© Institute of Veterinary Physiology Chart 1

Erythropoietin (Epo) regulates hypoxic ventilation by interacting with brainstem and carotid bodies

University of ZürichUniversité Claude Bernard Lyon1

© Institute of Veterinary Physiology Chart 2

O2

CO2

Diaphragm

Ven

tilat

ion

Breathing:

Carotid body

Brainstem

Cen

tral

nerv

ous

syst

emP

erip

hera

lne

rvou

s sy

stem

© Institute of Veterinary Physiology Chart 3

Ventilation underHYPOXIA (diminished PpO2)

PaO2

© Institute of Veterinary Physiology Chart 4

Molecules controlling the ventilation:

Epotarget genes

Hypoxia-activatedMolecules

(HIF-1)

NeurotransmittersAmines

Steroids

(Pascual, 2004)

(Kline, 2002)

© Institute of Veterinary Physiology Chart 5

Epo Epo expression occurs in:expression occurs in:

brain

testis

lung

kidney

liver

but also in other organs including:

© Institute of Veterinary Physiology Chart 6

3 “anti” roles of Epo:

anti-apoptotic

anti-cytotoxic

anti-oxidative

(Tann et al 1992; Masuda et al. 1993; Morishita et al 1997; Koshimura et al. 1999; Tanaka et al. 2001)

© Institute of Veterinary Physiology Chart 7

Epo and Epo receptor (Epo and Epo receptor (EpoREpoR) in brain) in brain

Epo &EpoR

are expressedin the

mammalian andhuman brain

are expressedin the

mammalian andhuman brain

cerebral Epo is induced by hypoxia

cerebral Epo is induced by hypoxia

are expressed by neurons

and astrocytes

are expressed by neurons

and astrocytes

© Institute of Veterinary Physiology Chart 8

Does brain-derived Epo

play a role in the control of ventilation under

hypoxia?

© Institute of Veterinary Physiology Chart 9

- Overexpress Epo ONLY in brain (4 x/Wt)- NORMAL Epo in plasma; HB ; Hct

Transgenic Tg21

Normoxiabasal Ve

20 min

Hypoxia 10%O2

20 min

Hypoxia 6%O215

min

15 m

in

© Institute of Veterinary Physiology Chart 10

Tg21 maintain high ventilation under at 6% hypoxia

∗ ∗ ∗ ∗ ∗

80

105

130

155

180

205

230

VE

(ml/m

in/1

00g)

exposure time (min)

Tg21WT

21% O2 10% O2 6% O2

© Institute of Veterinary Physiology Chart 11

Tg21 enhance ventilation after chronic hypoxia

• hypoxic chamber• 3 days at 10% O2

basal Ve

20 min

Hypoxia 10%O2

20 min

Hypoxia 6%O2

© Institute of Veterinary Physiology Chart 12

EpoR is expressed in the resp.areas of the brainstem

Legends:

4V, 4th ventricle; AP, area postrema; NTS, nucleus tractus solitarius; 7n, facial nerve; PBC, Pre-Bötzinguercomplex; NA, nucleus ambiguus; PY, pyramidal decussation; LC, locus ceruleus. Catecholaminergic areas in the medulla oblongata: A1C1 & A2C2 and in the pons: A5 & A6

© Institute of Veterinary Physiology Chart 13

Chemodenervation is life-threatening in Wtbut not in Tg21

Chemodenervation

© Institute of Veterinary Physiology Chart 14

Is the Epo impact on ventilation a

neuro-protectivephenomenon?

© Institute of Veterinary Physiology Chart 15

0

50

100

150

200

1 2

Activated STAT-5 35 kDa-

Perc

ent o

f con

trol

wt tg21

020406080

100120140

1 2

Activated ERK-1 44 kDa-

Perc

ent o

f con

trol

wt tg210

20406080

100120

1 2

Activated ERK-2

42 kDa-

Perc

ent o

f con

trol

wt tg21

0

15

30

45

60

1 2

JAK-2 45 kDa-

wt tg21

Perc

ent o

f con

trol

STAT5

JNK

PKBMAPK

AktERk1 ERk2

020406080

100120

1

46 kDa-

*P

erce

nt o

f con

trol

wt tg21

020406080

100120

1 2

*

62 kDa-

Perc

ent o

f con

trol

wt tg21

© Institute of Veterinary Physiology Chart 16

Are Catecholamines

involved in the impactof Epo

in ventilation?

© Institute of Veterinary Physiology Chart 17

Catecholaminergic group cells in the brainstem

© Institute of Veterinary Physiology Chart 18

WT Tg21

A1C

1A

2C2

A5

A6

© Institute of Veterinary Physiology Chart 19

pmol

esN

A/20

min

Nor

adre

nalin

e

0

10

20

30 *

Wt Tg

pmol

esD

OPA

/20m

in

pmol

esN

E

TH

act

ivity

NE

con

tent

A2C2 cell group

4

6

8

10

WT Tg21

*

10

15

20

25

WT Tg21

*

© Institute of Veterinary Physiology Chart 20

does plasma-derived Epo

impact hypoxic ventilation via CB?

© Institute of Veterinary Physiology Chart 21

TH

ECAICA

CCA

TH

EpoR is expressed in Carotid bodies glomic cells

Epo-REpo-R

ECAICA

CCA

Legends:

TH, tyrosine hydroxilase; Epo-R, erythropoietin receptor; ICA, internal carotid artery; ECA, external carotid artery; CCA, commun carotid artery

© Institute of Veterinary Physiology Chart 22

Epo injected (i.v.) in Wt mice modulates the hypoxic ventilatory pattern

Injection of Epo (iv)(2000 U/Kg: 50 U/mouse) Wt C57/Bl6

Epo does notcross the BBB

VE = RR x VT

© Institute of Veterinary Physiology Chart 23

Carotid body slices: Wistar rats (2 wk old)

Epo triplicates the CB evoked response under hypoxia

500 U/Epo

x 3

0 U/Epo

hypoxia

hypoxia

inclusion

slicing

culturingAmperometry

Spain: Lopez-Barneo

© Institute of Veterinary Physiology Chart 24

Conclusion

Brain-derived Epo (via EpoR in brainstem) and systemic Epo (via EpoR in carotid bodies) modulate ventilation under hypoxia

These results suggest that Epo has crucial role in the fine-tuning of oxygen homeostasis

High altitude dwellers?

Acute and Chronic mountain sickness?

Lance Amstrong…Epo doping?

© Institute of Veterinary Physiology Chart 25

C Soulage C BecskeiL Ogunshola J VogelT GorrS KellerD Furrer

Acknowledgements

Prof. M GassmannProf. V JosephProf. JM Pequignot

© Institute of Veterinary Physiology Chart 26

Is the Epo effect on ventilation a selective Is the Epo effect on ventilation a selective action?action?

Epo mediates general integrity to the tissue

retinal protection

brain injury(stroke)spinalcord

injury

heartinfarct

development?

gonads?

© Institute of Veterinary Physiology Chart 27

Hello, hello,…. how much oxygen do you have over there?