Neural Stem Cells in the Otocyst

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DESCRIPTION (provided by applicant): During development, cells lining on the rostro-ventral region of the otic epithelium acquire their identity as neuron precursors and migrate out of the otocyst to form the VIII (vestibulocochlear) ganglion. Little is known about the molecular mechanisms involved in the induction of VIII ganglion neurons. The primary goals of this study are to identify molecular factors that have neurogenetic effects on otocyst cells in vitro, and also to verify the physiological function of these factors during the course of neural induction in vivo. The long-term goal of this study is to delineate molecular mechanisms by which undifferentiated cells in the otocyst become competent to commit to neuronal cells, and subsequently differentiate into cochlear or vestibular ganglion neurons. The following two specific aims will be addressed: (1) to identify soluble proteins that have neurogenetic effects on undifferentiated otocyst cells. (2) To establish an in vivo model system to manipulate gene expression in undifferentiated otocyst cells. For Aim #1, primary otocyst cell cultures will be established and the neurogenetic effects of FGF2, FGF8, FGF10, TGFb2, BMP4, Noggin or retinoic acid, alone or in combination, on dissociated otocyst cells will be tested in vitro. For Aim #2, an in vivo gene delivery method using electroporation will be established and Phox2b will be overexpressed in the otocyst of live embryos before the formation of the VIII ganglion. The effects of Phox2b overexpression on cell proliferation and gene expression in otocyst cells will be tested in vivo. These experiments represent the first step in a systematic attempt to generate vestibular and cochlear ganglion neurons from otocyst stem cells to replace damaged or dead neurons. Furthermore, knowledge of the factors involved in neural induction and specification in the inner ear would advance our understanding of the cause of genetically inherited deafness or balance disorders.