G3P was reduced (S 2.5 U+ U 0.2; US 3.9 U+U 0.2 (p<0.05). These adaptations in GHSR-1a and G3P were further enhanced in offspring born to CE mothers that were NR. Spearman’s test showed a negative correlation between the GHSR-1a and G3P genes (p<0.001). In conclusion, the anabolic effects of maternal CE on fat deposition in the fetus are partly mediated by increased sensitivity to the GH secretagogue receptor -1α which also has anti-lipolytic effects. Pearce S et al. 2005 Adipocytes 1: 57-64. Symonds ME et al. 2003 J Endocrinol 179: 293-299.

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DB3. The Impact of Placental Restriction on the Expression of the Appetite Neuropeptide Regulators, NPY and POMC, in the Hypothalamus of the Fetal Sheep in Late Gestation. BHA Chuang, BS Muhlhausler, JL Morrison, SJ Williams, CL Adam, PA Findlay and IC McMillen Epidemiological studies have demonstrated that low birth weight infants are at increased risk of obesity in later life. It has been suggested that this may be the result of changes to the central neural network which regulates appetite in postnatal life. We have therefore determined the effect of fetal growth restriction on the expression of the appetite-regulators NPY (appetite stimulation) and POMC (appetite inhibition) in the hypothalamus of the fetal sheep in late gestation. Placental and fetal growth restriction was induced by removal of the majority of the uterine caruncles before mating (PR, n=6). Vascular catheters were inserted into Control (n=6) and PR fetal arterial vessels at between 117-118 d gestation and blood samples collected for determination of fetal arterial PO2 and glucose concentrations. At 139-141 d gestation, postmortem was performed and brains were collected. In situ hybridization and quantitative image analysis was then used to determine the level of expression of NPY and POMC in the hypothalamic arcuate nucleus. Fetal weight (2.63kg vs 5.07kg, P<0.01), plasma glucose (0.99mmol/l vs 1.70mmol/l, P<0.01), mean gestational arterial PO2 (12.96mmHg vs 20.85mmHg, P<0.01) and arterial PO2 on the day of postmortem (12.78mmHg vs 20.14mmHg, P<0.01) were each lower in PR fetuses compared to Controls. There was no significant difference in the expression of either POMC or NPY between the PR and Control groups. There was, however, an inverse relationship between hypothalamic expression of NPY and fetal weight (R2=0.36, P<0.05) and absolute brain weight (R2=0.41, P<0.05) at 139-141 d gestation when data from both groups were combined. Hypothalamic NPY expression was also inversely related to arterial PO2 (R2=0.49, P<0.05), but not plasma glucose concentrations, on the day of post mortem. In the PR group, but not Controls, hypothalamic expression of POMC and NPY were directly related (R2=0.88, P<0.02). We have therefore demonstrated that the hypothalamic expression of the appetite-stimulating neuropeptide, NPY is directly related to the extent of fetal growth restriction. This provides evidence that an important component of the appetite-regulating system is responsive to reduced fetal substrate supply, and this may have potential consequences for the regulation of appetite in low birth weight offspring in postnatal life. DB4. Timing of delivery of IUGR fetuses on the basis of hemodynamic changes EV Cosmi, G. Mari and E. Cosmi

Objective: to evaluate the morbidity and mortality rate of IUGR fetuses in relation to Doppler velocimetry, FHR tracing, BPP and AFI findings. A multicenter prospective study of 346 fetuses with ultrasound diagnosis of IUGR. Among them 286 met the study entry criteria. The study group underwent Doppler velocimetry study of UA, MCA, ductus venosus and umbilical vein twice weekly, BPP and AFI twice weekly and FHR tracing daily. IUGR fetuses were divided into two groups: Group 1 consisting of IUGR fetuses with Doppler velocimetry abnormality; Group 2 with normal Doppler velocimetry. In Group 1 the type of Doppler alteration and time passing from Doppler velocimetry alteration to birth were recorded. Apgar score, pH at birth, need of intubation, RDS, IVH, PVL, days of NICU hospitalization were available. Neonatal outcome was related in Group 1 with Doppler velocimetry, FHR tracings, BPP and AFI abnormality. Statistical analysis was performed using linear regression analysis to correlate the type of Doppler velocimetry alteration, the time passing from Doppler alteration and birth to neonatal outcome; Group 2 was taken as control group. Gestational age at time of admission ranged from 16.1 to 40 weeks of gestation. Group 1 consisted of two-hundred fetuses, while Group 2 consisted of 86 fetuses. Group 1 fetuses showed the following Doppler velocimetry alterations: increased resistance of the UA (100%); UA AEDF (52%), UA RF (26%); brain sparing effect in the MCA (38%); abnormality in the ductus venosus (absent or reversed flow) and umbelical vein pulsations (20%). BPP and FHR tracing were abnormal in 30% of the fetuses and in all of them these changes occurred after Doppler velocimetry alterations with a mean time of 3.8 days. AFI was below the normal ranges in 42% of fetuses. 4.7% of Group 2 fetuses presented FHR tracing alterations and BPP < 8; AFI was below the normal ranges in 36% of fetuses. Apgar score was < 7 in 49% in Group 1 and in 4.7% in Group 2 (p< 0.005); there was not a statistical difference in days of hospitalization and in NICU. Morbidity and mortality rate was significantly higher in Group 1(p< 0.005): 26% of Group 1 fetuses developed neurological sequelae, sepsis, IVH and RDS; 6.3% fetuses developed RDS because of prematurity; 24% of Group 1 fetuses died after NIUC hospitalization and one in utero. In Group 2 mortality rate was 4.7%. In Group 1 morbidity and mortality rate were significantly correlated with the type of Doppler velocimetry abnormality and with time passing from the latter finding and delivery being higher in fetuses with AEDF, RF in UA with a time > 72 hours from the Doppler abnormality and delivery. Doppler velocimetry should be performed in IUGR fetuses as its findings help to reduce morbidity and mortality rate. There is a significant correlation between the type of alteration and time passing from alteration to birth. Morbidity and mortality rate are strongly decreased if delivery is performed within 72 hours in IUGR fetuses with AEDF of RF in the UA, thus an aggressive management in these fetuses is suggested.

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DB5. The vasoreactivity of small placental arteries and systemic vascular bed Noriaki Imai, Minori Saito, Junichi Sugawara, Yoshitaka Kimura, Kunihiro Okamura Background: The control of the blood flow within the fetoplacental circulation has been poorly understood. To assess the characteristics of human placental chorionic plate small arteries, we examined those vasoreactivity compared with systemic resistance arteries by using wire myograph. Materials and methods: Twelve placentas were obtained from term vaginal deliveries or caesarean sections of women with uncomplicated pregnancies. Six omentums were obtained from women having omentectomy for ovarian cancer. The response to a variety of contractile agonists for example norepinephrine, arginine vasopressin and U46619 (thromboxiane A2 mimetics) was examined. Arteries were precontracted with U46619 and then exposed to the variety of agonists known to elicit endothelium-dependent or independent relaxation in other vascular beds that were acetylcholine, bradykinin, histamine and sodium. A series of experiments were performed examining the effect of preincubation in NG-nitro-L-arginine methyl ester (L-NAME) on bradykinin and histamine-induced relaxation. Results: Bradykinin produced a considerable relaxation to 91% and 29% of U46619 contraction in omental arteries and placental arteries, respectively. In omental arteries relaxation by bradykinin was not decreased significantly, but in placental arteries it was significantly reduced to 3% by L-NAME. Histamine produced a significant relaxation to 80% in omental arteries, but in placental arteries it did not elicit obvious relaxation. In placental arteries, histamine produced 18% vasoconstriction under L-NAME pretreatment. SNP produced a considerable relaxation in both omental and placental arteries. Conclusions: In this study we have demonstrated that bradykinin induced endothelium-dependent relaxation in isolated human placental small arteries. In placental small arteries, it is possible that NO-cGMP pathway is dominant, and it has been suggested that the pathway via H1 receptor affects vascular tone rather than H2 receptor. In conclusion, it has been suggested that analysis of vasoreactivity of placental small arteries would be important for clarifying the regulation of fetoplacental circulation. DB6. Neonatal Gender Influences Microvascular Function in Normal Newborns Stark MJ, Clifton VL, Wright IMR AIM: Previous studies by our group have demonstrated significant maternal physiological differences dependant upon fetal sex [1]. Neonatal mortality and morbidity are significantly related to gender. Our aim was to clarify normal neonatal microvascular response in relation to gender. METHOD: Peripheral microvascular responses were examined using laser Doppler flowmetry according to protocols we have published previously [2]. Healthy male (n=8) and female (n=11) neonates born near term were studied on day 3 of life. All data is expressed as medians (inter-quartile range). Statistical analysis was by Mann-Whitney testing and p<0.05 deemed to be statistically significant. RESULTS: Baseline blood flow in female neonates on day 3 of life was significantly lower than in males (7.6 + 6.0 vs 13.5 + 9.8, p=0.041). The hyperemic response to thermal stimulation at 44oC was also significantly lower in female neonates compared to males (9.2 + 6.7 vs 30.2 + 27.9, p=0.004). There was no significant difference in post-occlusive reperfusion between the genders. CONCLUSION: We have demonstrated that female neonates have a significantly lower peripheral microvascular baseline blood flow and a reduced capacity to respond to thermal hyperemia. Response to heat and baseline vascular tone are thought to be predominantly mediated by nitric oxide and acetylcholine. Thus, the observed difference in microvascular function supports a gender effect on various vasoactive pathways, or a common final pathway. These findings have implications for underlying long-term vascular programming in males and females. [1] M.V. Clifton VL, Placenta. 2003, 168(Supplement A), S45-52. [2] C.V. Crompton R, Bisits AT, Read MA, Smith R, Wright IMR, J Clin Endocrinol Metab. 2003,

88(11), 5427-5432.

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DB7. Leptin infusion does not alter cardiomyocyte development in the sheep fetus in late gestation UK.Botting,U I.C. McMillen, J. Dorosz, J.L. Morrison Fetal growth restriction is associated with an increased risk of cardiovascular disease in adult life. Leptin has been identified as an independent risk factor for coronary heart disease. Leptin receptors have been found in the heart and leptin treatment causes hypertrophy of cardiomyocytes in culture in the rat. We, therefore, have investigated the effect of leptin infusion during late gestation on cardiomyocyte development. Fetal vascular surgery was performed at 117-118 d gestation (term, 150d). At 129-130 d gestation, fetuses were infused with either 200µg/0.5ml sterile saline ovine leptin (n=5) or sterile saline (n=5) for 96 h. A blood sample was taken at the end of the 96 h just prior to post mortem. The heart was reversed perfused with Tyrode’s buffer and collagenase and cardiomyocytes collected by trituration. Cardiomyocytes were fixed in 1% paraformaldehyde. The percent of mononucleated cardiomyocytes in 200 cells and the length and width of 50 binucleated cardiomyocytes was determined. Fetal plasma leptin concentration was greater in the leptin infused fetuses (L, 5.1±0.5 ng/ml) compared to the saline infused fetuses (S, 2.4±0.9 ng/ml). There was no effect of leptin on fetal weight (S, 3.9±0.4 kg; L, 4.7±0.4 kg) or heart to body weight ratio (S, 8.4±0.8 g/kg; L, 7.4±0.4 g/kg). There was no significant effect in the percentage of mononucleated cardiomyocytes in the left or right ventricle between the two groups. Leptin infusion had no significant effect on the length or width of binucleated cardiomyocytes. These data show that 96 h leptin infusion during late gestation in the sheep fetus has no effect on the percentage of mononucleated cardiomyocytes or the size of binucleated cardiomyocytes. DB8. Role of Beta1-adrenergic receptor (β1AR) Pi3K/p70S6k signaling in the regulation of neonatal cardiomyocyte proliferation Yi-Tang Tseng, Naohiro Yano, Ting Zhao, Vlad Ianus, Joan P Stabila, Beth G McGonnigal, UJames F Padbury The growth of cardiomyocytes shifts from hyperplasia to hypertrophy shortly (4-5 days) after birth in rats. The mechanisms underlying this transition are not clear. We have shown that the β1AR is involved in regulation of neonatal cardiomyocyte proliferation. This is mediated in part via the PI3K/p70S6K pathway. We aimed to investigate the ontogeny of the PI3K/p70S6K signaling and to determine which β1AR subtype is involved in regulation of cardiomyocyte proliferation. We demonstrated regulated, coordinate expression of all components of the PI3K/p70S6K signaling pathway. There is a surge in PI3K activity from E20 to postnatal day 3. Its activity remained high in early postnatal life. After postnatal day 7, the activity decreased to low levels seen in adults. The expression and activity of the downstream kinases p70S6K, PDK1 and AKT (PKB) were all also strikingly elevated in late gestation E20/21, P2, P3 and decreased after postnatal day 7 to low levels in adulthood. In contrast, the expression and activity of the phosphatase PTEN, which inhibits PI3K signaling, was very low during the period of cardiomyocyte proliferation and very high in postproliferative stages and in adults. To confirm the role of β1AR regulation, three-day old animals (still in the proliferative period of cardiac growth) were injected with saline, propranolol, metoprolol or betaxolol hearts were collected and assayed for enzyme activity and proliferation index. Beta1AR blockade by metoprolol or betaxolol significantly reduced p70S6K activity and BrdU labeling. This effect was comparable to nonselective blockade by propranolol. We conclude, ontogeny of elements in the cardiac PI3K/p70S6K signaling pathway is synchronized and highly regulated throughout development in the heart. The Beta1AR is responsible for the mitogenic control of neonatal cardiomyocyte proliferation via PI3K/p70S6K signaling. We speculate recapitulation of this signaling pathway may underlie the recently demonstrated ability of adult cardiomyocytes to re-enter the cell cycle following injury. (Supported by NIH 1 P20 RR018728).

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DB9. Effect of Suppression of Neurosteroid Synthesis on Asphyxia-Induced Fetal Brain Injury UYawno TU, Yan EB, Walker DW and Hirst JJ Background: The neurosteroid allopregnanolone is a potent GABAergic agonist that suppresses CNS activity and may be neuroprotective by reducing excitotoxicity in adult animals. Allopregnanolone concentrations are higher in fetal compared to neonate and adult brain (1). Allopregnanolone concentrations in brain extracellular fluid of late gestation fetal sheep rise following asphyxia induced by 10 mins of umbilical cord occlusion (UCO), accompanied by increased expression of 5α-reductase type II in the fetal brain (2). Aim: To determine the effect suppressing neurosteroid synthesis using finasteride on the extent of fetal brain injury caused by 5 min UCO. Finasteride is a 5α-reductase inhibitor that suppresses neurosteroid production and was shown to raise CNS and postural muscle activities in fetal sheep (3). Methods: Ewes underwent surgery at 125-128 days of gestation (GA) for insertion of catheters and the placement of a cuff around the umbilical cord. At 130-132 GA, fetuses received finasteride (40mg/kg/h based on estimated fetal weight, n=4) or vehicle (40% hydroxypropyl-β-cyclodextrin, n=4) for 2 h via the right brachial catheter, the tip of which was placed in the common carotid artery. UCO for 5 mins was done 30 min after beginning the infusions. Brains were collected 24 h after UCO. Brains were dissected into defined blocks, immersion fixed in 4% paraformaldehyde, and 10m sections of the cortex and hippocampus cut for examination of activated caspase-3 by immunocytochemistry as an index of apoptosis. Results: Fetal POB2 B, OB2B saturation and pH were significantly reduced (P<0.05) from pre-UCO values (20.2U+ U1.4 mmHg 74.6 U+ U 3.5 mmHg 7.4U+ U0.01) after 5 mins UCO (6.6 U+ U 1.2 mmHg 7.3 U+ U 0.5 mmHg 7.07U+ U0.03); PCOB2 B was increased from 35.6 U+ U 0.8 to 72.6 U+ U 4.9. At 24 h the number of caspase-3 immunopositive cells in the subcallosal white matter (58373.2 U+U 5818.1 cells per mm2) and the hippocampal CA3 (1432.8 U+ U 236.1 cells per mmP

2P) regions

were increased in the finasteride treated, compared to the vehicle treated fetuses (22488 U+ U 622, 1078.6 U+ U 81.4 cells per mmP

2P, respectively).

Conclusion: Suppressing allopregnanolone production followed by UCO induces increased apoptosis in the hippocampus and white matter regions. Therefore, allopregnanolone might play a role in reducing the amount of cell death that occurs in the fetal brain following in utero asphyxia. 1. Nguyen et al 2003 Pediatr Res 53:956-964 2. Nguyen et al 2004 J. Physiol. 560(Pt 2): 593-602 3. Nicol et al 2001 Neuroscience Letts. 306:13-16. DB10. Secondary post-asphyxial hypoperfusion is associated with suppression of cerebral metabolism and increased tissue oxygenation in near-term fetal sheep UJensen ECU, Bennet L, Hunter CJ, Power GC, and Gunn AJ. Secondary cerebral hypoperfusion is very common during the early phase of recovery from global hypoxia-ischemia at term; however it remains controversial whether it represents true secondary ischemia, impairing tissue oxygenation, or simply is a consequence of reduced cerebral metabolism. We therefore examined the hypothesis that cerebral oxygenation would be reduced during hypoperfusion after severe hypoxia, and further, that blockade of endogenous activation of the adenosine A1 receptor would be associated with both greater hypoperfusion and deoxygenation. Sixteen near-term fetal sheep at 125U+U1 days gestation were infused with 8-cyclopentyl-1, 3-dipropylxanthine (DPCPX), followed by 10 min of asphyxia induced by occlusion of the umbilical cord. Infusions were discontinued at the end of the occlusion. The fetuses then recovered for 3 days. Transient hypoperfusion developed at 2 to 3 h after occlusion (p<0.001). The fall in carotid blood flow was greater in the DPCPX group compared to the vehicle group (p<0.05) at 1 h (81 U+U 7 vs 108 U+U 8 % of baseline) and 2 h (66 U+U 8 vs 95 U+U 12 % of baseline). Local cortical blood flow measured by laser Doppler showed a similar pattern of changes (p<0.05), and significantly correlated with carotid blood flow (r=0.65, p<0.0001). Cortical tissue PO2 was significantly increased at 1 h (7.9 U+U 1.1 Torr), 2 h (7.9 U+U 0.88 Torr), and 3 h (6.9 U+U 0.8 Torr) after occlusion compared to baseline values (4.7 U+U 1.0 Torr), with no significant differences between groups. Cerebral metabolism, measured by changes in cortical heat production, fell significantly 1 h after occlusion (26 U+U 3 vs 35 U+U 4 cal/min before occlusion), with no significant differences between groups. In conclusion, contrary to our initial hypothesis, delayed hypoperfusion following umbilical cord occlusion was associated with marked suppression of cerebral metabolism and a corresponding increase in cortical tissue PO2. DPCPX infusion was associated with a greater secondary fall in carotid blood flow, but no significant effect on either the fall in cortical metabolism or increase in tissue oxygenation. These data strongly suggest that post-asphyxial hypoperfusion is mediated by intact coupling between cerebral blood flow and suppressed cerebral metabolism.

