The long term goals of our laboratory are to understand the molecular mechanisms responsible for cell fate specification and sensory patch formation in the developing mammalian inner ear. Progress toward understanding these essential aspects of inner ear development have been hampered by the inaccessibility of the mouse embryo in utero. Equally confounding is the lack of molecular tools to dynamically manipulate developmental gene expression in situ. We apply experimental embryology to study the mouse inner ear in vitro and in vivo.
We aim : 1) to fate map mouse otic cup closure in wild type and mutant mice;2) to determine the lineage relationships among constituent cells in the inner ear;and 3) to probe the molecular mechanisms underlying mammalian sensory patch formation. Mouse whole embryo culture supports inner ear development from the placode through early otocyst stages. The fate map will be conducted by iontophoretic injection of fluorescent tracer dye into designated positions in the rim and concavity of the cup. Fate mapping will teach us the morphogenetic movements of otic epithelial progenitors during cup closure and permit their correlation with know domains of gene expression. Tranuterine microinjection of bioactive reagents into the early otocyst stage mouse embryo in vivo enables a broad range of studies in the developing and postnatal inner ear. Lineage analysis will be performed by transuterine microinjection of a retroviral construct encoding alkaline phosphatase and a complex 24 base pair library into the otocyst with subsequent clonal analysis in the mature, postnatal inner ear. Lineage analysis will show us the types and timing of cell fate choices made by otic epithelial progenitors that give rise to sensory and nonsensory cells. Sensory patch formation will be investigated by transuterine microinjection of expression plasmid into the early otocyst followed by in vivo electroporation for gain-of-function studies. We will misexpress transcription factors know to be involved in sensory patch formation in wild type and mutant mouse inner ears to gain insight into their mechanistic roles in establishing the auditory sensory epithelium. A clear understanding of otic vesicle morphogenesis, lineage relationships, and sensory patch formation is essential for the definition of regenerative medical approaches to ameliorate hearing loss and balance disorders in humans.

Agency
National Institute of Health (NIH)
Institute
National Institute on Deafness and Other Communication Disorders (NIDCD)
Type
Research Project (R01)
Project #
3R01DC008595-03S2
Application #
7857466
Study Section
Auditory System Study Section (AUD)
Program Officer
Freeman, Nancy
Project Start
2009-07-17
Project End
2011-12-31
Budget Start
2009-07-17
Budget End
2011-12-31
Support Year
3
Fiscal Year
2009
Total Cost
$183,285
Indirect Cost
Name
Oregon Health and Science University
Department
Otolaryngology
Type
Schools of Medicine
DUNS #
096997515
City
Portland
State
OR
Country
United States
Zip Code
97239
Gomes, Michelle M; Wang, Lingyan; Jiang, Han et al. (2016) A Rapid, Cost-Effective Method to Prepare Recombinant Adeno-Associated Virus for Efficient Gene Transfer to the Developing Mouse Inner Ear. Methods Mol Biol 1427:43-57
Jiang, Han; Wang, Lingyan; Beier, Kevin T et al. (2013) Lineage analysis of the late otocyst stage mouse inner ear by transuterine microinjection of a retroviral vector encoding alkaline phosphatase and an oligonucleotide library. PLoS One 8:e69314
Pokidysheva, Elena; Tufa, Sara; Bresee, Chris et al. (2013) Prolyl 3-hydroxylase-1 null mice exhibit hearing impairment and abnormal morphology of the middle ear bone joints. Matrix Biol 32:39-44
Huang, Alice H; Riordan, Timothy J; Wang, Lingyan et al. (2013) Repositioning forelimb superficialis muscles: tendon attachment and muscle activity enable active relocation of functional myofibers. Dev Cell 26:544-51
Giese, Arnaud P; Ezan, Jérome; Wang, Lingyan et al. (2012) Gipc1 has a dual role in Vangl2 trafficking and hair bundle integrity in the inner ear. Development 139:3775-85
Wang, Lingyan; Jiang, Han; Brigande, John V (2012) Gene transfer to the developing mouse inner ear by in vivo electroporation. J Vis Exp :
Wang, Lingyan; Bresee, Chris S; Jiang, Han et al. (2011) Scleraxis is required for differentiation of the stapedius and tensor tympani tendons of the middle ear. J Assoc Res Otolaryngol 12:407-21
Reisinger, Ellen; Bresee, Chris; Neef, Jakob et al. (2011) Probing the functional equivalence of otoferlin and synaptotagmin 1 in exocytosis. J Neurosci 31:4886-95
Brigande, John V; Gubbels, Samuel P; Woessner, David W et al. (2009) Electroporation-mediated gene transfer to the developing mouse inner ear. Methods Mol Biol 493:125-39
Brigande, John V; Heller, Stefan (2009) Quo vadis, hair cell regeneration? Nat Neurosci 12:679-85

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