““Biological Bases of AgeBiological Bases of Age-- Related Hearing LossRelated Hearing Loss””

Robert D. Robert D. FrisinaFrisina, PhD, PhD Professor and Assoc. ChairProfessor and Assoc. Chair

Otolaryngology DepartmentOtolaryngology Department University of Rochester Medical School,University of Rochester Medical School,

International Center for Hearing & Speech ResearchInternational Center for Hearing & Speech Research National Technical Institute for the DeafNational Technical Institute for the Deaf

Morning Session: Morning Session: PresbycusisPresbycusis –– What What Goes Wrong in the Ear and Brain?Goes Wrong in the Ear and Brain?

Why donWhy don’’t hearing aids work with many aged t hearing aids work with many aged listeners?listeners?Is it all about hair cell loss? Is it all about hair cell loss? –– No, butNo, but……Timing is everything! Timing is everything! ––Feedback loop from the brain to the ear Feedback loop from the brain to the ear declines, starting in middle agedeclines, starting in middle agePromising bioengineering avenues for Promising bioengineering avenues for prevention and therapeutics!prevention and therapeutics!

Gene Expression Experiment: Gene Expression Experiment: The Central ThemeThe Central Theme

The arrows represent the transfer or flow of The arrows represent the transfer or flow of information.information.DNA and RNA store information in a baseDNA and RNA store information in a base--4 code 4 code (the four nucleotides).(the four nucleotides).Proteins store information in a baseProteins store information in a base--20 code (the 20 20 code (the 20 amino acids). amino acids).

DNA mRNA ProteinTranscription Translation

Experimental Design For Experimental Design For GeneChipGeneChip StudyStudy

Hypothesis: Hypothesis: Gene expression changes in the ear and the brain Gene expression changes in the ear and the brain occur in occur in presbycusispresbycusis

Animal Model:Animal Model: CBA Mice CBA Mice –– slow progressive hearing lossslow progressive hearing loss

Tissue :Tissue : Cochlear and inferior Cochlear and inferior colliculuscolliculus (auditory midbrain) (auditory midbrain) RNA samples from individual mice on RNA samples from individual mice on individual individual microarraysmicroarrays

Investigative Tool:Investigative Tool: AffymetrixAffymetrix GeneChipGeneChip, one chip analyzes , one chip analyzes 22,600 gene probes for each sample, from each mouse22,600 gene probes for each sample, from each mouse

Project Strengths:Project Strengths: Number of replicates, N=80, strengthened the Number of replicates, N=80, strengthened the statistical analysis. One chipstatistical analysis. One chip--one mouse allows exploration of the one mouse allows exploration of the biological phenotype variance from mouse to mouse.biological phenotype variance from mouse to mouse.

PresbycusisPresbycusis MicroarrayMicroarray --Animal Animal Subject Group DetailsSubject Group Details

Groups of MiceGroups of Mice No. of MiceNo. of Mice No. of Chips,No. of Chips,1 chip/1 chip/mousemouse

Age Age -- MonthsMonths GenderGender

Young ControlYoung Control 99 99 3.5 3.5 ±± 0.40.4 Male=5Male=5Female=4Female=4

Middle aged Middle aged Good HearingGood Hearing

1717 1717 12.3 12.3 ±± 1.51.5 Male=8Male=8Female=9Female=9

Old Old -- Mild Mild PresbycusisPresbycusis

99 99 27.7 27.7 ±± 3.43.4 Male=4Male=4Female=5Female=5

Old Old -- Severe Severe PresbycusisPresbycusis

66 66 30.6 30.6 ±± 1.91.9 Male=2Male=2Female=4Female=4

C B A M o u s e Au d ito ry B ra in s te m Re s p o n s e

Au d io g ra m

0 1 0 2 0 3 0 4 0 5 00

1 02 03 04 05 06 07 08 09 0

1 0 0Yo u n g GH*

M id d le -A g e d GH*

Old M P*

Old SP*

* GH: Good He a ring M P: M ild Pre s byc us is SP: Se ve re Pre s byc us isError Ba r: Sta nda rd Error M e a n

Frequency (kH z )

Th

resh

old

(d

B S

PL

)

Auditory Brainstem Response (ABR) recordings for the CBA mice in the microarray experiments.. -Young adult and middle aged mice show good hearing- Whereas the auditory sensitivity declined in old age. -Note that the largest change was from the middle aged to the old presbycusis groups.