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DB11. Altered fetal brain development and growth following repeated, acute exposure to alcohol: potential mechanisms. Penelope A Dalitz, Sandra M. Rees, Kathryn L. Gatford, Julie A. Owens, Megan L Cock, Philip N Henschke, Richard Harding BACKGROUND: Repeated, acute patterns of maternal alcohol consumption may be particularly harmful to the fetal brain. Alcohol exposure has also been associated with reduced fetal growth. Mechanisms of altered fetal brain development and growth are unclear. OBJECTIVE: Using an ovine model of repeated, acute fetal alcohol exposure, we examined potential mechanisms of fetal cerebral injury and altered growth. METHODS: Ethanol (EtOH) (1g/kg of maternal weight) was administered i.v. to 8 twin-bearing ewes over 1 hour on 3 consecutive days at 1161 days of gestation (DGA: term ~147days); controls received saline (n=7). Fetal and maternal blood alcohol concentrations (BAC) and physiological parameters were measured. At post-mortem (1201 DGA) fetal brains were processed for structural or biochemical analysis. Fetal cerebral white matter (WM) and plasma collected prior to and 1h, 2h, 3h and 6h following EtOH on each treatment day were analyzed for levels of pro-inflammatory cytokines, by ELISA, as indices of fetal inflammation. Fetal plasma collected prior to and 6h after EtOH infusions and 1h, 2h, 3h and 4h after infusion on the first day of EtOH exposure was subjected to size exclusion HPLC at pH 2.5 to examine alterations in growth factors. IGFs and IGF- binding capacity were measured by RIA in fractions containing free IGFs and IGFBPs respectively. Fetal brain tissue was analysed for lipid hydroperoxide (LPO) levels as an indicator of oxidative stress. RESULTS: Maternal and fetal BAC peaked at 0.110.01 g/dL after EtOH infusion on each treatment day. Fetuses did not become hypoxemic or acidemic. At autopsy, mean EtOH body weights were ~400g lower than controls (p=0.02). Fetal plasma IGF-II tended (p=0.099) to be reduced 54h after the first EtOH infusion; IGF-I was unchanged. Cerebral WM injury was observed in 3/8 EtOH fetuses. Apoptosis increased throughout WM of EtOH treated fetuses. Fetal WM and plasma pro-inflammatory cytokine levels were not altered. LPO tended to be elevated in WM, corpus callosum, hippocampus, brainstem, thalamus and spinal cord of EtOH fetuses in comparison to controls. CONCLUSION: The fetal neuropathology following EtOH exposure could involve oxidative stress. Altered fetal IGF levels may contribute to alterations in growth following prenatal alcohol exposure. DB12. Regional specificity of detection of ischemic changes on magnetic resonance imaging in the preterm fetal sheep Mhoyra Fraser, Laura Bennet, Chris E Williams, Peter D Gluckman, Alistair J Gunn, and Terrie E Inder. Background and purpose: There is little information on the ability of magnetic resonance imaging (MRI) to define acute white and grey matter ischemic injury in the preterm infant. We therefore examined in a preterm fetal sheep model of cerebral hypoperfusion, with left-hemispheric placement of a microdialysis probe, whether post- T1 (spin-lattice) and T2 (spin-spin) weighted images predicted neural histopathology.(1) Methods: Mr imaging was performed on paraformaldehyde-fixed fetal sheep brains collected on days 99-100 of gestation, 72 hours after recovery from a 30 min period of bilateral cortical occlusion or sham occlusion. Mr images and histopathological findings of fetuses that exhibited a severe (persistent suppression of electroencephalogram (EEG), n=5) or a mild ischemic insult (brief suppression of EEG, n = 2) were compared with sham-control (n=3) and unoperated fetuses of the same gestational age (n = 4). Results: The ischemic brains demonstrated a variety of changes on MRI, ranging from significant global cerebral injury with diffuse white matter signal abnormality and marked irregularity of the cortical ribbon to mild white matter abnormality with indistinct cortical signal. The MRI findings for both severe and mild ischemic brains were consistent with histological findings and demonstrated the ability of conventional MRI to detect severe abnormalities such as parasagittal cortical infarction and necrosis and subtle pathologic white matter changes manifest by increased microglial cell proliferation. However, histological changes of deep structures such as the corona radiata were not reliably detected. No imaging abnormalities were detected in the unoperated group. Conclusion: This study suggests that in preterm fetal sheep conventional MRI in most cases accurately detected severe neuronal degeneration and white matter loss in many cerebral regions 72 hours following injury, but highlights for the first time some apparent specific, regional limitations. 1. Fraser M, Bennet L, Gunning M, Williams C, Gluckman PD, George S, Gunn AJ: Cortical electroencephalogram suppression is associated with post-ischemic cortical injury in 0.65 gestation fetal sheep. Brain Res Dev Brain Res 154:45-55, 2005

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DB13. Suppression of post hypoxic-ischemic EEG transients with delayed dizocilpine is associated with moderate striatal protection in the preterm fetal sheep Justin M Dean, Alistair J Gunn, Guido Wassink, Sherly George, Laura Bennet The immature brain appears to be particularly vulnerable to post hypoxic-ischemic damage resulting from abnormal glutamate receptor activation. Although the majority of studies are focussed on the preponderance of white matter injury and the role of the AMPA-type of glutamate receptor, recent clinical studies also demonstrate a high incidence of acute damage to subcortical neuronal structures. In vitro studies suggest that activation of the NMDA-type of glutamate receptor contributes to this pattern of injury, but this has not been directly tested. We therefore examined the role of the NMDA receptor in mediating neuronal injury during recovery from severe hypoxia-ischemia. Preterm fetal sheep at 70% of gestation received either sham-asphyxia or asphyxia and were studied for 3 days recovery. Dizocilpine maleate (2mg.kg-1 bolus plus 0.07mg.kg.h-1 i.v.) or saline was infused from 15min post-asphyxia until 4h. In the asphyxia-vehicle group abnormal EEG spiking activity was observed during the first 4h of reperfusion. Dizocilpine significantly suppressed this activity (2.30.9 versus 9.32.3 counts.min-1 at peak frequency, P<0.05) and significantly reduced average EEG intensity from 4h until 11h after occlusion (P<0.05). Dizocilpine was associated with a small increase in fetal extradural temperature during the period of infusion (40.10.2 versus 39.30.1oC at peak, P<0.05). At 3 days recovery there was a significant reduction in neuronal loss in the striatum (317 versus 582%, P<0.05), but not other neuronal areas, in dizocilpine treated animals. Protection was associated with a reduction in cleaved caspase-3 (1117 versus 15910 counts.area-1, P<0.05) and activated microglia (9216 versus 579 counts.area-1, P<0.05) expression. No change in loss of O4-postitive oligodendrocytes in the subcortical white matter was seen. In conclusion, the present studies suggest that abnormal NMDA receptor activation of in the first few hours of recovery from hypoxia-ischemia may be an important contributor to subcortical gray matter damage in premature infants.

DB14. Dose-response study of lipopolysaccharide-induced fetal brain injury in the guinea pig E.L. Harnett, M.A. Dickinson, and G.N. Smith. Chorioamnionitis is associated with an increased risk for fetal and neonatal brain injury. Cytokines, induced by maternal infection/inflammation, are thought to be involved through direct and indirect actions. To date, little is known regarding the association between severity of maternal infection/inflammation and resulting brain injury. It is hypothesized that brain injury will result only after a threshold infection, and concomitant inflammatory response, has been achieved. Maternal intraperitoneal injections of 1, 5, 25, 50, 100 or 200g lipopolysaccharide (LPS) per Kg of maternal body weight or sterile 0.9% saline were administered to guinea pigs (n=35) at 70% gestation. Animals were euthanized 7-days post-injection. Maternal serum, amniotic fluid and fetal brains were preserved for analysis. NeuroTACS, a TUNEL-like staining technique specific for neuronal tissue, was performed on fetal brains (n=9 for each LPS dose). Maternal serum (n= 10 for each LPS dose) and amniotic fluid (n= 10 for each LPS dose) samples were assayed for IL-6, IL-1 and TNF- concentrations using commercially available ELISA kits. By day seven after LPS-injection, cytokine concentrations in both maternal serum and amniotic fluid were not different (p>0.05) from controls. Up to 200 g/Kg, no significant increase in apoptosis in fetal brains, compared to controls, was observed. Based on previous work in our laboratory, the threshold maternal intraperitoneal LPS dose for fetal brain injury appears to be between 200 and 500g/kg; these studies are currently being completed. Based on our previous work, the peak in cytokine concentration occurs within 48 hours of maternal intraperitoneal LPS injection. The present study demonstrates that cytokine concentrations returned to baseline levels by seven days. Most infections during pregnancy do not result in fetal brain injury until a significant level of maternal inflammatory response is attained. This supports the concept that chorioamnionitis-associated fetal brain injury may be preventable if early detection of subclinical infection and resulting delivery was achieved. (Funded by the PSI Foundation).

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DB15. Pituitary and Adrenocortical Responses to Acute Hypoxemia in Highland and Lowland Newborn Llamas Riquelme RA, Herrera EA, Sanhueza EM, Reyes VR, Ebensperger G, Ebensperger R, Parer JT, Giussani DA, Blanco CE, Hanson MA, Llanos AJ. Chronic hypoxia at high altitude blunts the neonatal cortisol response to acute hypoxemic challenge when compared to lowland newborn sheep (Riquelme et al. 2004). However, the effects of pregnancy at high altitude on ACTH and cortisol responses to acute hypoxemic challenge in highland species, such as the neonatal llamas, remain unknown. The aim of this study was to determine ACTH and cortisol plasma concentration during basal and acute hypoxemic conditions in newborn llamas, which have undergone gestation at high and at low altitude. Under general anesthesia (ketamine 10 mg/kg i.m.), 10 newborn llamas at low altitude (580 m; LA) and 6 at high altitude (3,580 m; HA) were chronically catheterized with femoral vascular catheters. Three days after surgery, the llamas were submitted to experiments based on a 3h protocol: 1h basal (B), 1h of acute hypoxemia (H; PaO2 321 mmHg) and 1h of recovery (R). Blood samples were obtained to measure blood gases and ACTH and cortisol plasma concentrations (RIA). Data are expressed as means + SEM and differences were tested using two way ANOVA for repeated measures followed by the Newman-Keuls test (*P<0.05). Basal values of ACTH and cortisol were the same in LA and HA, but there was a small but significant ACTH increase in HA newborn llamas during acute hypoxemia (p<0.05), whereas ACTH in LA did not change. In contrast, plasma cortisol concentration augmented slightly in both groups during acute hypoxemia (p<0.05). The slopes of ACTH vs. cortisol correlation were not significantly different between HA and LA newborn llamas, and this correlation was not statistically significant in HA newborn llamas. These data suggest that chronic hypoxemia during fetal and neonatal life blunts pituitary-adrenocortical response to acute hypoxemia in lowland and particularly in highland newborn llamas. These responses are similar to those reported in HA newborn sheep. The lack of a marked increase in the cortisol plasma concentration during chronic hypoxia in highland species may help to avoid premature maturation of organs and preserve the cardiovascular health in later life. Supported by FONDECYT 1050479 and The Wellcome Trust CRIG 072256. Riquelme et al (2004) J Physiol 565P PC177

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32ND FNPS 2005 POSTER ABSTRACTS

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P1. The Role of Calcitonin Gene Related Peptide (CGRP) in Fetal Life UA.S. ThakorU and D.A. Giussani Introduction: In addition to being one of the most potent vasodilators known (Brain & Grant. Physiol Rev 84:903, 2004), accumulating evidence suggest that, in adult life, CGRP activates sympathetic outflow (Hasegawa et al. Jpn J Pharmacol 61:325, 1993) and affects the physiology of the adrenal cortex and medulla (Hinson & Vinson. Neuropeptides 16:129, 1990). The distribution of CGRP throughout the human placenta (Graf et al. Placenta 17:413, 1996) and its presence in human cord and neonatal blood (Parida et al. Pediatr Res 43:276, 1998), suggest a functional role for CGRP in development. This study tested the hypothesis that CGRP plays a role in fetal cardiovascular, metabolic and endocrine regulation during basal and stimulated conditions. Methods: Under halothane anaesthesia, 10 sheep fetuses (0.8 gestation) were instrumented with vascular catheters and femoral and umbilical Transonic flow-probes. Five days later, 5 of the fetuses received 2 and 5 g bolus doses of CGRP, before and during NO blockade, in randomised order. The remaining 5 fetuses were subjected to: 1 h normoxia, 0.5 h hypoxaemia and 1 h recovery, during either saline infusion or CGRP antagonist treatment (CGRP8-37; 42.6 U+U 1.8 mg.kg-1 bolus & 8.5 U+U 0.4 mg.kg-1.min-1 infusion), on separate days, in randomised order. Treatment started 0.5 h prior to hypoxaemia and ran continuously until the end of the challenge. Arterial samples were collected for blood gases, metabolic status and hormone analyses. Results: Exogenous CGRP caused hypotension, tachycardia and vasodilatation in both the femoral and umbilical vascular beds. During NO blockade, hypotension, tachycardia and the femoral vasodilatation were all significantly diminished and the umbilical dilator response was reversed to vasoconstriction. CGRP antagonism had no effect on basal cardiovascular, metabolic or endocrine function. However, during acute hypoxaemia, the umbilical vasodilatation, the femoral vasoconstriction, and the increases in blood glucose and plasma ACTH, cortisol, catecholamines and NPY were all significantly diminished. Conclusion: CGRP is a potent NO-dependent vasodilator in the fetal femoral and umbilical vascular beds. In addition, CGRP has an important role in the fetal cardiovascular, metabolic and endocrine defence responses to acute hypoxaemia. Supported by The Journal of Experimental Pathology and The Lister Institute for Preventive Medicine. P2. Melatonin and Fetal Physiology UA.S. ThakorU and D.A. Giussani Introduction: The actions of melatonin on fetal physiology have mostly been investigated at the level of the suprachiasmatic nucleus in relation to circadian rhythmicity (Sern-Ferre et al. Ann Rev Physiol 43: 141, 2003). However, the presence of melatonin receptors in diverse fetal tissues, suggest that melatonin may be involved in a wider array of fetal functions. In this study, we have tested the hypothesis that fetal treatment with melatonin will modulate cardiovascular, metabolic and endocrine regulation in the fetus, during basal and stimulated conditions. Methods: Under halothane anaesthesia, 11 fetal sheep (0.8 gestation) were instrumented with vascular catheters and umbilical and femoral Transonic flow-probes. Five days later, 5 of the fetuses received increasing bolus doses of phenylephrine, to elicit 1-adrenergic and baroreflex responses, before and during melatonin treatment for 1 h at 0.5U+U0.1 µg.kg P

-1P.min P

-1P. The remaining 6 fetuses were subjected to: 1.5 h

normoxia, 0.5 h hypoxaemia and 1 h recovery during saline infusion or melatonin treatment (0.5U+U0.1 µg.kg P

-

1P.min P

-1P), on separate days, in randomised order. Treatment started 0.5 h prior to hypoxaemia and ran

continuously through the challenge and 15 min into recovery. Arterial samples were collected for blood gases, metabolic status and hormone analyses. Results: During basal conditions, fetal treatment with melatonin led to marked vasodilatation in the umbilical, but not the femoral, vascular bed. In addition, melatonin treatment significantly attenuated pressor and vasopressor responses to β1-adrenergic stimulation in both the femoral and umbilical vascular beds and enhanced baroreflex sensitivity. Acute hypoxaemia during saline infusion led to increases in fetal arterial blood pressure, femoral vascular resistance, blood glucose and lactate concentrations and plasma catecholamine concentrations. All these responses were significantly diminished during fetal treatment with melatonin. Despite attenuation of the increase in plasma ACTH, melatonin treatment led to a greater increase in plasma cortisol concentration during acute hypoxaemia relative to saline-infused controls. Hence, fetal treatment with melatonin markedly increased adrenocortical sensitivity to ACTH. Conclusion: Melatonin has powerful effects on cardiovascular, metabolic and endocrine functions during basal and stimulated conditions in the late gestation fetus. Supported by The Journal of Experimental Pathology and The Lister Institute for Preventive Medicine

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P3. Effects of antenatal glucocorticoid treatment on fetal development in mice UCornelle W. NoorlanderU, Pierre N.E. de Graan, Gerard H.A. Visser Synthetic glucocorticoids are frequently administered to pregnant women at risk for preterm delivery to accelerate fetal lung development. However, little is known about the adverse side effects on brain function. Actions of glucocorticoids are mediated by corticosteroid receptors, which are highly expressed in the hippocampus, a brain structure involved in cognitive functions. To determine the effects of a single antenatal dexamethasone treatment on hippocampal development, cellular, molecular, electrophysiological and behavioral analysis was performed. Here we show that antenatal dexamethasone treatment affects the number of neurons by influencing apoptosis and proliferation in the hippocampus during development. Moreover, effects on cell proliferation were observed in the dentate gyrus of the adult hippocampus. Behavioral analysis revealed that learning and working memory was impaired in dexamethasone treated mice, indicating cognitive deficits. In addition, electrophysiological experiments showed alterations in long-term depression in hippocampal slices of dexamethasone treated mice, whereas long-term potentiation was unaltered. Interestingly, histopathological analysis on heart and liver, which express high levels of corticosteroid receptors, demonstrated moderate central fibrosis in the adult liver and interstitial fibrosis in the adult heart after dexamethasone administration. Taken together, antenatal dexamethasone treatment in mice results in long-term molecular changes, cognitive deficits, impaired synaptic plasticity and histological changes in liver and heart. These data support the notion that therapeutic doses of glucocorticoids may have adverse long-term effects on human brain function. Therefore, it may be important to re-evaluate the dose required for antenatal glucocorticoid treatment for women at risk for preterm delivery. P4. Activation of H9C2 Cell Proliferation by β-Adrenergic Receptors (AR) is via Involvement of PI3K Signaling Pathway Naohiro Yano, Vlad Ianus, Ting C. Zhao, Yi-Tang Tseng, UJames F. PadburyU

BACKGROUND: We have shown that blockade of the β-Adrenergic Receptors (AR) in vivo reduces neonatal cardiomyocyte proliferation with a concomitant reduction in cardiac p70S6K1 activity. The ontogeny of PI3K-p70S6K1 signaling was examined and there appeared to be a globally high input of cardiac PI3K signaling during late gestation and in early postnatal stages of development. OBJECTIVE: The goals of the current study were 1) to examine the roles of AR on regulation of cardiomyocyte proliferation using an in vitro model and 2) to determine if PI3K signaling pathway is directly involved in this regulation. DESIGN/METHODS: H9c2 cells, a rat fetal Cardiomyocyte cell line, were treated with isoproterenol (ISO 10 M) alone, ISO + wortmannin (200 nM) or ISO + LY294002 (10 M) for 24 hours. Cell proliferation was monitored with a non-radioactive cell proliferation assay (Promega). RESULTS: ISO treatment induced a significant increase in cell proliferation index (OD490 0.2283 U+ U 0.0073 vs 0.1612 U+ U 0.0037 in control, p < 0.01). The increase was completely abrogated by co-treatment with either wortmannin or LY294002. Treatment with ISO also activated expression of phosphoactive Akt and p70S6K1. CONCLUSIONS: We conclude that activation of βAR results in an increase in cardiomyocyte proliferation in vitro via direct involvement of the PI3K signaling pathway. (Supported by NIH 1 P20 RR018728)

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P5. Influence of Porcine Genotype on the Abundance of Thyroid Hormones and Leptin in Sow Milk and its Impact on Plasma Concentrations in the Resulting Offspring A. Mostyn, J. C. Litten, K. S. Perkins, J. Laws, M. E. Symonds & L. Clarke Neonatal mortality is greater in the leaner commercial porcine genotypes compared to the ancient Meishan breed. Pigs do not express the uncoupling protein 1 responsible for heat production in most other mammals and rely on alternative strategies for thermoregulation. Thyroid hormones and leptin are potential regulators of heat production in the pig and differences in plasma triiodothyronine (T3) have been observed between the commercial and Meishan genotypes on the first day of life (1), but whether these are influenced by maternal milk supply is unknown and was therefore the focus of this study. Seven Meishan and 6 commercial sows were entered into the study and had a milk sample taken daily from the day of parturition until the forth day of postnatal life in conjunction with daily blood sampling from the median sized piglet in each litter. T3, thyroxine (T4) and leptin were measured using RIA or ELISA. Milk from Meishan sows contained significantly more T3 than milk from commercial sows (Day 1 milk, Meishan, 4.6 + 2.0; commercial, 0.89 + 0.06 ng/ml (P<0.05)) a difference not reflected in the piglet plasma T3. Sow milk T4 was similar between breeds although commercial piglets had significantly higher plasma T4 (Meishan, 55.7 + 3.0; commercial, 106.6 + 14.5ng/ml (P<0.05)). Leptin was higher in commercial sow milk throughout the study (Meishan, 9.8 + 1.3; commercial, 26.0 + 6.0 ng/ml (P<0.05)); but there was no difference in plasma leptin between piglets. In conclusion, we have found a significant disparity in the provision of thyroid hormones and leptin between Meishan and commercial sows’ milk although these differences are not translated into piglet plasma concentrations for these hormones. This work was supported by a BBSRC research grant. J.C.L. also wishes to thank Wye College for the provision of a PhD studentship. 1. Mostyn A, Litten JC, Perkins K.S, Alves-Guerra M, Pecqueur, C, Miroux B, Symonds ME, Clarke L. (2004) Journal of Endocrinology 183, 121-131 P6. Effect of maternal cold exposure on hepatic glycogen content, peroxisome proliferator-activated receptor alpha and insulin-like growth factor-II mRNA expression in the neonate. E.A.Butt, S. Pearce, H. Budge, T. Stephenson and M.E. Symonds Maternal cold exposure induced by winter shearing promotes fetal growth, particularly that of the liver by increasing maternal glucose supply to the fetus. A major component of the fetal liver is glycogen which is rapidly mobilized at birth in order to provide an endogenous energy source. Hepatic glycogen content is regulated in part by the lipid activated transcription factor, peroxisome proliferator-activated receptor alpha (PPAR), whilst insulin-like growth factor (IGF)-II regulates glycogen synthase. The aim of the present study was to determine the effect of maternal CE on hepatic glycogen content in the newborn and whether its abundance was related to PPAR or IGF-II mRNA abundance. Fifth teen multiparous ewes of similar body weight were entered into the study of which eight were shorn (S) at 70 days gestation and seven remained unshorn (US). All mothers were fed to fully meet the metabolisable energy requirements of an US sheep. The offspring were humanely euthanased at one day of age to enable liver sampling in which the relative abundance of PPAR and IGF-II mRNA to 18S rRNA were determined using RT-PCR and are expressed in arbitrary units (a.u.) whilst liver glycogen content was measured enzymatically. Despite CE offspring having larger livers (S 128+11; US 110+11 g (P<0.05)) they had a reduced glycogen content (S 67+21; US 132+11 mmol (P<0.01)) in conjunction with a decreased mRNA abundance for IGF-II (S 75+7; US 191+27 a.u. (P<0.01)) and PPAR (S 5+0.09; US 14+1a.u. (P<0.05)). In addition PPAR mRNA abundance was positively correlated (P<0.01) with hepatic glycogen content. In conclusion, maternal CE has a pronounced effect on liver mass and content which may be related to a decrease in mRNA abundance for IGF-II and PPAR that act to promote glycogen mobilization in the newborn.

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P7. Intrauterine growth restriction in the fetal lamb: effect on the size of the macula densa and the renal renin-angiotensin system Vladislava Zohdi, Karen M Moritz, Kristen Bubb, Megan L Cock, Richard Harding, UM Jane BlackU

Preliminary observations from our laboratory suggest that the maculae densae in the kidneys of intrauterine growth restricted (IUGR) fetuses are enlarged. Since, the maculae densae play a key role in the regulation of renin production, we hypothesised that the renal renin-angiotensin system is up-regulated in the IUGR fetal kidney. Hence, the aims of this study were to determine (1) whether the volume of maculae densae is increased and (2) whether the renal renin-angiotensin system is up-regulated in the fetal lamb kidney following IUGR in late gestation. IUGR was induced in fetal lambs by umbilico-placental embolization from 110-130 days of gestation (full-term 147days). At 130 days of gestation the ewes and fetuses were humanely killed and the kidneys excised. Samples of cortex and medulla from the left kidney were snap frozen and the right kidney perfusion fixed. In the fixed-kidney, total kidney volume, nephron number, renal corpuscle volume, total maculae densae volume and the volume of macula densa per glomerulus were stereologically estimated. In the frozen kidney samples the relative gene expression of the angiotensin II receptors (type 1, AT1 and type 2, AT2), renin and angiotensinogen were determined using real time PCR. Body weight, kidney volume and nephron endowment were significantly reduced in the IUGR fetuses compared to controls. Interestingly, there was a significant increase in the number of glomeruli per kidney volume in the IUGR group. There was no difference in the total volume of maculae densae between the groups. However, there was a tendency for the volume of macula densa per glomerulus to be higher in the IUGR kidneys. There was a four and five fold increase in AT1 and AT2 receptor expression, respectively in the cortex of the IUGR fetal kidneys compared to controls. Angiotensinogen mRNA levels also appeared to be up regulated in the cortex of the IUGR group. There was a significant increase in renin mRNA expression in the IUGR renal medulla. In conclusion, induction of IUGR in late gestation leads to reduced birth weight and nephron endowment but up-regulation of the renal renin-angiotensin system. P8. N-acetyl cysteine, a potential neuroprotectant, induces severe fetal hypoxemia in the presence of endotoxin. UMegan ProbynU, Megan Cock, Jhodie Duncan, Sandra Rees, Richard Harding Introduction: Maternal or intrauterine infections may induce inflammatory states leading to fetal brain injury. We have shown that bacterial endotoxin (lipopolysaccharide, LPS) delivered i.v. to the immature ovine fetus induces white matter injury. N-acetyl cysteine (NAC) is potentially neuroprotective as it blocks the production of inflammatory cytokines by scavenging the reactive oxygen species produced at the onset of an inflammatory state; however, effects of NAC on the fetus are unknown. Objective: To determine the effects of NAC on the ovine fetus exposed to LPS. Methods: Catheterised fetal sheep were entered into one of four treatment regimens repeated on a daily basis between 95 and 100d (term 147d): 1. Control (saline; n=5); 2. LPS alone (1µg/kg estimated fetal weight; n=6); 3. LPS+NAC (1 µg/kg LPS + 50-200mg/day NAC; n=8); 4. NAC alone (50-200mg/day; n=3). LPS was delivered as an i.v. bolus; NAC was delivered as an i.v. infusion over 5 hours. Fetal blood pressure and heart rate were recorded continuously and arterial blood samples collected prior to treatment and hourly during the 5 hour NAC infusion. Results: LPS alone: from 3 to 7 hours after the first exposure to LPS, SaO2 fell from 674% to 463%; controls remained at 674%. A similar response to LPS occurred on day 2 but there was little response on subsequent days. The hypoxemia was exacerbated in the LPS+NAC group, with the SaO2 falling from 711% to 382%; this effect of LPS+NAC was seen on days 1-3 but not on days 4 or 5. Fetal deaths (n=2) occurred in the LPS+NAC group. NAC alone did not alter fetal SaO2 (median 701%). Conclusion: When used in the presence of LPS in the ovine fetus, NAC (50-200mg/day) exacerbates the hypoxemia caused by LPS. The adverse physiological effects of NAC suggest that it may not be a suitable treatment for fetal inflammation.

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P9. Human Maternal-Fetal Placental Transfer of Proinflammatory Cytokines UL.K. MooreU, G.N. Smith. The chorioamnionitis-induced proinflammatory response results in elevated maternal and fetal serum, amniotic fluid and fetal brain cytokines. However, it is unknown to what extent the maternal, placental and fetal compartments each contribute to the fetal inflammatory response which is thought to directly and/or indirectly cause brain injury. It is hypothesized that maternal proinflammatory cytokines cross the placenta into the fetal circulation; these cytokines then contribute to the fetal inflammatory response leading to brain injury. The objective of this study was to examine placental transfer of maternally-administered proinflammatory cytokines using the in vitro human placental perfusion set-up. Term human placentas (n=10) were obtained from Kingston General Hospital immediately following elective caesarean section. The in vitro human placental perfusion set-up was used, as previously described in our laboratory. A dose of IL-6 (100pg/mL), TNF-α (100pg/mL) or IL-1β] (400pg/mL) was administered to the maternal arterial (MA) reservoir at time zero. Doses were determined by a review of the literature pertaining to measurement of amniotic or serum cytokine levels in mothers and fetuses with chorioamnionitis and/or periventricular leukomalacia. Samples were obtained from the maternal venous (MV) and fetal venous (FV) lines prior to the start of the experiment and for 180 minutes after the individual cytokines were administered. The samples were analyzed by ELISA to determine specific cytokine concentrations. For placentas dosed with IL-6 (n=5), MV IL-6 concentrations increased from baseline throughout the timecourse, reaching 6.6pg/mL at 135 minutes. No significant increase in IL-6 concentration from baseline was observed in FV samples. For placentas dosed with TNF-α (n=5), MV TNF- α concentrations increased from baseline throughout the timecourse, reaching 5.1 pg/mL at 150 minutes. No significant increase in TNF- α concentration from baseline was observed in FV samples. Based on the current study, it appears that TNF- α and IL-6 do not cross the human placenta into the fetal circulation. ELISAs for IL-1] are currently being run on the samples obtained. In addition, determination if these maternally administered cytokines turn on placental cytokine production is also being assessed. (Supported by a grant from The PSI Foundation). P10. Maternal parity determines the abundance of mitochondrial proteins in the lung and kidney of developing sheep D. Yakubu, A. Mostyn, T. Stephenson and UM.E. SymondsU

Maternal parity and nutrition independently affect fetal development. Previous studies have shown growth and mitochondrial protein abundance in the lung is strongly influenced by maternal diet in late gestation (1). The aim of the present study was to determine whether comparable effects are found in the kidney and the extent to which this may impact on the abundance on two important mitochondrial proteins, namely voltage-dependent anion channel (VDAC) and cytochrome c. Twenty-four twin-bearing ewes of similar body weight were entered into the study of which 14 were primiparous (P) and 10 multiparous (M). Eight P and five M ewes were nutrient restricted, consuming 50% of total metabolisable energy requirements (NR; 110d V term) whilst the remainder were fed to requirements. At 30 days of age one lamb from each ewe was humanely euthanased and tissues collected. The relative abundance of each mitochondrial protein was determined by immunoblotting with results expressed as a % reference of the same sample included in all gels. Despite there being no effect of parity on total body weight, P offspring had smaller kidneys (P 31 U+U 2; M 42 U+U 1 g (p<0.01)) and lungs (P 187 U+U 34; M 262 U+U 15 g (p<0.01)) at 30 days of age. These changes in organ weight were accompanied by a decreased abundance of VDAC (e.g. in the lung ⟨V P 60 U+U 6.4: M 79 U+U 5.4 % ref (P<0.05)) but greater content of cytochrome c in both tissues compared with offspring of M mothers. This influence of maternal parity was unaffected by maternal nutrient restriction over the final month of gestation. In conclusion, maternal parity has an organ specific effect on growth in the neonate which is paralleled by changes in mitochondrial protein abundance that may relate to alterations in metabolic activity in these tissues during postnatal development. 1. Gnanalingham MG, Mostyn A, Dandrea J, Bispham J, Symonds ME, and Stephenson T. Ontogeny and nutritional programming of uncoupling protein-2 and glucocorticoid receptor mRNA in the ovine lung. J Physiol, Lon 565.1: 159-169, 2005.

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P11. Peripheral vascular response to local cooling in pregnancy as measured by ultrasound and laser doppler UHartgill TWU, Bergersen TK, Wall E L, Pirhonen J During pregnancy profound changes occur in maternal cardiovascular physiology, most already in the first trimester. Blood volume increases by 50% with a total increase in the heart’s minute volume by 30-50%. Despite this, blood pressure falls early in the first trimester reaching its nadir in the second trimester before rising again towards term. The fall in blood pressure is due to a reduction in peripheral vascular resistance; which part of the peripheral vascular tree is affected, is unknown. The thermoneutral zone is the range of body temperature that does not trigger shivering or sweating (Lopez et al 1994), where core temperature is controlled solely by vasomotor activity in the skin. Subjects in their thermoneutral zone, show large fluctuations of arterial bloodflow to the hands and feet, caused by simultaneous opening and closing of arteriovenous anastomoses (AVA) in these areas (Thoresen et al 1980). Vasomotor activity of AVAs shows a close relationship with heart rate and mean arterial pressure variability (Lossius et al 1993). To date there is little data on the effect of human pregnancy on vasomotor activity in the skin and no data on AVAs. This study investigated the blood velocity patterns in the digital arteries using doppler ultrasound and their response to local cooling during pregnancy and for 1 year postpartum. Twenty healthy normotensive volunteers were investigated 4 times during pregnancy (weeks 8, 16, 26 and 36) and 3 times postpartum (weeks 12, 24 and 52). 12 were primipara. Average age was 31.6 years (26-36), BMI averaged 23 (19-29). The experiments were carried out in a climactic chamber, temperature adjusted to the patient’s thermoneutral zone. After 30 minutes acclimatisation, we performed local cooling (from 35P

oPC to 19P

oPC) of

the right hand in a waterbath with simultaneous blood velocity recording from both hands and laser doppler flux from the left forearm. Blood pressure and tympanic temperature were also recorded. A 3 lead ECG enabled beat for beat averaging of velocities and all data was fed online to a computer for analysis. Preliminary results will be presented for discussion

P12. Brain metabolic changes in the late gestation fetal sheep in response to umbilical cord occlusion UEdwin B. YanU, Suzanne L. Miller, Margie Castillo-Melendez, Phuong N Nguyen, Alexis A Adamides, David W. Walker Background: Neurologic injuries induced by hypoxia-ischemia accounts for a large proportion of perinatal morbidity. A previous study using microdialysis techniques showed that a delayed increase in hydroxyl radical (OH) production occurred at 8-10 h after 10 min umbilical cord occlusion (UCO). This delayed OH increase may reflect secondary metabolic changes in the brain after asphyxial insult. The aim of the current study is to determine fetal brain metabolic changes in response to asphyxia induced by UCO. Method: Pregnant sheep carrying singleton fetuses were used in this experiment. At 124-127 days of gestational age, the fetus was implanted with a brachial artery catheter for obtaining blood samples. An inflatable cuff was placed around the umbilical cord to induce asphyxia. A microdialysis probe was inserted into parasagittal grey matter for determining extracellular glucose; lactate, glycerol and pyruvate concentrations using a CMA microanalyser. Fetal asphyxia was induced by 10 min complete UCO and the cuff was not inflated in a sham group. The microdialysis probe was perfused with artificial CSF at 2 µl/min and samples collected at 30 min intervals starting 6 h before and finishing 24 h after the real or sham occlusion. Result: Glucose, lactate, pyruvate and glycerol concentrations were all increased immediately after UCO. The increase was transient for lactate and glycerol lasting 1-2 h after the occlusion. Glucose increases 2-fold and reached maximum at 4.5 h after occlusion, and did not return to the pre-occlusion levels until +10 h. Extracellular pyruvate increased from 6-7 h after UCO, and the levels remained elevated until the end of the 24 h experiment. Conclusion: Glycerol is released from cell membranes in the condition of cellular injury. The immediate increase in glycerol concentration may be indicated the immediate brain cell damage after asphyxia. The prolonged increase of glucose and the delayed increase of pyruvate in extracellular fluid indicated prolonged disruption of normal cellular metabolism.

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P13. The sensitising effects of inflammation on hypoxic-ischaemic brain injury G Kendall, A Clements, H Varley, G Raivich, D Peebles The fetal brain is protected from the effects of hypoxia-ischaemia (HI) by haemodynamic and metabolic compensatory mechanisms, so that pure HI is a relatively unusual cause of perinatal brain injury. A growing body of epidemiological and experimental animal data suggests that infection may sensitise the developing brain to mild HI1, reducing the threshold at which HI leads to brain injury. The aims of this study were to investigate the interaction between a bacterial product (endotoxin) mediated inflammation, and hypoxia-ischaemia on neuronal cell death and glial activation in a mouse model of neonatal HI. Animal procedures were approved by the Home Office, and were carried out according to the UK Animals (Scientific Procedures) Act 1986. Experiments were performed on C57/Bl6 mice at postnatal day 7. An inflammatory reaction was provoked by i.p. injection of 0.3mg/g endotoxin (LPS) from E.coli, 055:B5. Mild HI was induced by left-sided carotid occlusion under isoflurane anaesthesia followed by exposure to 8% oxygen for 30 minutes. These insults were performed independently and in combination. Histological brain injury after HI was assessed at 48 hours by measuring infarct volume (expressed as a percentage of the contralateral hemisphere). The effect of LPS was assessed after 12 and 48h using histology, TUNEL and immunohistochemistry for microglial (M 2-integrin) & astrocyte (GFAP) activation. Mild HI resulted in minimal histological brain jury with neuronal loss confined to the cortex and hippocampus. LPS alone (0.3mg/g) without subsequent HI did not result in any cell death (Nissl, TUNEL). LPS alone did, however, activate microglia in all areas of the forebrain at 12h following injection (F1,25=65.6 p<0.01); this activation was still present at 48h. LPS alone did not result in astrocyte activation (GFAP) over controls at 12h, but there was increased GFAP immunoreactivity by 48h. LPS given 4h before HI resulted in an overall increase in brain injury compared with animals exposed to HI alone (F3,19=6.37, p<0.01 n=10). This sensitising effect was evident when LPS was given within a window of 4-12 hours before the onset of HI. No significant sensitisation was observed when LPS was given immediately or 24 hours prior to the onset of HI. These data support the hypothesis that inflammation sensitises the brain to subsequent HI; further, the magnitude of this interaction depends on the time interval between exposure to LPS and HI. Interestingly, endotoxin alone already resulted in microglial activation within the critical 12 hour period. Activation of the innate immune system by LPS may therefore be an important mediating factor in the synergistic interaction between LPS and HI2. In comparison, astrocyte activation only occurs relatively late and may be secondary to microglial activation, following infectious and/or hypoxic stimuli. 1. Peebles DM, Wyatt JS. BJOG. 2002 Jul;109(7):737-9. 2. Lehnardt S et al. Proc Natl Acad Sci. 2003 Jul 8;100(14):8514-9. P14. Expression of leptin and the leptin receptor in the ovine placenta D. M. O’Connor, E. Brookes, F.B.P. Wooding, N. Hoggard, A.L. Fowden and A.J. Forhead. Leptin is a hormone produced by adipose tissue. The leptin receptor (Ob-R) has five isoforms with the long form being the main signal transduction form of the receptor. The placenta has been found to express the leptin receptor protein and therefore is a possible site of leptin action. The placenta as a source of leptin possibly signalling either or both the mother and the fetus has been investigated. Localisation of leptin protein and mRNA has been demonstrated in the mouse placenta [1]. Leptin and the leptin receptor have been detected in the ovine placenta [2]. However the localisation and abundance of both proteins and mRNAs for leptin and leptin receptor has not been reported previously in the ovine placenta. Placentomes were collected from ewes at two gestational ages 130 (n=7) and 145 (n=5) days of gestation, after maternal administration of a lethal dose of barbiturate (term 1452d). Immunohistochemistry was carried out on paraffin-embedded sections with primary anti-bodies raised against leptin (1:1000) and Ob-Rb (1:100). Leptin and Ob-Rb localisation were detected using a 4nm gold-labelled secondary antibody with silver enhancement. In situ hybridisation was carried on frozen sections using DIG-labelled oligonucleotide probes detecting leptin and leptin receptor mRNA. Leptin and leptin receptor proteins were detected in placentomes at both gestational ages studied. Both proteins co-localised to the interdigiting trophoblast layer in the placentome. No differences in distribution or abundance of either protein were observed with gestational age. Expression of leptin and leptin receptor mRNA was detected in the ovine placenta at both ages. Leptin mRNA was localised in the fetal trophectoderm and in the blood vessels. Leptin receptor mRNA co-localised with leptin but its expression appeared more widespread within the placentome. No differences were observed in abundance of either leptin or the leptin receptor mRNA with gestational age. These results demonstrate that the ovine placenta is both a source and a target for the actions of leptin. This raises the possibility that ovine placental leptin may signal to either or both the mother and fetus. Supported by the BBSRC. 1. Hoggard et al., 1997 2. Thomas et al., 2001

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P15. Antenatal Glucocorticoids Suppress Cytochrome C Oxidase Activity in the Fetal Ovine Brain M. Schwab, G. Wichmann, M. Loehle, P.W. Nathanielsz, I. Maurer, O.W. Witte Synthetic glucocorticoids administered to accelerate fetal lung maturation in pregnant women who threaten preterm delivery decrease metabolism-controlled cerebral blood flow, electrocortical brain activity and the brain expression of high-turnover neuroskeletal and synaptic proteins. We hypothesized that these changes are at least in part mediated by catabolic glucocorticoid effects. Previously, we have shown in fetal sheep that glucose uptake across the blood brain barrier and the neuronal membrane is not affected. In the present study, we examined glucocorticoid effects on the respiratory chain, the major neuronal pathway of energy generation. At 112 days gestation (0.73 of gestation), pregnant sheep were treated I.M. with one course of betamethasone phosphate consisting of two injections betamethasone 24 h apart at doses of 170 (n=8), 85 (n=8) and 42,5 gkg-1 (n=7) or saline (n=7). The betamethasone doses are equivalent to 12, 6 or 4 mg betamethasone, respectively, administered to a 70 kg woman. 24 h after the second injection, we determined spectrophotometrically the activities of mitochondrial respiratory chain enzymes complex I+III, complex II+III, complex IV (cytochrome c oxidase, COX), succinate dehydrogenase, and citrate synthase in brain specimens of the parietal cortex, basal ganglia, thalamus and hippocampus. Activities were expressed per non-collagen protein content and corrected for citrate synthase activity, i.e. for number of mitochondria. Betamethasone treatment reduced COX activity in the parietal cortex (p<0.05) independently of the betamethasone concentration. We could not evaluate the effect of betamethasone on COX activity in other brain regions examined probably due to the lower content of neurons in these specimens. Moreover, betamethasone did not have effects on the other enzyme activities studied in any brain region investigated. In conclusion, the clinical dose of betamethasone is supramaximal with respect to the specific inhibition of the COX which may lead to impaired energy generation in the fetal brain. We hypothesize that the inhibition of the energy generation contributes to the steroid effects on the fetal brain. We suggest that risk-benefit studies involving the use of even lower doses are needed to evaluate both the beneficial and multisystem unwanted effects of antenatal steroid therapy. P16. Spiny Mouse – A Model for Perinatal Research Hayley Dickinson, David W Walker The spiny mouse (Acomys caharinus) is native to the Mediterranean region where it inhabits desert and rocky terrains. The relatively long gestation (40 days), small litter size (1-4) and precocial pups suggest this species would be appropriate for many questions currently considered in perinatal research. In a colony established at Monash University we have made the following observations: Adult weight is 35-45 g, resting mean arterial pressure is 76 mm Hg, heart rate is 400 beats/min, 24 h food and water consumption is 2.6 + 0.3 g and 3.9 + 0.5 ml respectively; Metanephrogenesis begins at approx. day 18 of gestation and is completed before birth. The adult spiny mouse has 7245280 nephrons per kidney, a glomerular filtration rate [under anaesthesia] of ~135ul/min, and produces 1.75 + 0.27 ml urine/24 h which is hyperosmotic (1599 + 177mOsm/kgH2O) compared to urine of laboratory mice (841 + 41mOsm/kgH2O). Female spiny mice enter post-partum oestrus within 24 h of parturition and will conceive again during this time. Pre-term labour can be induced by inflammatory stimulation from at least day 30 - delivery of non-viable fetuses occurred 12 and 20 h after 2 and 0.25 mg/kg LPS (i.p), whereas pregnancy was maintained after 0.125 mg/kg LPS i.p. Birth weight of the spiny mouse is 5.6 + 0.07 g, and term placental weight is 0.53 + 0.05 g. The placenta increases in weight throughout pregnancy, but the inner labyrinthine region increases relative to the junctional zone from day 25, correlating with accelerated fetal growth from approx day 30 of gestation. The brain is incompletely myelinated at birth, and mature (CNPase positive) oligodendrocytes are not present until after postnatal day 10. Brain weight increases from approx 0.06 g on day 18 to 0.475 + 0.009 g at term, with the brain:body weight ratio decreasing from 0.502 + 0.017 to 0.084 + 0.005 over this time. Neonates gain postural tone within an hour of birth, and may be tested behaviourally for motor coordination, locomotion, balance, and open field exploratory behaviour within the first 24 hours. The incomplete development of white matter in the brain at birth suggests the Spiny Mouse is comparable to the human in this respect, and less developed than the sheep and guinea pig where substantial myelination has occurred by the time of birth.