Functional Anatomy and Ion Channel Exchange in the Mammalian Inner Ear – Cochlear Tissue

For Gene Arrays-Organ of Corti-Lateral Wall

Gene Microarray Findings

GABA – Important Inhibitory Neurotransmitter of the Efferent Feedback System from the Central Auditory System to the Cochlea:

Key Cochlear GABA Receptor Declines with Age – Starting in Middle age, like the Efferent System

From: D’Souza et al., J. Neuroscience Methods, 2008

Atf3 - activating transcription factor3

Bcl2 - B-cell leukemia/lymphoma2

Bcl2l1 - Bcl2-like1

Casp4 - caspase4 apoptosis-related cysteine protease 4

Cochlear Apoptotic Pathways Show Up- Regulation with Age and Hearing lossApoptosis Apoptosis –– Programmed Cell DeathProgrammed Cell Death

From: Tadros et al., Apoptosis, 2008

CentralImpairment

Perception of speech and complex Perception of speech and complex sounds in cortical centerssounds in cortical centers

BrainstemAuditorySystem

PeripheralAuditorySystem

Peripherally - Induced central changesInsult-Age

Peripheral Hearing

Loss

Summary of Central Auditory System Changes with AgeSummary of Central Auditory System Changes with Age

From: Frisina et al., Functional Neurobiology of Aging, 2001

Glutamate – The Primary Excitatory Neurotransmitter of the Auditory System

Pycs plays a role in converting glutamate to proline- Its deficiency in old age may lead to:- Glutamate increases and proline deficiencies in the auditory midbrain - Playing a role in the subsequent inducement of glutamate toxicity and loss of proline neuroprotective effects

From: Tadros et al., Brain Research, 2007

Slc1a3 is a glutamate transporter- Gene expression changes with age and hearing loss may reflect a cellular compensatory mechanism to protect against age-related glutamate or calcium excitoxicity

Upregulation of Serotonin Receptors with Age and Hearing Loss in the Inferior Colliculus – Auditory Midbrain

Gene Expression

Protein Expression

From: Tadros et al., Neurobiology of Aging, 2007

Upregulation of Serotonin Receptors with Age and Hearing Loss in the Inferior Colliculus – Auditory Midbrain-Could help compensate for declines in Serotonin with age

- Could result in age- related Ca++ toxicity by increasing the intracellular concentration of IP3

From: Tadros et al., Neurobiology of Aging, 2007

From: Zettel et al., Hearing Res. 2001

- Compensatory up- regulation of calretinin

Genetically Cross the CBA and C57 Mouse Strains: Discovered a new mouse model for aged human listeners who have audiograms within the normal hearing range: mice with “Golden Ears”

From: Frisina et al., Neurobiology of Aging, On Line

Genetically Cross the CBA and C57 Mouse Strains: Discovered a new mouse model for aged human listeners who have audiograms within the normal hearing range: mice with “Golden Ears”

From: Frisina et al., Neurobiology of Aging, On Line

Work Supported by NIH:National Institute on Aging,

Sensory and Motor Disorders of Aging,Behavioral & Systems Neuroscience Branch,

Division of NeuroscienceNational Institute on Deafness &

Communication Disorders

Rochester Hearing/Deafness Research GroupRochester Hearing/Deafness Research Group

Otolaryngology Otolaryngology -- U. RochesterU. RochesterDr. Kathy Dr. Kathy BarszBarsz ––NeurophysiolNeurophysiol..Dr. Owen Dr. Owen BrimijoinBrimijoin -- PhysiologyPhysiologyDr. Mary Dr. Mary D'SouzaD'Souza –– Molecular Molecular Biology, Gene Biology, Gene MicroarraysMicroarraysSusan Susan FrisinaFrisina, RN, RN –– Med. GeneticsMed. GeneticsDr. Robert Dr. Robert FrisinaFrisina --NeuroscienceNeuroscienceDr. Patricia Dr. Patricia GuimaraesGuimaraes –– Hormonal Hormonal Effects on AuditionEffects on AuditionDr. UDr. U--Cheng LeongCheng Leong -- PhysiologyPhysiologyDr. Olga Dr. Olga VasilyevaVasilyeva –– PharmacolPharmacol..Dr. Joseph WaltonDr. Joseph Walton ––Auditory Auditory NeurophysiologyNeurophysiologyMartha Martha ZettelZettel, MS, MS ––ImmunocytochemistryImmunocytochemistryDr. Dr. XiaoxiaXiaoxia ZhuZhu –– Emissions, Emissions, ABRsABRs, Micro, Micro--SurgerySurgery

University of RochesterUniversity of RochesterDr. Paul AllenDr. Paul Allen -- Behavior, Behavior, NeurophysiologyNeurophysiologyJohn HouselJohn Housel -- Animal CoreAnimal CoreDr. James Dr. James IsonIson -- Animal BehaviorAnimal BehaviorDr. William ODr. William O’’NeillNeill –– ABRsABRs, , Auditory NeuroscienceAuditory Neuroscience

Rochester Institute TechnologyRochester Institute TechnologyDr. Robert Dr. Robert FrisinaFrisina, Sr, Sr. . –– Speech Speech Perception, PET, Perception, PET, AudiologyAudiologyDr. David Dr. David BorkholderBorkholder –– MicroMicro--Systems BioengineeringSystems BioengineeringDr. David Dr. David EddinsEddins -- PsychoacousticsPsychoacousticsFray Fray MapesMapes, MA, MA –– Res. Res. AudiologyAudiologyDr. Dina NewmanDr. Dina Newman –– GeneticsGenetics