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P17. Gonadotrophic hormones reduce pregnancy rates and perturb fetal development due to changes in the maternal uterine and hormonal environment in mice RL Kelley, KL Kind, M Lane, RL Robker, JG Thompson, LJ Edwards Despite its routine use in clinical IVF, the adverse effects of gonadotrophin stimulation using recombinant human follicle stimulating hormone (rhFSH) are largely unknown. Evidence from clinical studies indicates that rhFSH stimulation may be detrimental to the endometrium and implantation. We have previously shown in mice that stimulation with rhFSH or equine chorionic gonadotrophin (eCG) reduces embryo quality and developmental potential, and also alters maternal preimplantation progesterone levels and uterine expression of leukemia inhibitory factor (LIF) mRNA and progesterone receptor (PR) protein, both of which are markers of endometrial receptivity. In this study we investigated the effect of gonadotrophin stimulation on pregnancy outcomes in mice. Adult female mice were stimulated with 10 IU rhFSH (n=23) or 5 IU eCG (n=19), followed by 5 IU human chorionic gonadotrophin (hCG). Control mice (n=16) received saline injections. Mice were mated with vasectomised males to induce pseudopregnancy. Embryos collected from stimulated pre-pubertal donors and cultured to blastocyst stage were transferred to the uterus of pseudopregnant mice on day 4 of pregnancy. Mothers were sacrificed on day 15 to assess pregnancy outcomes. Mice administered eCG or rhFSH had lower pregnancy rates compared to controls (11, 35 and 75 % respectively), but litter size was unaffected by treatment. Stimulation with rhFSH reduced mean fetal weight (94 + 6 vs 176 + 8 mg), length (11 + 0.2 vs 12 + 0.1 mm) and maturity (14.1 + 0.09 vs 14.6 + 0.05 days) compared to controls. Placental weight was unaffected by stimulation. Consequently the fetal: placental weight ratio was reduced compared with controls (1.1 + 0.2 vs 2.3 + 0.1). This study shows that gonadotrophins can cause acute changes in the maternal preimplantation hormonal and uterine environment and that these may contribute to the longer-term defects in establishment of pregnancy and fetal development reported here. P18. IGF-II and IGF2R Expression in Early Murine Implantation Sites: A Role in Both Placentation and Decidual Angiogenesis? Pringle KG & Roberts CT The highly invasive activity of the human placenta is tightly regulated by a variety of growth factors and other molecules. Insulin-like growth factor-II (IGF-II) is widely expressed in early human pregnancy and is critical for optimal fetal and placental growth. Important roles for IGF-II in murine placental transport and growth have been shown by IGF-II gene ablation. However to date, IGF-II and its specific receptor, the type II IGF receptor (IGF2R) have not yet been localised to early murine implantation sites when trophoblasts are most invasive. This study provides a photo micrographic account of early placental development and the decidual vasculature in the mouse, and localises IGF-II and the IGF2R from days 5.5 to 10.5 of pregnancy. During early pregnancy, the decidua undergoes extensive angiogenesis and remodelling, displaying a paucity of blood vessels, so that by day 10.5 the decidua has become a highly vascularized structure, with an extensive network of dilated endothelial lined vessels that contain no smooth muscle and presumably enable maximal blood supply to the placenta. Throughout these early stages of pregnancy, there is no evidence of individual trophoblasts migrating into the maternal tissue, as is the case in humans, suggesting that murine trophoblasts are not directly involved in the remodelling of the maternal vasculature. However, it is possible that the trophoblasts play an indirect role through the secretion of an array of molecules that may promote angiogenesis in this region. IGF-II and its receptor were present throughout early pregnancy in the embryo and the trophoblasts, supporting their roles as regulators of fetal and placental development. Most interesting however, was their association with the developing maternal blood vessels in the mesometrial, but not the antimesometrial, decidua. It seems likely then, that in mice IGF-II and IGF2R may have two very distinct roles in early murine pregnancy; the first to promote the growth of the developing fetus and placenta, and the second to promote decidual angiogenesis.

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P19. Transient NMDA receptor-mediated hypoperfusion following umbilical cord occlusion in preterm fetal sheep Justin M Dean, Alistair J Gunn, Guido Wassink, Sherly George, Laura Bennet Exposure to severe hypoxia commonly leads to the delayed appearance of cerebral and peripheral hypoperfusion. There is evidence that transient abnormal glutaminergic receptor activity can occur during this phase of recovery. We therefore examined the role of the NMDA receptor in mediating this secondary hypoperfusion. Preterm fetal sheep at 70% of gestation received either sham-asphyxia or asphyxia and were studied for 3 days recovery. Dizocilpine maleate (2mg.kg-1 bolus plus 0.07mg.kg.h-1 i.v.) or saline was infused from 15min post-asphyxia until 4h. In the asphyxia-vehicle group abnormal EEG spiking activity was observed during the first 4h of reperfusion, the peak of which corresponded with the nadir in peripheral and cerebral hypoperfusion. Dizocilpine significantly suppressed this activity (2713 versus 11227 counts.10 min-1 at peak frequency, P<0.05) and markedly delayed and attenuated the rise in vascular resistance in both peripheral and cerebral vascular beds observed after asphyxia, effectively preventing the initial deep period of hypoperfusion maximal at 1 h (CBF: 29.5 + 2.5 versus 16.0 + 3.0 ml.min-1, P<0.01; FBF: 8.8 + 1.9 versus 3.0 + 1.1 ml.min-1, P<0.01). However, while continued infusion did attenuate subsequent transient tachycardia, it did not prevent the development of a secondary phase of persistent but less profound hypoperfusion. In conclusion, the present studies suggest that the initial phase of delayed cerebral and peripheral hypoperfusion following exposure to severe hypoxia is mediated by NMDA receptor activity. The timing of this effect in the cerebral circulation corresponds closely with the abnormal EEG activity, suggesting a pathological glutaminergic activation that we speculate is related to evolving brain injury.

P20. Changes in force of contraction of the papillary muscle with increasing gestational age in the sheep fetus T. Spencer, J.L. Morrison, G. Posterino Sheep fetal hearts, after 110 days gestation, begin binucleation of cardiomyoyctes, which then increase in size across late gestation. The force of contraction generated by papillary muscle of the heart may be dependent on the degree of binucleation. This study aims to investigate the force of contraction of the papillary muscle at three times in late gestation and to understand the impact of fetal oxygenation on these forces. Reduced fetal oxygenation in test sheep will be established by carunclectomy surgery performed on ewes prior to conception. Comparisons will be made with age-matched fetal control sheep. Fetal vascular surgery has been performed on control fetal sheep at 110-118 days gestation to allow measurement of fetal blood gases and post mortem performed at 126 (n=3), 131 (n=5) and 139 (n=3) days gestation. Fetal hearts were dissected, weighed and reverse perfused through the aorta with Tyrodes buffer. The right ventricle was incised for the removal of papillary muscle samples. Under microdissection, small bundles of cardiomyocytes of diameter 100-300µm and length ~1.5mm were isolated and attached to a force-transducer. Force responses were recorded via Powerlab/8Sp (ADInstruments, Castle Hill, Australia) onto a chart recorder (Ross Recorders, USA) and Powerlab Chart v4.2.1 software (ADInstruments, Castle Hill, Australia). All surface and compartmental membranes were permeablised using a diluted Trinton-X 100 solution (Sigma Chemicals, Australia), leaving the contractile apparatus intact, then to be exposed to solutions of increasing free calcium concentrations (pCa). Force responses were normalised against the cross-sectional area of the cardiomyocyte bundle and as a force required to produce 20% (EC20), 50% (EC50) and 80% (EC80) of maximum force (pCa 4.2). The data demonstrate a decrease in the sensitivity of the contractile apparatus to free calcium as the gestational age of the fetus increases from 126 to 140 days. The EC50 value decreases from a pCa of 6.22±0.03 at 126-127 days gestation to 6.11±0.03 at 130-132 days gestation. The expected relative force and pCa relationship was established for the control fetal hearts, in preparation for comparison with fetal hearts developed under placental restriction.

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P21. Classification of FHR Patterns according to risk of acidemia and evolution to higher risk patterns J.T. Parer and Tomoaki Ikeda OBJECTIVES: Numerous national and international bodies have attempted development of guidelines for FHR pattern management; none has yet been demonstrated to be fully successful. Two major problems have been agreement about patterns signifying fetal acidemia and management of the evolution of milder patterns to those with a higher risk of acidemia. Our aim is to provide insight into these 2 aspects using the best data available. M&M: We have identified 134 FHR patterns based on baseline rate, baseline variability and type of deceleration. Based on the best available evidence from the literature (Parer et al, WCPM, Osaka, 2003) we have assigned a risk of fetal or newborn acidemia, or low 5 min Apgar scores, to these patterns and coded them in 5 levels of risk, from I (no intervention required) to V (immediate intervention). In addition, we have assigned a risk of these patterns evolving to those patterns with a higher risk of acidemia, based on further observations from the literature. Table: Risk of acidemia (I-V), and risk of evolution to higher risk acidemia patterns in 1 hour period (Low, moderate, high) FHR pattern: Risk of Acidemia Risk of evolution Nl rate, Nl FHRV 0 decels,early decels, mild VD I Low

Mod VD, Mild LD,Mod LD, Mild PD I Mod Sev VD,Sev LD, Sev PD I High

Nl rate, Min FHRV 0 decels, early decels, mild VD II(or I) Unk/mod Mod VD, sev VD III Mod Mild LD III High Mod LD IV High Sev LD V High

Nl rate and 0 decels IV/less Unk Undetect FHRV All others V High * VD, variable decels; LD, Late decels; PD, prolonged decels; FHRV, FHR variability DISCUSSION: This classification, based on uncontrolled observational studies, can be used as a basis for confirmation of its utility by prospectively gathered data. We have omitted several patterns, e.g., sinusoidal, tachycardia, for want of sufficient data. Bradycardia is in general similar to PD. Meanwhile, this schema can be tentatively used for imposing a hierarchy of conservative interventions, or for delivery, and for determining patient acuity in a unit. P22. Relation between ECG change and alteration of low frequency domain in the spectral analysis of R-R intervals in the progress of fetal acidemia. Kimura Y, Sugawara J, Imai N, Chisaka H, Okamura K Objectives: As one of the adjunctive fetal monitoring during labor, fetal ECG waveform including T/QRS ratio has been applied in many countries. T/QRS ratio varies with the activity of sympathetic receptor in the ventricle. On the other hand, low frequency (LF) domain, around 0.1Hz, in spectral analysis of heart beat variability has been considered for years as an indicator of cardiac sympathetic nerve activity. We investigated the relation between T/QRS and cardiac sympathetic nerve activity reflected in the change of LF in the fetal sheep acidemia model. Materials and Methods: Femoral artery was cannulated, an occluder was placed around the umbilical cord and ECG electrodes were stitched in five sheep fetuses. Gradual fetal acidemia was induced by repetitive umbilical cord compression. The change of fetal ECG was expressed as P wave amplitude measured from the baseline and T wave amplitude measured from the starting point of Q wave to the T wave peak. Inversed T was counted ?g minus?h. A wavelet transform was used to evaluate temporal change of the power of LF during cord compression. Results: In fetal heart rate deceleration caused by cord compression, both of T/QRS ratio and LF increased when arterial pH > 7,20. When pH<7.20, both of them decreased together, and inversed P wave remained even after the release of cord compression. Discussion: Autonomic nervous control in cardiac ventricle is mainly conducted by sympathetic nerve. Increase of T wave in ischemia should be an expression of compensation of fetal heart to hypoxic stress. Inversion of T wave emerges at the collapse of the compensation mechanism. To evaluate LF may lead us to know the change of cardiac sympathetic activity due to ischemia which could be observed in T/QRS change.

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P23. Circadian Rhythm in Breathing Characteristics in Fetal Lambs Marta Wlodarska, John Challis, Stephen Lye, William Gibb, Nancy Gruber, Dan Rurak Fetal breathing movements (FBM) in sheep exhibit a circadian rhythm, with an increase in breathing in the evening. This has been linked with the maternal circadian rhythm in plasma melatonin. However, there are limited data on FBM characteristics in terms of rate and depth, largely because of the difficulty in analyzing the FBM over long periods of time. The objective of the current study was to use PowerLab data acquisition software to analyze FBM rate and depth on-line from chronically instrumented fetal lambs (n=6) at 126.5 + 1.0 d gestation. Fetal arterial, amniotic and tracheal pressures were continuously recorded using PowerLab 5.2 (ADInstruments). Fetal heart rate and FBM rate and depth were determined over 15 sec epochs using the cyclic variables function for the arterial and tracheal-amniotic pressure traces, respectively. The data were analyzed over 48 h periods, 4-5 days after surgery and before any experimental procedures. There was a clear circadian rhythm in FBM rate, with a trough (39.5 + 3.5/min) at midnight and a peak (62.5 + 7.5/min) at 1500, with a similar trend for FBM depth. To examine the FBM pattern more closely, the FBM episodes closest to the trough (00:39 0:04) and peak (15:21 0:04) of the circadian rhythm in each animal were analyzed in detail, with the duration of each episode being normalized. In episodes occurring at the peak, rate increased significantly from 47.2 + 1.8 to 80.6 + 1.2 /min after 32% of the episode had elapsed. FBM depth also increased significantly (3.4 + 0.3 to 5.0 + 0.2 mm Hg) at the same time. In contrast, there were no consistent changes in FBM rate or depth during the episodes that occurred at the circadian rhythm trough. It is concluded that there is a circadian rhythm in FBM rate and depth, whose timing is different from the rhythm in FBM incidence. Moreover, the rate and depth rhythm is associated with consistent changes in the rate and depth characteristics within episodes. The cause of this rhythm is not yet apparent, but it may relate to the progressive rise in fetal blood glucose concentration (peaking at 1300) which occurs with the feeding regimen we employ with our sheep. P24. Temporal antenatal testing alterations in severe IUGR fetuses EV Cosmi, G Mari and E Cosmi Objective: to assess the longitudinal changes in Doppler velocimetry, FHR tracing and BPP in severe IUGR fetuses. Methods: A prospective longitudinal study of 286 IUGR fetuses. In all fetuses a daily FHR tracing, Doppler velocimetry twice weekly has been performed to assess the umbilical artery (UA), middle cerebral artery (MCA), ductus venosus (DV) and umbilical vein (UV) blood flow. The BPP and amniotic fluid index were evaluated twice weekly. Results: of 286 fetuses enrolled in the study, 86 did not show Doppler alteration and were not considered into the study. Gestational age at time of admission ranged from 19 to 39 weeks of gestation. 200 IUGR fetuses showed Doppler velocimetry alteration after the enrolment. In all fetuses antenatal testing alterations consisted in the following temporal sequence: increased resistance of the UA (100%); UA AEDF (52%), UA RF (26%); brain sparing effect in the MCA (38%); abnormality in the DV (absent or reversed flow during atrial contraction) and UV pulsations (20%). BPP and FHR tracing was abnormal in 30% of the fetuses and in all of them these changes occurred after Doppler velocimetry alterations with a mean time of 3.8 days. AFI was below the normal ranges in 42% of fetuses. Conclusions: severe IUGR fetuses show a specific temporal sequence antenatal testing alterations consisting of Doppler velocimetry alterations, while FHR tracing become abnormal too late, and are the last changes seen in IUGR fetuses as well as the BPP and AFI.

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P25. The Pituitary-Adrenal Axis In The Lamb Is Re-set Following Placental Restriction In Utero S.L. Dunn, J.A.Duffield, I.C. McMillen and C.L. Coulter It has been proposed that growth restriction in utero results in a reprogramming of the fetal hypothalamic-pituitary-adrenal axis and hypercortisolism in postnatal life. Previously, we have shown that restriction of placental growth (PR) and hence fetal growth, results in a relative increase in adrenal weight, increased adrenal expression of the steroidogenic enzyme cytochrome P450 side-chain cleavage (CYP11A1) and elevated fetal plasma cortisol concentrations (Phillips et al, 1996; Ross et al 2000). The aim of this study was to determine if the effects of PR in utero on the pituitary-adrenal axis were persistent in the early postnatal period. We investigated the effects of PR on adrenal steroidogenic capacity and basal plasma cortisol concentrations in lambs at 21 days after birth (21d). Normally grown (n=10) and PR (n=6) lambs were killed at 21d, adrenals collected and frozen for determination of expression of steroidogenic Acute Regulatory Protein (StAR), melanocortin type-2 receptor (MC2R), CYP11A1 and CYP17MrNA. Placental restriction resulted in reduced birth weight (3.84 + 0.19kg) compared to normally grown lambs (5.4 + 0.09kg), while relative adrenal weights were increased in PR lambs (Control = 81.31 + 4.52, PR = 111.41 + 6.74mg/kg). There was no significant difference in the expression of CYP11A1 or CYP17 in the adrenal glands from normally growth lambs or growth-restricted lambs at 21days after birth (CYP11A1: Control = 35.8 + 2.4, PR = 32.0 + 2.8; CYP17; Control = 76.7 + 17.4, PR = 64.6 + 10.4). Similarly there were no differences between treatments for MC2R (Control =61.4 + 6.9, PR = 86.3 + 19.0) or StAR (Control = 94.2 + 8.8, PR = 99.9 + 19.6). In addition, basal plasma cortisol concentrations were not significantly different between normally grown and PR lambs (Control =23.16 + 4.94nmol/L; PR=37.31 + 11.34nmol/L). In summary, whilst relative adrenal growth remains increased in the post-natal lamb following fetal growth restriction, there has been a normalisation of adrenocortical synthetic capacity and basal cortisol concentrations. P26. Changes observed in Doppler studies of the fetal circulation in pregnancies complicated by pre-eclampsia or intrauterine growth restriction. Nayana Parange, Gustaaf Dekker, Chris Wilkinson. Objectives: To compare changes in Doppler ultrasound studies of the fetal circulation in normal pregnancies with a group of pregnancies complicated by preeclampsia or intrauterine growth restriction. Study Design: Ongoing Prospective longitudinal observational study. Methods: The PI of fetal ductus arteriosus, PI of foramen ovale, PIV of ductus venosus and RI of Middle cerebral artery, and Umbilical artery and maternal uterine arteries was measured by Doppler ultrasonography, from 16 weeks onwards till delivery in high risk pregnancies as well as controls, and outcomes compared. Results: There were serial haemodynamic dynamic changes throughout pregnancy in the PI of Foramen ovale and Ductus arteriosus, in addition to the changes in Ductus Venosus, Umbilical artery and Middle cerebral artery. The trends of sequence of haemodynamic changes observed in adverse outcomes appeared to be different to the changes seen in normal fetuses. The changing trends in the pulsatility of Ductus Arteriosus suggested that these changes appeared early in the disease, versus the trends seen in pulsatility in umbilical artery and ductus venosus which appeared much later. Conclusions: The human fetal cerebral response to intrauterine growth restriction is being investigated. However, the exact sequence of haemodynamics changes occurring in fetal circulation, whereby the fetal circulation switches from “physiological adaptation” to “decompensation”, in the presence of stress, is still not clear. Our study suggests that the intrauterine cardiac shunts may play an important role in redistribution of fetal flows before growth restriction sets in and the mechanism of fetal adaptation to stress may be more complex than previously thought.

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P27. Ovine Placental Igf2 and Igf2r Gene Expression is Reduced by Repeated Antenatal Betamethasone Treatment Roberts CT, Moss TJ, Button JJ, Nitsos I, Harding R, Newnham JP Maternal treatment with antenatal glucocorticoids improves neonatal outcome following preterm birth. In sheep, both maternal and fetal intramuscular betamethasone injections improve preterm lung function. However, only maternal treatment causes fetal growth restriction suggesting impaired placental function. We aimed to quantify placental Igf2, Igf2r and Glut-1MrNA expression in sheep following single or 3 repeated maternal (M1 and M3) or fetal (F1 and F3) intramuscular injections of betamethasone (1-beta and 3-beta). Methods: Forty-one ewes were injected with 150mg medroxy-progesterone acetate on day 98 of pregnancy (term is 150 days) and randomised to maternal (MS) or fetal saline (FS) (104, 111, 118 days) or 1-beta (104 days, followed by saline at 111, 118 days) or 3-beta (104, 111, 118 days) treatment groups. At 146 days, ewes were killed and fetuses weighed. All placentomes were weighed and snap frozen. To determine if betamethasone treatments caused sustained changes in placental gene expression, RNA was extracted from B and C type placentomes from MS, M3, FS and F3 groups. Gene expression was quantified by real time RT-PCR. Results: We have previously reported that only M3 reduced fetal weight (22%, p=0.038) but total placental weight was unaffected. Nevertheless, fetal and placental weights were positively correlated (r=0.722, P<0.0005). Placental Igf2 and Igf2r mRNA were reduced by 35% and 26%, respectively in M3 compared to controls (MS, p<0.05). Both genes were expressed more abundantly in placentas from fetal compared to maternal treatment groups (P<0.03). Placental Glut-1 expression was not altered by treatment. Placental igf2 expression was correlated with that of igf2r (r=0.634, P=0.002) and Glut1 (r=0.622, P=0.002). Placental igf2r expression was correlated with fetal weight (r=0.542, P=0.009). Conclusion: These results suggest that repeated maternal betamethasone treatment causes sustained reductions in fetal growth and in placental expression of genes involved in tissue remodelling (igf2 and igf2r) and placental transport and growth (igf2). Acute reductions in IGF-II expression following glucocorticoid treatment have previously been reported in a range of other tissues and cell lines. This is the first report of glucocorticoid inhibition of igf2 transcription sustained nearly one month following cessation of treatment and therefore has significant implications for repeated glucocorticoid treatment in human pregnancy. P28. Accelerated Apoptosis and Differentiation of Trophoblasts Isolated from Intrauterine Growth Restricted Pregnancies Sarah Newhouse, Gary Chan, Sandra Davidge, Larry Guilbert. Intrauterine growth restriction (IUGR) is a serious pregnancy complication in which the fetus fails to reach its full growth potential, possibly leading to a number of health problems during child and adult life. Preeclampsia, hypertension accompanied by proteinuria during the latter half of pregnancy, is often associated with IUGR. The villous placenta is bathed in maternal blood, thus is essential for ensuring proper fetal development. Specifically, trophoblasts are responsible for the exchange of oxygen and nutrients between the maternal and fetal circulation. However, very little is known about cell turnover and differentiation in trophoblasts isolated from IUGR pregnancies (IUGR) or from IUGR pregnancies complicated with preeclampsia (IUGR-PE). In agreement with other studies, we show that trophoblasts isolated from IUGR pregnancies undergo higher rates of apoptosis than normal trophoblasts. Next, we examined trophoblast differentiation using human chorionic gonadotropin (hCG) expression, placental alkaline phosphatase (PLAP) surface expression, and multinucleation. Trophoblasts isolated from IUGR pregnancies express higher levels of hCG, PLAP, and spontaneous syncytialization when compared to normal age-matched control trophoblasts. Others have shown p38 mitogen-associated protein kinase (MAPK) to be involved in trophoblast syncytialization and differentiation. We find that trophoblasts isolated from IUGR pregnancies express higher levels of phosphorylated p38 MAPK than normal trophoblasts. Inhibition of p38 phosphorylation prevents spontaneous cell fusion in both IUGR and normal trophoblasts. Furthermore, trophoblasts isolated from IUGR-PE pregnancies show similar rates of syncytialization and hCG and PLAP expression to normal. Our results indicate that trophoblasts isolated from IUGR pregnancies, in contrast to those isolated from IUGR-PE pregnancies, have increased rates of trophoblast turnover and differentiation, possibly leading to decreased placental function.

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P29. Hypoxic/Ischemic models in newborn piglet: a constant versus variable approach S. Tracey Bjorkman, Kelley A. Foster, Stephanie M. O’Driscoll, Genevieve N. Healy, Barbara E. Lingwood and Paul B. Colditz A severe hypoxic/ischemic insult to the developing brain can lead to permanent neurological impairment and in some cases result in infant death. In Australia, approximately 3.5% of perinatal deaths can be attributed to asphyxia annually {Laws, 2004 #9}. Current clinical practice fails to detect the occurrence of hypoxia antepartum, the period in which hypoxic/ischemic neuropathology has been shown to primarily occur {Low, 2004 #20}. Rapid identification and intervention of asphyxia during and following birth is crucial for reducing the risk of lifelong disability or death. The numerous biochemical cascades through which neuronal cell death occurs offers the potential for minimizing neuropathological damage due to the many different points in these pathways where cellular injury can be interrupted or in some cases reversed. A comparison of a constant and variable hypoxic/ischemic insult was performed to determine which insult is more effective in producing a consistent degree of survivable neuropathological damage in a newborn piglet model of perinatal asphyxia and, to identify which physiological responses may contribute to obtaining this ‘desirable outcome’. Fifty-one day old piglets were subjected to either a constant hypoxic/ischemic insult (FiO2 4%; n = 23) of 30-37 min duration or a variable hypoxic/ischemic insult (FiO2 2-10%; n = 16) of 30 min low amplitude cerebral function (LAEEG) including 10 min of low mean arterial blood pressure (MABP; <70% of baseline). Control animals were subjected to FiO2 21% (n = 12). At 72 h the piglets were euthanased, their brains removed and fixed in 4% paraformaldehyde and assessed for hypoxic/ischemic injury by histological analysis. Based on histology scores, piglets were grouped as undamaged or damaged; piglets that did not survive to 72 h were grouped separately as dead. The variable insult resulted in a greater number of piglets with neuropathological damage (undamaged = 12.5%, damaged = 68.75%, dead = 18.75%) while the constant insult resulted in a large proportion of undamaged piglets (undamaged = 50%, damaged = 22.2%, dead = 27.8%). Our results suggest that for a reliable degree of hypoxic/ischemic induced damage; a variable approach is the recommended paradigm. P30. Adenosine Receptor Modulation Effects on Behavioural State Activity and Cerebral Blood Flow in the Ovine Fetus Near Term Neesha Rao, Shannon Hemstreet, Brad Matushewski and Bryan Richardson Objective: In the ovine fetus near term, one hour infusion of an adenosine (Ado) receptor (R) agonist enhances high-voltage (HV)/non rapid movement (NREM) behavioural activity, while an Ado R antagonist increases low-voltage (LV)/REM behavioural activity, implicating Ado as a sleep-promoting agent, as in adults. This study investigated longer term effects of Ado R modulation on behavioural state and cerebral blood flow velocity (CBFv). Chronically catheterized fetal sheep were studied at ~0.9 of gestation for a 2 hour control period and 6 hour experimental period: Control group (N=10) intra-arterial (IA) saline. CPA (Ado A1R agonist) group (N=9) IA N(6)-cyclopentyladenosine (CPA);0.008 mg/min/kg estimated fetal weight (EFW). ZM (Ado A2R antagonist) group (N=5) IA ZM-241385 (ZM); 0.056 mg/min/kg EFW. Fetal electrocortical (ECOG), electroocular (EOG), breathing (FBM), nuchal muscle (NM) activities, peak CBFv, fetal blood gases and pH were monitored. Data are presented as grouped means + SEM with significance determined by paired and non paired t tests (p<0.01 vs. control group unless stated otherwise). Control animals showed LV and HV ECOG activity incidences of 53 1% and 44 1%, respectively, and of EOG, FBM, and NM activity at 562%, 433%, and 322%, respectively. Mean peak CBFv was 15517 mm/sec, increasing during LV ECOG compared with HV ECOG, averaging 142% (p<0.01). CPA animals showed LV and HV ECOG activity incidences of 75% and 64%, respectively, but a marked increase in indeterminate ECOG at 877%; EOG, FBM, and NM activity decreased (234%, 253%, and 142%, respectively). Peak CBFv averaged 618mm/sec, but was indeterminable relative to changes in ECOG activity. ZM animals showed LV and HV ECOG activity incidences of 682% and 302%; EOG, FBM, and NM activity increased (743%, 713%, and 478%, respectively). Peak CBFv (9412 mm/sec) was lower than control period and control animals (p<0.05), but increased during LV ECOG (2610%, p=0.06). Fetal blood gases and pH remained unchanged between groups or over time. Extended infusion of Ado R modulators in the ovine fetus near term affects behavioural state activity with ZM producing wakefulness, as seen in adults. CPA, in contrast, increases indeterminate ECOG but decreases EOG, FMB, NM activity, thereby producing uncoupling of these state parameters. Ado modulators may also have direct effects on CBFv in addition to those mediated through behavioural state.

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P31. Effect of sex, size at birth and metyrapone inhibition of cortisol production on body composition in young adult guinea pigs. Sanita Grover, Cathie L. Coulter, Melissa R. Walker, Karen L. Kind, Jeffrey S. Robinson and Julie A. Owens Low birth weight leads to elevated circulating glucocorticoids, reduced lean tissue and relative obesity in adult humans. Excess glucocorticoid exposure such as in Cushing’s syndrome also induces obesity. We therefore investigated the effect of birth weight on adult body composition in the guinea pig and whether the increased circulating glucocorticoids observed in this species following low birth weight contributes to any changes and whether these can be reversed by the administration of metyrapone to inhibit cortisol production. Young adult guinea pigs of known size at birth were classed into low, average or high birth weight tertiles. Vascular catheters were implanted at 100 days of age. Animals received metyrapone (100mg/kg/day IV), vehicle or no treatment for three consecutive days. Six hours after the last dose, animals were weighed (BW) then sacrificed and organs and tissues including a range of individual skeletal muscles and adipose depots weighed. Males were heavier than females (+13.4%, p=0.0001), tibialis (p=0.045), semitendinosus (p=0.001) and extensor digitorum longus (EDL) (p=0.027) were heavier, and that of biceps brachii lighter (p=0.007), in males compared to females, but not different as %BW. Adult body weight was affected by tertile (p=0.032), being reduced in low compared to average (p=0.011) and high (p=0.001) tertile animals. Tertile also altered weight of individual skeletal muscles with gastrocnemius (p=0.046) and biceps brachii (p=0.003) being reduced compared to average and/or high birth weight animals (p<0.05), but not as %BW. Tertile did interact with skeletal muscle as a repeated measures factor to alter their weights as %BW (p=0.014), with those of low and high birth weight having increased abundance compared to average (p<0.001). Treatment altered biceps brachii weight (p=0.016) and %BW (p=0.001), with metryapone increasing these compared to vehicle. Sex altered summed total fat weight (p=0.009) and summed visceral fat (p=0.021), with males showing reduced fat weight compared to females, but not as %BW. Tertile altered summed total fat weight (p=0.037), such that this was reduced in low compared to average (p=0.007) and high birth weight animals (p=0.011), but not as %BW. Treatment altered subcutaneous fat as %BW (p=0.03), with metyrapone reducing this (p=0.008) compared to no treatment. In the young adult guinea pig, low birth weight increased the abundance of lean tissue and did not later that of adipose tissue. Inhibition of cortisol production did increase the abundance of lean tissue and reduce that of adipose tissue however, suggesting that cortisol has similar actions on these and body composition as in other species. In conclusion, prenatally induced elevations in circulating cortisol observed in the young adult male guinea pig are not accompanied by increased adiposity, but whether this develops as a consequence longer term remains to be determined. P32. Uterine vessel ligation induces placental restriction and impairs mammary function but does not impact on perinatal growth following a second pregnancy Westcott, K.T. and Wlodek, M.E. Placental restriction, induced by uterine vessel ligation, is a characteristic of human intrauterine growth restriction. We have shown that placental restriction in the rat reduces birth weight, impairs maternal mammary gland function and lactation, further restricting postnatal growth. The fetal and lactational environments are implicated as critical periods in the programming of adult diseases. Our aim was to determine whether the adverse maternal effects of uterine vessel ligation, including reduced uterine blood flow and impaired mammary development, alters perinatal growth in a subsequent untreated pregnancy. In a mother’s first pregnancy, bilateral uterine vessel ligation (placental Restriction, R1) or Sham (Control, C1) surgery was performed on day 18 of gestation in Wistar Kyoto rats. In a second cohort, mothers were exposed to bilateral uterine vessel ligation or Sham surgery in their first pregnancy. Pups were born and separated from their mothers at day 35. These females where then mated 10-12 weeks after their first pregnancy with no surgical intervention (Second pregnancy, Restriction surgery in first pregnancy, R2; Second pregnancy, Control surgery in first pregnancy, C2). Mothers and pups were killed on day 6 after birth with maternal mammary and pup weight and pup dimensions measured. Placentally Restricted pups (R1) were smaller than Control (C1) (p<0.05). Mothers exposed to placental restriction in their first pregnancy (R2) produced pups in a second pregnancy that were the same size as Controls from the first (C1) or second pregnancy (C2). Placental restriction (R1) reduced litter size by approximately 50% (p<0.05) but this was not observed following a second pregnancy (R2). Maternal mammary weight and milk volume and composition were lower (p<0.05) following placental restriction (R1) compared to Control (C1) further compromising postnatal growth. The mother’s second pregnancy following the insult (R2) did not alter mammary or pup weight on day 6 indicating that lactation was not altered compared to Controls (C1 or C2). These results indicate that the effects of uterine vessel ligation causing placental and perinatal growth restriction are not translated to a mother’s second pregnancy. Placental restriction impairs mammary development during pregnancy and lactation but this effect is specific to the pregnancy in which the insult is imposed. The adverse mammary effects associated with fetal growth restriction induced by placental restriction can recover to ensure that lactation and pup growth are not affected in future pregnancies.

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P33. In Vitro Embryo Culture Affects Ovine Placental Gene Expression Fletcher CJ, MacLaughlin SM, McMillen IC, Walker S, Sibbons J, Roberts CT In vitro embryo culture (IVC) is an important feature of reproductive technologies. However, culture conditions impact on the metabolism and physiology of the developing embryos, as well as on pregnancy outcome. Culturing rodent embryos in simple defined media decreases post implantation viability and fetal growth rate 1,2. Cytokines and growth factors are present in vivo but are absent in media which may be causal in the perturbed fetal growth observed. Furthermore, the occurrence of large offspring syndrome (LOS) following embryo IVC in ruminants is associated with loss of genomic imprinting 3,4. Abnormal placental development following IVC is also likely to involve perturbed expression of imprinted genes including insulin-like growth factor II (IGF-II) and its receptor (IGF2R), as well as glucose transporters (GLUTs). This study included a control naturally mated group (NM n=10), an embryo transfer group without culture (ET n=11), an in vitro embryo culture in serum free defined medium group (IVC-NS n=10) and an in vitro embryo culture in defined medium with human serum group (IVCHS n=8). The cultured embryos were transferred to recipient ewes and placentomes were collected at 144-145d gestation. Placental weight for IVC-NS (396.26 + 35.82g) and IVCHS (436.88 + 36.7g) was similar to NM (473.58 + 35.82g), however, it was decreased in ET (374.25 + 31.43g) compared to NM (P<0.05). Real Time RT-PCR was used to quantify IGF2, IGF2R, GLUT1, GLUT3 and GLUT8 mRNA expression normalized to housekeeper RpP0 in the ET, IVC-NS and IVCHS groups (results shown in table).

IGF-II mRNA IGF2R mRNA GLUT1 mRNA GLUT3 mRNA GLUT8 mRNA ET 1.22 + 0.37 0.003 + 0.001 40.52 + 8.74ab 40.52 + 8.74a 0.53 + 0.29a IVC-NS 1.17 + 0.43 0.004 + 0.001 28.43 + 5.71a 28.43 + 5.71a 0.98 + 0.39ab IVCHS 2.27 + 0.44 0.008 + 0.003 82.82 + 21.94b 111.13 + 26.47b 3.59 + 1.58b 2-way ANOVA with bonferroni post-hoc, letters denote difference between groups P<0.05 IGF-II and IGF2R expression were increased in the IVCHS group when compared to ET and IVC-NS groups, however, this was not significant. GLUT1, 3 and 8 expression was effected by culture conditions as shown in the table. The presence of serum in IVC restored fetal growth to normal and increased expression of GLUT1, GLUT3 and GLUT8 mRNA in placental tissue near term. Our ongoing research is comparing placental gene expression from naturally mated pregnancies with the ET, IVC-NS and IVCHS groups to further dissect the role of serum in IVC media on placental and fetal development. 1. Bowman & McLaren, 1970; 2. Kaye & Gardner, 1999; 3. Thompson et al., 1995; 4. Young et al., 1998. P34. Hypertension in adult male rats is prevented by cross-fostering a growth restricted pup onto a mother with normal lactation. Mibus, A.L., Westcott, K.T., O'Dowd, R., Owens, J.A. and Wlodek, M.E. Accelerated growth in infancy and childhood following fetal growth restriction is implicated as contributing to increased risk of hypertension in the adult. In Western society, impaired placental blood flow and subsequent reduced oxygen and nutrient delivery across the placenta is a characteristic of human intrauterine growth restriction. We have shown that uteroplacental restriction in the rat impairs mammary function and pup milk intake, compromising postnatal as well as prenatal growth. Our aim was to assess the roles of the prenatal placental and/or the postnatal lactational environment in the programming of adult hypertension. Bilateral uterine vessel ligation (Restriction, R) or Sham (Control, C) surgery was performed on day 18 of gestation in Wistar Kyoto rats. Cross-fostering occurred on day 1 after birth. Pup weight as well as milk intake during lactation and food intake after week 5 were measured. Blood pressure by tail cuff was measured on weeks 8, 12 and 20. Data was analysed by ANOVA and Duncan post-hoc test. Cross-fostering a restricted male pup, who are born small, onto a mother with impaired lactation (R-on-R) reduced absolute pup milk intake on day 10 of lactation. R-on-R pups consumed less food (per kg body weight) at 5 weeks and more food at week 10 compared to pups with a normal placental and lactational environment (C-on-C). By week 12 and at week 20, blood pressure was higher (by 8mmHg) in R-on-R than C-on-C pups (p<0.05). Cross-fostering a restricted pup onto a mother with normal lactation (R-on-C), increased absolute milk intake on day 10 of lactation above that of R-on-R pups but to less than that of C-on-C pups (p<0.05). The accelerated R-on-C pup growth during lactation normalised body weight to that of C-on-C by week 4, and subsequent food intake, weight and blood pressure up to 20 weeks were similar to C-on-C. The growth and blood pressure of control pups suckled on mothers with impaired lactation (C-on-R) were also similar to that of C-on-C pups. This study demonstrates that being born small and providing adequate nutrition throughout lactation accelerates pup growth prior to but not after week 5 and protects against the programming of hypertension.

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JEFFREY ROBINSON SYMPOSIUM ABSTRACTS

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Jeffrey Robinson:

Jeffrey Robinson is an obstetrician, scientist, teacher and administrator. He has been Professor of

Obstetrics and Gynaecology and Head of Department at University of Adelaide since 1986, after

previous appointments as Professor of Reproductive Medicine at University of Newcastle NSW and

Research Program Leader at the Nuffield Institute, Oxford, England. Professor Robinson is co-author

of over 130 research articles published in peer-reviewed journals and 90 reviews, symposia or book

chapters. He has also accepted 29 invitations to present seminars at national and international

meetings over the last eight years. Since 1992 he has served as member, secretary or chair of 29 state

or national committees concerned with provision of community health services, development of

health policy, examination of medical specialists or assessment and evaluation of research project

applications and institutions. He has served as President of major national and international perinatal

societies including the Australia and New Zealand Perinatal Society and the Asia-Oceania Perinatal

Society. Jeffrey is an international leader in research in reproductive biology and medicine, and will

present the keynote Dawes lecture at this years FNPS meeting in Adelaide.

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The “fatal fetus” Jeffrey Robinson Department of Obstetrics and Gynaecology, University of Adelaide SA 5005 The “fatal fetus” refers to English as spoken by an Ulsterman and perceived by an Englishman.

However, it has a more serious aspect. Although the stillbirth rate has fallen dramatically in the last

forty years, this decline is slower than that of neonatal or infant death rates in developed societies. As

a result, the incidence of unexplained stillbirths is now higher than the incidence of other conditions,

such as SIDS. A crucial question is how can we further reduce the stillbirth rate.

Experimentalists have shown that prolonging pregnancy or reducing placental function can cause fetal

death. Barcroft originally proposed a concept sometimes referred to as “Everest in utero”. This was

not supported by later studies in the chronic fetal sheep in which there is no fall in oxygen tension

with increasing gestation, unless there is a restriction of placental growth. Placental restriction is

accompanied by fetal growth restriction, with adverse perinatal and long-term outcomes for health.

The hormonal, metabolic and physiological responses of the fetus to restriction of placental function

by a variety of means and in different species have been well defined over the last four decades.

In human growth restricted fetuses, there is a decline in oxygen tension accompanied by a rise in

haemoglobin to maintain oxygen delivery. There are numerous endocrine and metabolic changes in

the growth-restricted fetus, which closely resemble those in experimental studies, including

hypoglycaemia, hypoinsulinaemia and increased cortisol. Such fetuses are more likely to go onto

develop a range of adult onset disease, including the metabolic syndrome and to be at increased risk of

CVD. Follow up studies have also shown that a significant proportion of severely growth restricted

preterm babies from normotensive mothers have a higher incidence of medical disorders.

Setting of the growth rate of the fetus and the length of its gestation appears to begin at or before the

beginning of pregnancy. A current challenge is to define the mechanisms by which the growth of the

fetus and its placenta are set initially and how this or later growth of the placenta can be enhanced.

Dawes (1968) concluded “in normal circumstances maternal nutrition is not a limiting factor in foetal

growth”. More recent evidence indicates that fetal growth and development is sensitive to variation in

maternal nutrition within the normal range of intakes and with effects persisting across more than one

generation. Defining and manipulating these regulatory mechanisms to improve health outcomes

remains a challenge for the future.

Dawes (1968) essentially reviewed fetal and neonatal physiology over a forty-year period, beginning

with Hugget’s early work and that of others. To witness and to contribute a little to the rapid progress

since then has very exciting. In the next forty years researchers must address or resolve how the fetus

and the mother communicate within and across generations to ensure healthy survival of our species.

Ref: Dawes GS (1968) Foetal and Neonatal Physiology. Year Book Medical Publishers Inc, Chicago.

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Dr Chittarajan Yajnik (MBBS , MD), is the pre-eminent investigator of metabolic disease and its

pathophysiology in India today, with an outstanding track record in cohort and other studies. He is the

Director of the Diabetes Unit King Edward Memorial Hospital, Pune, India, which is the only unit in

India to have undertaken prospective studies with complex physiological and endocrinological

assessment. Major studies co-ordinated by him include:

Wellcome Diabetes Study in adults of varying glucose tolerance (now at 15 year follow up, with more

than 80% follow up at 10 years)

Pune Children Study in urban Pune: body size and IR syndrome in 477 children at 4 and 8 years of

age and their parents, now being followed up for study at 18 years of age (follow up rates of more

than 90%)

Pune Maternal Nutrition Study in 6 villages near Pune with pre-pregnant and gestational

measurements of body size, nutrition, insulin resistance, fetal growth, and detailed measurements of

newborn at birth. Follow up for 6 monthly growth in children, and more than 90% evaluated at 6 years

of age for body composition (BIA, DXA) and full cardiovascular risk testing - OGTT, blood pressure,

metabolic-endocrine testing both in children and parents (Maternal Origins of Diabetes in India,

MODI study, supported as a program grant by the Wellcome Trust)

CRISIS study (Coronary Risk of Insulin Sensitivity in Indian Subjects), a population-based study in

rural, urban slums and urban middle class to study body fat by anthropometry, bioimpedance,

deuterated water, MRI, DXA and CT scan, as well as insulin resistance syndrome variables and

inflammatory and endothelial function markers.

He is also an honorary founding member and Vice President of Sneha, (The Society for the Natal

Effects on Health in Adults). The main objectives of Sneha are to study the impact of maternal, fetal

and childhood factors on adult health and disease in India, to discover safe and effective ways of

improving the health and growth of the fetus and young child to get the best possible 'start in life', as

well as to train and encourage young Indian doctors and scientists to enter this field of research, and to

join the international exchange of knowledge at the highest level and to raise funds to support further

research, training and the public and scientific dissemination of knowledge.

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Maternal nutrition and offspring body composition and insulin resistance in childhood. C S Yajnik, Diabetes Unit, King Edward Memorial Hospital, Pune, India.

The ‘fetal origins’ hypothesis is based on the findings of an inverse relation between birthweight and

risk of type 2 diabetes and cardiovascular disease (CVD). The corollary was that improved maternal

nutrition leading to larger offspring size will reduce the risk of these conditions. Pune Maternal

Nutrition Study (PMNS) is one of the first purpose driven ‘prenatal cohort’ to test and elaborate this

hypothesis. We studied over 800 pregnancies in 6 villages near Pune and measured maternal nutrition,

physical activity, circulating levels of nutrients and metabolites in relation to fetal growth. The babies

have been measured every 6 months after birth and at 6y the children and the parents were assessed

for cardiovascular risk factors.

The mothers weighed 42 kg before pregnancy and had a BMI of 18.1 kg/m2. Their babies weighed 2.7

kg, and were quite ‘thin’ (ponderal index 24.1 kg/m3) but ‘fat’ (preserved skinfolds). Maternal

macronutrient intake was not predictive of neonatal size but higher frequency of eating micronutrient

rich food items (green leafy vegetables, fruits and milk) predicted larger neonatal size. Maternal

circulating levels of nutrients (vit C, folate etc) and metabolites (glucose, triglycerides and

cholesterol) also predicted neonatal size. None of the mothers smoked or drank alcohol and only a few

had gestational diabetes. Higher maternal physical activity predicted smaller offspring size.

At 6y these children were short and thin by international standards (1.22m, 16kg, BMI 13.5 kg/m2)

but were more adipose when compared with age and gender matched British children. Their adiposity

(DXA) was predicted by higher maternal folate status in pregnancy. Childrens’ insulin resistance

(HOMA-R) was predicted by higher maternal folate and vitamin C status, and by higher frequency of

intake of dairy products during pregnancy. In other words higher maternal nutrition in pregnancy

predicted adiposity and insulin resistance in the offspring. These counterintuitive findings could be

due to low vitamin B12 status in these mothers. Offspring of mothers with low vitamin B12 and high

folate status during pregnancy were the most insulin resistant at 6y of age. Our results highlight the

importance of ‘one carbon’ metabolism in nutrient partition during fetal life, possibly acting through

epigenetic control through methylation. Comparable demonstration of nutritional programming in

Agouti mice support this contention. Thus, PMNS results have provided a testable hypothesis of

‘fetal programming’ of adiposity and insulin resistance in Indians. Improving the low vitamin B12

status in mothers may help curtail the epidemic of type 2 diabetes and CV disease.

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Professor David Henderson-Smart is Professor of Perinatal Medicine at the University of Sydney

and Director of the NSW Perinatal Services Network and the Centre for Perinatal Health Services

Research. He has represented paediatrics as a member or head of numerous committees for the NSW

Department of Health and the National Health and Medical Research Council. He is a past president

of the Australian Perinatal Society and the Paediatric Research Society of Australia. He has published

widely and his internationally recognised research ranges from laboratory to population-based studies.

He is best known for his work in: regulation of breathing in the newborn; cardio-respiratory function

during perinatal development; development of standardised methods for neurological assessment of

preterm infants; auditory brainstem potentials and brain dysfunction in infants; and sensory processing

in the infant brain. David is active in the Cochrane Collaboration and has been involved in a number

of systematic reviews in the area of perinatal care.

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Clinical networks to enhance clinical care and research in neonatology

David Henderson-Smart

Centre for Perinatal Health Services Research, University of Sydney, NSW 2006

Clinical problems in neonatology generate the need for valid research to guide effective care. A wide

range of information generated by different research methods contributes to this. A framework for

generating, evaluating and applying research will be presented (adapted from Henderson-Smart et al

Clinics in Perinatology 2003;30:333-342).

The history of neonatology is littered with examples of the results of observational and basic research

results being applied directly into practice, often with adverse effects. This approach might have been

a necessity when there was little information available to guide practice. Now it is more appropriate to

test in applied research whether an intervention with theoretical benefit, works in practice.

One way to assist in ensuring that the full research cycle is completed and the results are implemented

is to develop collaborative networks. These are between clinicians of different disciples to define the

clinical problems and to ensure the best research evidence is used in practice. Also between clinicians,

epidemiologists, basic scientists and applied researchers to ensure the questions and answers are

relevant and translated into practice.

Examples of epidemiological (neonatal outcomes variation in the Australian and Neonatal Network)

and applied research (oxygen saturation targeting in preterm infants) will be presented.

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Free Oral Communications 1 Blood pressure during physiological stress in adults after prenatal exposure to the Dutch famine. R.C. Painter, S.R. de Rooij, T.J. Roseboom, P.M.M. Bossuyt, D.I.W. Phillips, C. Osmond, D.J.P. Barker, O.P. Bleker. Prenatal exposure to maternal famine in early gestation is associated with a threefold increase in coronary heart disease at age 50. Programming of stress response may explain the link between famine exposure an subsequent coronary heart disease. We performed continuous blood pressure and heart rate measurements during a battery of three 5-minute physiological stress tests (Stroop test, mirror-drawing test and a public speech task) in 723 men and women, aged 58, born as term singletons in Amsterdam around the time of the Dutch 1944-1945 famine. During the stress tests, the systolic blood pressure rose from baseline by 18 mmHg during the Stroop, by 30 mmHg during the mirror-drawing test and by 46 mmHg during the public speech task. Men were more stress responsive than women (2.0 mmHg extra systolic increase, p=0.03). Systolic and diastolic blood pressure increase during stress were highest among people exposed to famine in early gestation compared to unexposed subjects (3.2 mmHg extra systolic increase adjusted for sex, p=0.06; 1.3 mmHg diastolic increase adjusted for sex, p=0.1). Exposure during mid and late gestation were not associated with stress-related increment of blood pressure (p adjusted for sex >0.7). Correcting for potential confounders (baseline blood pressure, anti-hypertensive medication use, smoking, adult body mass index and size at birth) in a multivariable model did not attenuate the association between famine exposure in early gestation and systolic and diastolic blood pressure increment. Heart rate increment was not related to famine exposure during any part of gestation. We found a more pronounced blood pressure increase during stress among people exposed to famine in early gestation. Increased stress responsiveness may underlie the association between coronary heart disease and exposure to famine in early gestation. In future studies we aim to determine the origin of sympatho-adrenal programming after prenatal exposure to the Dutch famine. Thirty Percent Global nutrient Restriction Slows Development of the Cortical Neuronal Network in Fetal Baboons at 0.5 of Gestation M. Schwab, I Antonow-Schlorke, N. Schlabritz-Loutsevitch, K. Stuchlik, M. Brodhun, O.W. Witte, P. Nathanielsz Aim: To estimate the effects of 30% global maternal nutrient restriction on early maturation of the fetal primate brain. Methods: Pregnant baboons were either fed ad libitum (n=8) or 70% of the weight adjusted ad libitum diet (n=6). After delivery by Cesarean section at 0.5 gestation we evaluated cell proliferation (anti-ki67) and apoptotic cell death (TUNEL), the protein expression of glial (GFAP for immature and S-100 for mature astroglia) and neuronal cell lineages (DCX for immature and NeuN for mature neurons). In addition, neurite density (silver amplification) and myelination (levanol) was estimated. Specific labeling was semi-quantified using a computer-assisted image analyzing program. Results: Control and nutrient restricted fetal brains did not differ in weight or gross anatomy at 0.5 gestation. The proliferative subventricular zone thickness was reduced to 56.6 + 11.5% after maternal nutrient restriction (p<0.05). Maternal nutrient restriction led to both an increase of the number proliferating cells by 42.5 + 7.9% and apoptotic cells by 118.99 + 7.6% in the subventricular zone. Therefore, the increase in the ratio of programmed cell death to cell proliferation rose by 76.4 % approaching significance (p<0.1). In the subventricular zone, fetuses of nutrient restricted mothers showed a reduction of S-100 immunoreactivity by 65.6 + 19.6% (p<0.05). In contrast, GFAP immunoreactivity and the neuronal expression for DCX and NeuN did not change. Moreover, nutrient restriction led to a decline of neuronal arborization and myelination in the prospective neocortex by 55.5 + 23.6% and 41.0 + 29.6%, respectively (p<0.05). Conclusion: These observations indicate that moderate maternal nutrient restriction impairs brain maturation at the levels of cell proliferation, migration and development of the cortical neuronal network.

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Professor Roger Smith (MBBS; PhD). Director Mother and Babies Research Centre Endocrinology

Unit, John Hunter Hospital. Professor Smith studied at the University of Sydney (MBBS 1974) and

London, UK (PhD 1981; on opioid peptides in man). He has published extensively on the

endocrinology of pregnancy. In 1995 his work, which established the presence of a biological clock

determining the length of human pregnancy, achieved not only scientific acclaim but also widespread

interest in the media. More recently a review of parturition research in Scientific American gained

worldwide coverage. The American College of Obstetricians and Gynaecologists convened a meeting

in 1998 specifically to discuss the implications of his group's research for obstetric practice in the

United States. In 2000 he was awarded the Society for Endocrinology medal for Australia.

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Turning on the Uterus

Roger Smith

Mothers and Babies Research Centre, Hunter Medical Research Institute, John Hunter Hospital,

University of Newcastle, Newcastle, NSW, Australia.

The mechanism that drives uterine activation in the human has been a challenging enigma throughout

the last century. Gradually evidence is accumulating that inflammatory pathways are a crucial

component of the process. Histological studies reveal that macrophage infiltration is pronounced in

labouring human myometrium although the aetiology of this infiltrate in the human is presently

unclear. Work in the rat suggests that in that species fetal lung maturation is linked to production of

surfactant protein A released into the amniotic fluid, within the amniotic fluid surfactant protein A

activates macrophages which then migrate across the membranes and into the myometrium provoking

an inflammatory response which initiates labour. In the human an analogous process may occur with

fetal lung development driven by placental CRH production. Using myometrial samples obtained at

caesarian section and quantitative mRNA measurements of inflammatory and contraction associated

genes a model for human myometrial activation has been generated and tested using recently

developed artificial intelligence techniques. The results strongly support an inflammatory origin for

human myometrial activation. The pathway that leads to increased myometrial force development

remains to be clarified. Progress in this area is identifying potential targets for new tocolytic

therapeutics.

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Professor Leslie Myatt (PhD U London) is Professor of Obstetrics and Gynecology, Director of the

Physician Scientist Training Program, Program Director, Women's Reproductive Health Research

Scholars Program at the University of Cincinnati, College of Medicine. He is the current President of

the International Federation of Placenta Associations and former President of the Perinatal Research

Society. He is an international authority on the placental development and the intra-cellular

mechanisms leading to birth and has published over 160 primary research papers. He served as the

Editor of Placenta from 1997-2004, which is the major international journal in this field. Throughout

his career he has received a number of awards including a Royal Society Fellowship and Fulbright

Travel Grant in 1984 to enable him to undertake research at the University of Louisville, Kentucky,

USA. In 1998, he was the Dozor Visiting Professor at Ben Gurion University of Negev, Israel. In

2002, the J Ross Vant Memorial Visiting Lecturer at the University of Alberta, Canada and the Vice

Chancellor's Distinguished Visitor at the University of Auckland.

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The Placenta: Active or Passive Participant in Programming?

Les Myatt

Alterations in fetal development or adaptations of the fetus to the amount or pattern of substrate

delivery across the placenta leads to cardiovascular or metabolic disease in adult life. Adaptations of

placental structure/function may regulate the amount and composition of substrate transferred i.e the

placenta may play and active rather than a passive role in fetal programming. Manipulations in animal

models that lead to altered placental structure/function identify a critical role for the placenta and

associative data from human studies support this concept. Mechanisms whereby structure/function

might be altered include abnormal placental vasculogenesis leading to altered placental blood flows

and nutrient transfer, abnormal trophoblast differentiation/function leading to altered barrier thickness,

altered expression of transporter proteins and synthesis of peptide and steroid hormones or insults

such as oxidative/nitrative stress that occur at critical times in placental development and alter

function.

Adequacy of blood flows is a major determinant of nutrient transfer. Indeed in IUGR, preeclampsia

and diabetes abnormalities in both maternal- and fetal-placental blood flows are seen and associated

with programming. Oxygen is a major determinant of vasculogenesis as hypoxia regulates trophoblast

invasion of spiral arteries and development of the villous vascular tree. Hypoxia acting via angiogenic

factors causes branching angiogenesis and hypercappilarization. Recent work has highlighted the role

of the placental barrier (trophoblast) function in modulation of growth. There is evidence from human

studies of adaptive alterations in expression and activity of glucose and amino acid transporters with

pregnancy pathologies, eg diabetes or IUGR, associated with programming. The timing of the insult

that leads to the pathology may be important in this regard. Covalent modifications of regulatory

proteins by oxidative and nitrative stress may affect placental development/function via their effects

on specific placental tissues. Mouse studies have shown the inter-relationship of altered barrier

thickness, altered passive permeability and regulation of specific transporter systems, highlighting the

role of barrier function in modulation of fetal growth.

Imprinted genes also regulate overall placental growth, growth of specific cell types and expression of

specific transporters and are thus able to exert a role in placental development/function.

Environmental factors and maternal nutrition regualte imprinting by epigenetic mechanisms.

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Free Oral Communications 2 Male seminal factors influence fetal and placental development, postnatal growth and adult onset obesity in mice. John J Bromfield, Claire T Roberts and Sarah A Robertson The development of an optimal uterine environment early in pregnancy is critical for the future development of the fetus and placenta. Constituents within seminal plasma, derived primarily from the seminal vesicle glands, are known to be powerful modulators of the early uterine environment, through regulating female tract cytokine expression and tissue remodeling. It is unclear however whether the absence of seminal plasma at the time of conception has detrimental effects on the outcomes of pregnancy or the subsequent progeny. In this paper we investigated the impact of initiating pregnancies in the absence of seminal plasma by means of utilising male mice to which the seminal vesicles had been surgically removed. Seminal vesicle deficient males displayed a highly significant perturbation in their ability to initiate pregnancies when compared to intact males, due primarily to a failure in fertilization potential. Outcomes for viable implantation were also adversely affected. Near term placentas from these matings displayed a 15% increase in weight compared to those from control matings. While fetal and new born weights were significantly reduced, the progeny of seminal vesicle deficient males displayed ‘catch-up growth’ during puberty. These progeny proceeded to become heavier than control progeny, with male progeny displaying an increased abdominal fat deposition. Male progeny displayed reduced plasma adiponectin and increased levels of plasma leptin, a profile associated with obesity. Female progeny exhibited increased plasma insulin and significantly increased levels of glucose and free fatty acids. This study is the first to show that uterine exposure to seminal plasma at the time of conception has a major impact on the development of the fetus, placenta and progeny of these pregnancies.

The Effect of Hypoxia to Promote Human Cytotrophoblast Outgrowth is Mediated by Insulin-like Growth Factor-II AN Sferruzzi-Perri, JS Robinson, CT Roberts Poor placental development has been associated with a variety of pregnancy-associated disorders, including preeclampsia. Placentation involves spatial and temporal regulation of cytotrophoblast (CTB) proliferation, differentiation and invasion into the maternal decidua. These processes, during early pregnancy occur in a relatively hypoxic environment and insulin-like growth factor (IGF)-II is abundantly expressed by the invading extravillous CTBs. Transforming growth factor (TGF)-1 is predominantly synthesised by decidual cells and requires activation to have biological effects. TGF 1 has opposing effects to IGF-II as it inhibits CTB invasion. Therefore, this investigation aimed to determine the effect and interaction of O2 concentration and IGF-II on placental outgrowth and activation of TGF1 during the first trimester in vitro. First trimester (7-8 weeks gestation) human placental villous explants were cultured in serum-free media with and without the addition of 125ng/ml IGF-II, in 1%, or 20% O2 (4 groups). On day 6, explants were morphometrically analysed, assayed for TGF-b1 protein activation and IGF-II, TGF-b1 and type 1 and 2 IGF receptor (IGF1R, IGF2R) gene expression relative to 18S was quantitated (Real Time RT-PCR). Culture in 1% O2 or with IGF-II stimulated extravillous CTB outgrowth (p<0.02) and inhibited TGF1 activation (-90% and -65%, respectively p<0.05). IGF-II gene expression was increased 3.5-fold in 1% O2 (p=0.04), however there was no effect of treatment on TGF 1, IGF1R or IGF2R transcription. CTB outgrowth and migration were positively correlated with IGF-II and IGF2R transcription, whereas activation of TGF1 was negatively correlated with these parameters. In conclusion, our data suggest that the low oxygen environment of the placenta during early pregnancy stimulates placental outgrowth and IGF-II expression, while it inhibits TGF 1 activation. IGF-II appears to mediate the low oxygen response as addition of IGF-II to 20% O2 explants promotes CTB outgrowth and inhibits TGF1 activation. This novel interaction of oxygen, IGF-II and TGF 1 during pregnancy is an important determinant of CTB invasive behaviour. This research may be important in understanding pregnancy disorders characterised by impaired placental invasion of the maternal decidua, such as preeclampsia.

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Influence of Placental Restriction and the Lactational Environment on Vascular Function in Adulthood M. Tare, H.C. Parkington, A.L. Mibus, J.A. Owens, M.E. Wlodek Intrauterine growth restriction occurs in 10% of pregnancies and in many cases arises from a poorly functioning placenta. This has been associated with increased risk of cardiovascular disease in adult offspring. However, the extent of programmed morbidity may be further accentuated by nutrition during lactation. Recent evidence in the rat indicates that milk production and quality is impaired in lactating dams that had experienced placental restriction during pregnancy. Here we have investigated the separate influences of placental restriction and the lactational environment on the reactivity of mesenteric arteries in adult male rats. Bilateral uterine vessel ligation (Restriction, R) or sham surgery (Control, C) was performed on day 18 of gestation in Wistar Kyoto rats, and pups were cross-fostered one day after birth. Male offspring were killed at 6 months of age and ring segments of small mesenteric artery were mounted on a wire myograph to enable assessment of smooth muscle and endothelial function. In placentally restricted rats suckled on a mother with impaired lactation (R-on-R), vascular contraction to phenylephrine was suppressed, compared with controls (C-on-C). Sensitivity and maximal response to the vasodilator sodium nitroprusside (SNP) was reduced in arteries from R-on-R compared with C-on-C offspring. Stimulation of the endothelium evoked equivalent relaxation in arteries from all offspring. However, after blockade of nitric oxide synthase and cyclooxygenase activity, the remaining relaxation attributable to endothelium-derived hyperpolarizing factor, was reduced in arteries from R-on-R compared with C-on-C offspring. All of the programmed alterations in vascular reactivity of R-on-R offspring could be completely prevented by cross-fostering onto control dams with normal lactation (R-on-C). Mean arterial pressure (MAP) in R-on-R offspring was 8mmHg higher than in C-on-C pups and this was prevented by cross-fostering. Control offspring subjected to lactational restraint (C-on-R) had unchanged MAP and vascular responses to phenylephrine and endothelium-dependent relaxation, but increased vascular sensitivity to SNP. In conclusion, this study demonstrates that placental restriction followed by lactational restraint causes hypertension and vascular dysfunction in adult male rats, and that a normal lactational environment prevents these adverse consequences. Furthermore, an impaired lactational environment induced significant, albeit subtle, vascular changes in adult offspring, further substantiating early postnatal life and lactation as determinants of later cardiovascular function.

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Professor Jane Harding BSc, MBChB, DPhil, FRACP, FRSNZ is an international authority on the

regulation of fetal growth, placental function and the treatment and consequences of undernutrition in

the womb. Both a fetal physiologist and paediatrician, she trained in Oxford as a Rhodes scholar and

in Auckland and San Francisco. She was Visiting Assistant Professor in the University of California 's

San Francisco Division of Neonatology before returning to Auckland as a Senior Lecturer in

Paediatrics (Neonatology). She does both clinical and experimental research, and spends a portion of

each year working at Auckland National Women's Hospital's newborn intensive care nursery.

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What should women eat during pregnancy, and does it matter?

Jane E Harding

Liggins Institute, Faculty of Medical and Health Sciences, University of Auckland, Private Bag

92019, Auckland, New Zealand.

Diet during pregnancy is a subject of folklore and divisive debate, particularly within families. There

is growing evidence that maternal nutrition before and during pregnancy can alter the growth of the

fetus, and influence its lifelong health. However the accumulation of evidence on which to base

rational recommendations regarding maternal diet during pregnancy is much slower.

Pre-pregnancy maternal weight and body composition influence the growth of the fetus, although the

underlying mechanisms remain uncertain. Maternal diet during pregnancy also influences fetal

growth. Total macronutrient intake, balance of macronutrients, and a number of micronutrients have

all been shown to alter size of the baby at birth, and their subsequent postnatal physiological

functioning.

Maternal undernutrition around the time of conception alters early growth of the offspring, and also

changes many aspects of fetal metabolism and endocrinology in late gestation. In principle such

changes could be due to effects of periconceptional undernutrition on the overall rate of fetal

maturation, on placental development and function, or on maternal physiological adaptation to

pregnancy. There is some evidence that all of these aspects may be involved, although their relative

contributions remain unclear. The postnatal growth and physiology of fetuses exposed to maternal

undernutrition in the periconceptional period remain to be examined.

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Professor David Hill (PhD), Scientific Director, Lawson Health Research Institute, University of

Western Ontario, Canada and Professor in Medicine, Physiology and Paediatrics at The University of

Western Ontario. Dr. Hill is also Vice President of Research at St. Joseph 's Health Care, London. He

has a number of administrative roles on local and international Scientific boards, committees and

organizations, such as the Chair of the National Research Committee of the Canadian Diabetes

Association. He also serves as a Director for 3 companies in London, Canada; the Stiller

Biotechnology Commercialisation Centre, FRALEX Therapeutics and Multi-Magnetics, Inc. Dr. Hill

is an active researcher recognized internationally for his work in the area of diabetes. David's research

program is focused at the interface of diabetes, pancreatic development and stem cell biology, fetal

programming of maturity onset diabetes, and the environmental impact of nutrition on tissue

plasticity. In particular, t he fetal programming through nutrition of the endocrine pancreas as a risk

factor for type 2 diabetes in later life. The identification and biology of endocrine pancreatic stem

cells and their differentiation into functional ß-cells. Pharmaceutical and nutraceutical strategies for

the survival of ß-cells in type 1 diabetes. The expression, control and action of peptide growth factors

such as insulin-like growth factors and fibroblast growth factors.

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The plastic pancreas: Early developmental decisions with life-long consequences.

*David J Hill, Lawson Health Research Institute, London, Ontario, Canada.

* Faculty of Health Sciences Senior Visiting Research Fellowship

Altered development of the endocrine pancreas in the fetus and neonate, as can result from maternal

nutritional insult, leads to impaired metabolic control in adulthood. Understanding the origins of

pancreatic β-cells is key to revealing where the primary insult is directed, and in designing reversal

strategies that could also be applied to a more general regeneration of β-cells in individuals with

insulin deficiency. The pancreas develops from the embryonic mid-gut as a ductal tree from which

presumptive endocrine cells are generated and defined through the expression of a sequence of

transcription factors. Expression of Pdx-1 is necessary to enable the endocrine lineage, and also for

the maturation of the functional β-cell. Morphogenesis and differentiation of the islets also depends

on growth factors derived from the supporting vascular network, such as vascular endothelial growth

factor, the mesenchyme as for fibroblast growth factors –7 and –10, and from the endocrine cells

themselves such as insulin-like growth factor-II. Glucagon-like polypeptide-1 (GLP-1) is synthesized

by α-cells during development and can control β-cell differentiation, proliferation and survival. While

multi-potent stem cells exist within the developing pancreas, endocrine cell plasticity is also possible

by trans-differentiation of acinar or ductal cells into β-cells. Mature endocrine cells can also de-

differentiate into a multi-potential, proliferative precursor phenotype. However, the initiation of islet

regeneration may require interaction with endothelial precursor cells derived from bone marrow.

Manipulation of this population results in an increased β-cell mass. The capacity for compensatory

growth of endocrine cells decreases with age, and might be linked to the presence of endogenous islet

or endothelial precursor cells. The effects of maternal nutritional insufficiency on fetal islet

development can be rescued either by selective nutritional supplementation, or by the use of trophic

peptides, such as GLP-1.

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Free Oral Communications 3 Molecular markers of early-stage lung injury in ventilated, very preterm lambs. UM.J. Wallace U, M.E. Probyn, F. Sozo, A.M. Thiel, C.J. Morley, R. Harding and S.B. Hooper Background: Infants born very preterm (<30 weeks GA) are at high risk of developing chronic lung disease (CLD) resulting from an inflammatory process in response to lung injury induced by resuscitation and artificial ventilation. CLD is characterised by disrupted alveolarization resulting in fewer, larger alveoli and abnormal growth of the alveolar capillaries. However, determination of ventilation strategies that are not injurious to the lung is presently restricted by a lack of early response markers that reliably reflect ventilator-induced lung tissue injury. Aim: To identify molecular markers that are rapidly induced in response to lung tissue injury in ventilated preterm lambs Methods: Premature lambs (n=16) were delivered by C-section at ~132 days GA (term is ~147 days), intubated and ventilated for 15 minutes using an injurious ventilation protocol (tidal volume (VT) of 20 ml/kg and 0 cm H20 PEEP). After 15 min, lambs were ventilated using a VT of 5 ml/kg and a PEEP of 8 cm H20 and then killed 15, 30, 60 or 120 min later (n=4 for each) for lung tissue collection. Markers of lung tissue injury examined included NFκB, TNFα, TGF1, CTGF, CYR61, EGR1, IRF2-BP2 and VEGF; levels were measured using gel shift assays, ELIZA assays, Northern and Western blots and immunohistochemistry, depending on the marker. Results: Injurious lung ventilation induced a rapid elevation in lung tissue mRNA levels for CYR61 (645% at 15 min), EGR1 (432% at 15 min), CTGF (344% at 30min) and IRF2-BP2 (244% at 30min), in comparison with unventilated controls. The proportion of lung tissue recognised by a CYR61 antibody was also increased 6.5 fold, 2 hours after injurious ventilation. NFκB activation, TNFα protein levels, TGF 1 mRNA and VEGF mRNA levels were not significantly elevated within 2 hours of inducing lung tissue injury in preterm lambs. Conclusions: These data indicate that the mRNA levels for CTGF, CYR61, EGR1 and IRF2-BP2 in lung tissue are rapidly (within 30min) and markedly elevated by ventilator induced lung injury in very premature lungs. As CTGF and CYR61 are secreted proteins, they may serve as useful early response markers of ventilation-induced lung injury in ventilated very preterm infants.

Maternal nutrient restriction in late gestation up regulates mRNA abundance for the insulin receptor and peroxisome proliferator-activated receptor gamma in adipose tissue of newborn sheep. E.A.Butt, S Pearce, T. Stephenson, M.E. Symonds and UH BudgeU

Maternal nutrient restriction targeted over the final month of gestation results in a reduced fat mass in the newborn but greater fat deposition in the resulting offspring P

(1)P. These adaptations may be mediated in part by

increased sensitivity of fat in the newborn to insulin although this has yet to be established. Insulin also regulates fat growth by stimulating peroxisome proliferator-activated receptor gamma (PPARγ) that is involved in the cascade of events leading to adipogenesis and transcribes genes like phosphoenol pyruvate carboxykinase (PEPCK), the rate limiting enzyme involved in adipocyte glycerogenesis. The aim of this study was to determine whether maternal nutrient restriction in late gestation results in enhanced insulin sensitivity in adipose tissue in the newborn. Eighteen pregnant sheep of similar age, body weight and condition score were entered into the study of which nine were nutrient restricted (NR - 50% of metabolizable energy) from 110 days gestation until term (145 days) and nine (controls; C) were fed to requirements. All lambs were born spontaneously and were humanely euthanased at one day of age to enable perirenal adipose tissue sampling. The relative abundance of insulin receptor (IR), PPARγ, and PEPCK mRNA to 18S rRNA were determined using RT-PCR and all results are expressed in arbitrary units (a.u.). The mRNA abundance for all three genes examined were significantly upregulated in the NR offspring compared to C i.e. IR - NR 179U+U18; C 33U+U4 a.u. (P<0.01; PPARγ - NR 112U+U8; C 67U+U4 a.u. (P<0.05), and PEPCK - NR 8.6 U+U0.7; C 1.3 U+U0.4 a.u. (P<0.01). In conclusion, maternal nutrient restriction coincident with the period of maximal fat deposition in the fetus increases insulin sensitivity within the adipocyte in conjunction with an increase in abundance of adipogenic-related genes. These adaptations may explain in part why these offspring have increased fat mass in later life P

(1)P.

1. Gardner DS, Tingey K, van Bon BWM, Ozanne SE, Wilson V, Dandrea J, Keisler DH, Stephenson T, and Symonds ME. Am J Physiol: doi:10.1152/ajpregu.00120.02005, 2005.

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Maternal Overnutrition Programmes Postnatal Appetite Regulation: Increased Nutrition Before Birth Increases Hypothalamic Expression of the Appetite-Inhibiting Neuropeptide POMC in the Neonatal Lamb BS Muhlhausler, CL Adam, PA Findlay, JA Duffield, IC McMillen Introduction: Epidemiological studies have demonstrated that infants of diabetic mothers, who are exposed to an elevated nutrient supply during development, are at increased risk of altered appetite regulation and obesity in later life. In the present study we have therefore determined the effect of increasing maternal nutrient supply on components of this appetite-regulating neural network in the early postnatal period. Methods: From 115 d gestation until delivery pregnant ewes were fed a diet which provided either 100% (control, n=12) or ~150% (Well Fed, n=9) of maintenance energy requirements (MER). Lambs were delivered spontaneously and birthweight was recorded within 6 h of birth. Blood samples were collected every 72 hours for the determination of glucose, insulin and leptin concentrations. On postnatal day 30, postmortem was preformed and brains were collected. In situ hybridisation was then used to determine the expression of the appetite-stimulating neuropeptides NPY and AGRP and appetite-inhibiting neuropeptide precursor Proopiomelanocortin (POMC) and neuroeptide Cocaine-Amphetamine-Related Transcript (CART) in the hypothalamic arcuate nucleus. Results: Maternal nutrient intake between 115 d gestation and delivery was higher in the Well Fed group (129 + 4 % vs 90 + 2 % MER, P<0.001). Plasma concentrations of glucose, but not insulin or leptin, were significantly higher in lambs of Well Fed ewes compared to Controls (F=5.93;P<0.05). Hypothalamic expression of the appetite-inhibiting neuropeptide precursor POMC was higher in the Well Fed group (0.48 + 0.09 vs 0.28 + 0.04, P<0.05) and tended (P=0.07) to be positively correlated with plasma glucose concentrations when data from all lambs were combined. CART mRNA expression was positively correlated with plasma glucose, insulin or leptin concentrations in Control, but not Well fed lambs. NPY and AGRP were inversely correlated with total relative fat mass in both control and Well Fed groups (NPY, r=0.53 P<0.04; AGRP, r=0.63 P<0.01). There was no difference in hypothalamic expression of CART, NPY or AGRP mRNA between the Control and Well fed groups. Conclusion: We have therefore demonstrated that increasing maternal nutrient intake in late gestation results in changes to the appetite regulating neural network after birth. This provides evidence that important components of the appetite regulating system may be programmed by exposure to increased nutrient supply before birth.

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List of Delegates

Dr Amit Agrawal Training Department Baudha, Timchule PO Box 1144 KATHMANDU NEPAL [email protected] Mr Paul Allen Accompanying Person, Land Agent 19 Bakers Lane Shutlanger TOWCESTER NORTHAMPTONSHIRE UK [email protected] Ms Beth Joyce Allison PhD Student C/O Physiology Department Wellington Rd CLAYTON VIC AUSTRALIA [email protected] A/Prof Laura Bennet Associate Professor Private Bag 92019 AUCKLAND NEW ZEALAND [email protected] Dr Tracey Bjorkman NHMRC Research Officer Level 6 Ned Hanlon Building Royal Brisbane And Women's Hospital Butterfield Street HERSTON QLD AUSTRALIA [email protected]

Dr Jane Black Senior Lecturer Dept Of Anatomy PO Box 13C CLAYTON VIC AUSTRALIA [email protected] Dr Frank Bloomfield Senior Lecturer In Neonatology Private Bag 92019 AUCKLAND NEW ZEALAND [email protected] Dr. Alan Bocking Professor And Chair Mount Sinai Hospital, Room 1006 600 University Avenue TORONTO ONTARIO CANADA [email protected] Mrs Barbara Bocking Accompanying Person 6 Nesbitt Drive TORONTO ONTARIO CANADA [email protected] Ms Kimberley Botting Honours Medical School South Level 4 University Of Adelaide ADELAIDE SA AUSTRALIA [email protected]

Mr John Bromfield Postgraduate Candidate Department Of Obstetrics And Gynaecology 6th Floor Medical School Nth, Frome Road ADELAIDE SA AUSTRALIA [email protected] Dr Russell Brown Post Doctoral Fellow Dept Of Physiology SYDNEY NSW AUSTRALIA [email protected] Dr Helen Budge Academic Division Of Child Health E Floor, East Block University Hospital NOTTINGHAM UNITED KINGDOM [email protected] Ms Natasha Campbell PhD Student Department Of Obstetrics University Of Adelaide ADELAIDE SA AUSTRALIA [email protected] Miss Priscila Cassaglia PhD Student Ritchie Centre Level 5 Monash Medical Centre 246 Clayton Rd CLAYTON VIC AUSTRALIA [email protected]

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Prof John Challis Vice-President, Research And Associate Provost 27 King's College Circle TORONTO ON CANADA [email protected] Miss Joanne Chua PhD Student Frome Road ADELAIDE SA AUSTRALIA [email protected] Mr Bernard Chuang Student 709 Residential Wing, Royal Adelaide Hospital ADELAIDE SA AUSTRALIA [email protected] Prof Paul Colditz Director Royal Brisbane And Women's Hospital Butterfield Street HERSTON QLD AUSTRALIA [email protected] Dr Kristin Connor PhD Graduate Student 1 King's College Circle Medical Sciences Building Room 3344 TORONTO ONTARIO CANADA [email protected] Dr Catherine Coulter Discipline Of Physiology School Of Molecular University Of Adelaide ADELAIDE SA AUSTRALIA [email protected]

Prof Caroline Crowther Professor Of Obstetrics And Gynaecology Department Of Obstetrics And Gynaecology ADELAIDE SA AUSTRALIA [email protected] Ms Penelope Dalitz Building 13F, Department Of Physiology Monash University Wellington Road CLAYTON VIC AUSTRALIA [email protected] Dr Miles De Blasio Postdoctoral Research Officer Department Of Obstetrics University Of Adelaide ADELAIDE SA AUSTRALIA [email protected] Prof Jelte De Haan Research Institute Growth & Development Postbus 5800 MAASTRICHT NETHERLANDS [email protected] Mr Justin Dean PhD Park Avenue Grafton AUCKLAND NEW ZEALAND [email protected]

Dr. Kate Denton Senior Research Officer Department Of Physiology Monash University Wellington Road CLAYTON VIC AUSTRALIA [email protected] Ms Hayley Dickinson PhD Student Wellington Rd CLAYTON VIC AUSTRALIA [email protected] Dr Greg Duncombe Staff Specialist In Maternal Fetal Medicine Raymond Terrace South Brisbane BRISBANE QLD AUSTRALIA [email protected] Miss Stacey Dunn Research Assistant Frome Road ADELAIDE SA AUSTRALIA [email protected] Dr Susan, Y.S. Feng PhD Student Monash Medical Centre 246 Clayton Rd CLAYTON VIC AUSTRALIA [email protected] Mrs Cherise Fletcher Level 6, Medical School North Frome Rd ADELAIDE SA AUSTRALIA [email protected] Dr Alison Forhead University Lecturer Department Of Physiology Downing Street CAMBRIDGE CAMBRIDGESHIRE UK [email protected]

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Ms Kathryn Franko PhD Student Downing St CAMBRIDGE CAMBRIDGESHIRE, UK [email protected] Dr Kathryn Gatford Postdoctoral Research Officer Department Of Obstetrics University Of Adelaide ADELAIDE SA AUSTRALIA [email protected] Dr Karen Jane Gibson Senior Lecturer Dept Of Physiology School Of Medical Sciences, UNSW SYDNEY NSW AUSTRALIA [email protected] Ms Patricia Grant Research Officer Department Of Obstetrics University Of Adelaide ADELAIDE SA AUSTRALIA [email protected] Ms Sanita Grover PhD Student Department Of Obstetrics University Of Adelaide ADELAIDE SA AUSTRALIA [email protected] Ms Lina Gubhaju PhD Student Dept Of Anatomy PO Box 13C CLAYTON VIC AUSTRALIA [email protected]

A/Prof Alistair Jan Gunn Department Of Physiology, University Of Auckland Private Bag 92019 AUCKLAND GRAFTON NEW ZEALAND [email protected] Hedieh Hamrahi PhD Student 268 Grosvenor St. Room G513 St. Joseph's Hospital LONDON ONTARIO CANADA [email protected] Prof Jane Harding Private Bag 92019 AUCKLAND NEW ZEALAND [email protected] Prof Richard Harding Professorial Fellow Wellington Rd CLAYTON VIC AUSTRALIA [email protected] Ms Erica Harnett MSc Candidate Department Of Anatomy Botterell Hall, Rm 915 Queen's University At Kingston KINGSTON ONTARIO CANADA [email protected] Dr Tom Hartgill PhD Student Ullevaal University Hospital Kirkeveien 166 OSLO NORWAY [email protected]

Prof David Henderson-Smart Director, NSW Pregnancy And Newborn Services Network D02 - Queen Elizabeth II Research Institute And Coppleson In The University Of Sydney SYDNEY NSW AUSTRALIA [email protected] Prof David Hill Scientific Director, Lawson Health Research Institute And La St. Joseph's Health Care 268 Grosvenor Street LONDON ON CANADA [email protected] Dr Stuart Hooper Assoc. Prof. Department Of Physiology MONASH UNIVERSITY VIC AUSTRALIA [email protected] Dr Noriaki Imai Obstetrics & Gynaecology 1-1 Seiryomachi Aobaku SENDAI JAPAN [email protected] Dr Benjamin Jackson 11 October Ln WESTON MA USA [email protected] Dr Juanita Jellyman Postdoc 212 Dickinson Street Ctf B211 SAN DIEGO CA USA [email protected]

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Professor Graham Jenkin Associate Director Monash University Level 3, STRIP 1 Wellington Road CLAYTON VIC AUSTRALIA [email protected] Dr Ellen Jensen Research Fellow Department Of Physiology Faculty Of Medical And Health Science AUCKLAND NEW ZEALAND [email protected] Ms Sylvia Johnson PO Box 1024 DC BROOKVALE NSW AUSTRALIA [email protected] Dr. Neerja Kamal Registrar(Obs. Unit 5 74 Ward Street North Adelaide SA AUSTRALIA [email protected] Ms Rebecca Kelley Research Assistant The Queen Elizabeth Hospital 28 Woodville Rd WOODVILLE SA AUSTRALIA [email protected] Dr Michelle Kett Senior Research Officer Dept Physiology PO Box 13F MONASH UNIVERSITY VIC AUSTRALIA [email protected]

Dr Karen Kind Research Officer Level 4, Maternity The Queen Elizabeth Hospital WOODVILLE SA AUSTRALIA [email protected] Professor Brian Koos 10833 Le Conte Avenue 22-128 Chs LOS ANGELES CALIFORNIA USA [email protected] Ms Poonam Ligam PhD Student Physiology Department Wellington Road CLAYTON VIC AUSTRALIA [email protected] Dr Barbara Lingwood Postdoctoral Research Fellow Royal Brisbane And Women's Hospital Butterfield Street HERSTON QLD AUSTRALIA [email protected] Prof Eugenie Lumbers Professor High St Randwick SYDNEY NSW AUSTRALIA [email protected] Prof Stephen Lye Vice-President, Research Mount Sinai Hospital 600 University Ave, Room 982 TORONTO ONTARIO CANADA [email protected]

Mr. Severence MacLaughlin Student Department of Physiology University Of Adelaide Frome Rd ADELAIDE SA AUSTRALIA [email protected] Ms Devaki Maduwegedera PhD Student Building 13F, Department Of Physiology Monash University Wellington Road CLAYTON VIC AUSTRALIA [email protected] Dr Carina Mallard Box432 GOTEBORG SWEDEN [email protected] Dr Simon Malpas Private Bag 92019 Auckland AUCKLAND NEW ZEALAND [email protected] Ms Adelle McArdle PhD Student Department Of Physiology Monash University, Wellington Road CLAYTON VIC AUSTRALIA [email protected] Jeremy McCallum Trainee 268 Grosvenor Street LONDON ONTARIO CANADA [email protected]

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Ms Amy Mibus PhD Student Royal Pde PARKVILLE VIC AUSTRALIA [email protected] Ms. Laura Moore MSc Candidate Botterell Hall Stuart Street KINGSTON ONTARIO CANADA [email protected] Dr Karen Moritz Research Fellow Wellington Rd CLAYTON VIC AUSTRALIA [email protected] Dr Janna Morrison Research Fellow Medical School South Frome Rd ADELAIDE SA AUSTRALIA [email protected] Dr Timothy Moss Senior Research Fellow School Of Women's M094, 35 Stirling Highway CRAWLEY WA AUSTRALIA [email protected] Ms Beverly Muhlhausler PhD Student Discipline Of Physiology School Of Molecular And Biomedical Science ADELAIDE SA AUSTRALIA [email protected]

Prof Leslie Myatt Professor Of Obstetrics And Gynecology College Of Medicine PO Box 670526 CINCINNATI OH USA [email protected] Sarah Newhouse M.Sc. Candidate 625 Heritage Medical Research Center University Of Alberta EDMONTON ALBERTA CANADA [email protected] Prof. Dr. Jan G. Nijhuis Prof Of Obstet, Head Of Dept. Mat. Fetal Medicine, POBox 5800 MAASTRICHT-NL LIMBURG THE NETHERLANDS [email protected] Dr Hidekazu Nishigori 268 Grosvenor Street LONDON ONTARIO CANADA [email protected] Dr Ilias Nitsos Research Fellow M094 35 Stirling Highway CRAWLEY WA AUSTRALIA [email protected] Cornelle Noorlander PhD Student Stratenum, RMI Universiteitsweg 100 UTRECHT THE NETHERLANDS [email protected]

Miss Rachael Nowak Department of Obstetrics and Gynaecology University of Adelaide Frome Rd ADELAIDE SA AUSTRALIA [email protected] Miss Amanda E O'Connell PhD Student Dept Of Physiology School Of Medical Sciences, UNSW SYDNEY NSW AUSTRALIA [email protected] Dr Deirdre O'Connor Post-Doctoral Research Associate Downing Street CAMBRIDGE CAMBRIDGESHIRE UNITED KINGDOM [email protected] Ms Rachael O'Dowd PhD Student Royal Pde PARKVILLE VIC AUSTRALIA [email protected] Prof. Kunihiro Okamura Professor 1-1,Seiryomachi, Aobaku SENDAI MIYAGI JAPAN [email protected] Dr. David M. Olson 220 H.M.R.C. University Of Alberta EDMONTON ALBERTA CANADA [email protected]

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Prof Julie Owens Professor Department Of Obstetrics University Of Adelaide ADELAIDE SA AUSTRALIA [email protected] Dr. James Padbury Professor And Associate Chair Of Pediatrics 101 Dudley Street PROVIDENCE RI USA [email protected] Dr Rebecca Painter PhD Student Room J1b-227 PO Box 22660 AMSTERDAM NORTH HOLLAND THE NETHERLANDS [email protected] Md Mervi Pakkila Specialist Doctor/obstetrics Kajaanintie 50 OULU FINLAND [email protected] Dr Nayana Parange PhD Candidate Department Of Obstetrics And Gynaecology Women's & Children's Hospital ADELAIDE SA AUSTRALIA [email protected] Prof Julian (Bill) Parer Professor, Dept Of OB/GYN 505 Parnassus Avenue M-1489 Box 0132 SAN FRANCISCO CA USA [email protected]

Lindsay Patrick PhD Student Botterell Hall 863 Stuart St KINGSTON ONTARIO CANADA [email protected] Dr Donald Peebles Reader 86-96 Chenies Mews LONDON UNITED KINGDOM [email protected] Prof. Jouko Pirhonen Head Of Department Ullevaal University Hospital Kirkeveien 166 OSLO NORWAY [email protected] Dr Julia Pitcher NHMRC Peter Doherty Research Fellow 6th Floor, Medical School North Frome Rd ADELAIDE SA AUSTRALIA [email protected] Dr Graeme Polglase Research Associate 35 Stirling Highway (M094) CRAWLEY WA AUSTRALIA [email protected] Ms Kirsty Pringle Level 6, Medical School North Frome Rd ADELAIDE SA AUSTRALIA [email protected]

Dr Megan Probyn Research Officer PO Box 13F MONASH UNIVERSITY VIC AUSTRALIA [email protected] Professor J. Usha Raj Professor Of Pediatrics, Chief, Div of Neonatology RB-1, 1124 W. Carson Street TORRANCE CA USA [email protected] Dr Manindra Raj Training Department Baudha, Timchule PO Box 1144 KATHMANDU NEPAL [email protected] Neesha Rao Graduate Student St. Joseph's Health Centre, Dept. Of Obstetrics And Gynecology 268 Grosvenor Street LONDON ONTARIO CANADA [email protected] Dr Juha Rasanen Kajaanintie 50 OULU FINLAND [email protected] Dr Bryan Richardson Professor St Joseph Health Care 268 Grosvenor Street LONDON ONTARIO CANADA [email protected] Dr Claire Roberts Level 6, Medical School North Frome Rd ADELAIDE SA AUSTRALIA [email protected]

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A/Prof Sarah Robertson NHMRC Senior Research Fellow University of Adelaide Frome Road ADELAIDE SA AUSTRALIA [email protected] Prof Jeffrey Robinson Professor And Head Of Department Of Obstetrics University Of Adelaide ADELAIDE SA AUSTRALIA [email protected] Dr. Matthias Schwab Dept. Of Neurology JENA GERMANY [email protected] Miss Amanda Sferruzzi-Perri 6th Floor, Medical School North Frome Rd ADELAIDE SA AUSTRALIA [email protected] Dr Penelope Sheehan Obstetrician C/o Pregnancy Research Centre Royal Womens Hospital 132 Grattan Street CARLTON VIC AUSTRALIA [email protected] Dr Andrew Siebel Research Officer Royal Pde PARKVILLE VIC AUSTRALIA [email protected]

Ms Reetu Singh PhD Student Wellington Rd CLAYTON VIC AUSTRALIA [email protected] Dr Deborah Sloboda Research Fellow King Edward Memorial Hospital Bagot Road SUBIACO WA AUSTRALIA [email protected] Prof Roger Smith Director HMRI, Level J3, JHH Locked Bag 1, HRMC NEWCASTLE NSW AUSTRALIA [email protected] Mr Timothy Spencer Student (Honours) ROSE PARK SA AUSTRALIA [email protected] Dr Jacqueline Stack Staff Specialist - Neonatology Liverpool Bc NSW AUSTRALIA [email protected] Dr Michael Stark Clinical Research Fellow Hunter Medical Research Institute John Hunter Hospital NEWCASTLE NSW AUSTRALIA [email protected]

Dr Junichi Sugawara Obstetrics & Gynaecology 1-1 Seiryomachi Aobaku SENDAI JAPAN Miss Brooke Summers PhD Student Frome Road Clinical Biochemistry ADELAIDE SA AUSTRALIA [email protected] Prof Michael Symonds Academic Division Of Child Health E Floor, East Block University Hospital NOTTINGHAM UNITED KINGDOM [email protected] Ms Nicolette Hodyl Doctoral Student Department Of Psychology, School Of Behavioural Sciences University Of Newcastle CALLAGHAN NSW AUSTRALIA [email protected] Dr Marianne Tare Senior Research Officer Department Of Physiology PO Box 13F Monash University CLAYTON VIC AUSTRALIA [email protected] Dr Joseph Thomas Perinatal Medicine 72 King William Road NORTH ADELAIDE SA AUSTRALIA [email protected]

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Dr David Todd Westmead Hospital Hawkesbury Road WESTMEAD NSW AUSTRALIA [email protected] Ms Melanie Tran B Sc Honours Student Department Of Obstetrics University Of Adelaide ADELAIDE SA AUSTRALIA [email protected] Dr Mark Vickers Research Fellow 2-6 Park Avenue Grafton AUCKLAND NEW ZEALAND [email protected] Assoc Prof David Walker Principal Research Fellow Department Of Physiology Wellington Road CLAYTON VIC AUSTRALIA [email protected] Dr Megan Wallace Senior Research Officer Department Of Physiology P.O. Box 13F MONASH UNIVERSITY VIC AUSTRALIA [email protected] Ms Kerryn Westcott Senior Research Assistant Royal Pde PARKVILLE VIC AUSTRALIA [email protected]

Prof E.Marelyn Wintour-Coghlan Honorary Professor Monash University Wellington Road CLAYTON VIC AUSTRALIA [email protected] Assoc Prof Mary Wlodek Associate Professor Royal Pde PARKVILLE VIC AUSTRALIA [email protected] Prof Charles Wood Professor And Chair Of Physiology And Functional Genomics PO Box 100274 GAINESVILLE FLORIDA USA [email protected] Dr Ian Wright Staff Specialist Lookout Road NEW LAMBTON HEIGHTS NSW AUSTRALIA [email protected] Prof Chittaranjan Yajnik Director, Diabetes Unit Rasta Peth PUNE INDIA [email protected] Mr Edwin Yan Department Of Physiology Wellington Road CLAYTON VIC AUSTRALIA [email protected]

Ms Tamara Yawno PhD Student Building 13F Wellington Road CLAYTON VIC AUSTRALIA [email protected]

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List of Presenting Authors

Allison, BA ………. 28, 53

Bjorkman, ST ………. 27, 106

Black, MJ ………. 25, 34, 68, 95

Bloomfield, FH ………. 34, 69

Botting, K ………. 23, 29, 86

Bromfield, J ………. 38, 121

Brown, RD ………. 21, 46

Budge, H ………. 23, 29, 34, 70, 83, 127

Cassaglia, PA ………. 22, 50

Challis, J ………. 26, 39, 103

Chua, JSC …….… 36, 80

Chuang, BHA …….… 23, 29, 84

Cosmi, E …….… 26, 103

Cosmi, EV …….… 23, 29, 84

Coulter, CL …….… 21, 43

Dalitz, PA …….… 24, 29, 32, 65, 88

De Blasio MJ …….… 35, 75

Dean, JM …….… 24, 26, 30, 89, 101

Dickinson, H …….… 26, 35, 73, 99

Dunn, SL …….… 26, 104

Feng, YS …….… 32, 64

Fletcher CJ …….… 27, 108

Forhead, AJ …….… 35, 75

Franko, KL …….… 23, 28, 83

Fraser, M …….… 24, 30, 88

Gatford, KL …….… 36, 80

Gibson, KJ …….… 21, 45

Gluckman, PD …….… 39

Grover, S …….… 27, 107

Gubhaju L …….… 21, 46

Hamrahi, H …….… 32, 63

Harding, J …….… 39, 123, 124

Harnett, EL …….… 24, 30, 89

Hartgill, TW ………. 25, 97

Henderson-Smart, D….. 37, 114, 115

Hill, D ………. 39, 125, 126

Hodyl, NA ………. 36, 78

Hooper, SB ………. 28, 55

Ikdea, T ………. 31, 61

Imai, N ………. 23, 29, 85

Jensen, EC ………. 21, 23, 29, 43, 87

Kelley, RL ………. 26, 100

Kind, KL ………. 31, 58

Knight, B ………. 34, 70

Koos, BJ ………. 31, 61

Ligam, P ………. 28, 55

Lumbers, ER ………. 21, 39, 44

MacLaughlin, SM….….34, 69

Maduwegedera, D……..22, 48

Mallard, C …….… 32, 63

Malpas, SC …….… 22, 51

McCallum, J …….… 32, 66

Mibus, AL …….… 27, 108

Moore, LK …….… 25, 96

Moritz, K ….…… 21, 45

Morrison, JL ….…… 31, 59

Moss, TJM ….…… 28, 54

Mostyn, A ….…… 24, 94

Muhlhausler, B ….…… 39, 128

Myatt, L ….…… 38, 119, 120

Newhouse, S ….…… 27, 105

Nijhuis, JG ….…… 32, 66

Nishigori, H ….…… 31, 60

Nitsos, I ….…… 31, 59

Noorlander, CW…….…24, 93

O’Connell, AE …….… 21, 94

O’Connor, DM …….… 25, 98

O’Dowd, R …….… 34, 71

Okamura, K …….… 26, 102

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Olson, DM …….… 34, 68

Padbury, JF ………. 22,23,24,29,50,86,93

Painter, P ………. 38, 116

Pakkila, M ………. 22, 49

Parange, N ………. 27, 104

Parer, JT ………. 24, 26, 30, 90, 102

Patrick, L ………. 32, 64

Peebles, D ………. 25, 32, 65, 98

Polglase, GR ………. 28, 35, 54, 74

Pringle, KG ………. 26, 100

Probyn, M ………. 25, 95

Rao, N ………. 27, 106

Rasanen, J ………. 22, 51

Roberts, CT …….….27, 105

Robinson, JS …….… 36, 37, 110, 111

Schwab, M …….… 25,35,38,76,99,116

Sferruzzi-Perri, AN…....31, 38, 58, 121

Sheehan PM ….…… 28, 56

Singh, RR ….…… 35, 76

Sloboda, DM ….…… 35, 74

Smith, R ………. 38, 117, 118

Spencer, T ………. 26, 101

Stark, MJ ………. 22, 48

Summers, BL ………. 36, 81

Symonds, ME ………. 25,35,36,73,79,94,96

Tare, M ………. 22, 38, 49, 122

Thakor, AS ………. 24, 92

Tran, M ………. 36, 79

Usha Raj, J ………. 31, 60

Vickers, MH ………. 36, 78

Wallace, M ………. 39, 127

Westcott, KT ………. 27, 107

Wood, CE ………. 28, 53

Wright, IMR ………. 23, 29, 85

Yajnik, R ………. 37, 112, 113

Yan, EB ………. 25, 97

Yawno, T ………. 23, 29, 87

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The 32nd Annual Meeting of the Fetal and Neonatal Physiological Society 2005 September 25-28

Stamford Grand Hotel Glenelg South Australia, Australia

Cover image supplied by the South Australian Tourism Commission