PROJECT SUMMARY Mechanosensory hair cell loss in the inner ear is a leading cause of hearing and balance disorders. Unfortunately hair cell loss is permanent as humans have little or no capacity for regenerative recovery of hair cells. Although there has been recent progress in the restoration of hair cells in mammals by promoting expression of genes and pathways used during development, functional restoration is largely incomplete and within a limited window of postnatal development. By contrast non-mammalian species from fish to birds readily replace hair cells from surrounding supporting cell precursors. Critical to understanding hair cell regeneration is the identification of the subpopulations of supporting cells that sustain regeneration. The zebrafish is well established as a system to study regeneration with the advantages of advanced genetics and in vivo imaging of living animals. Considerable progress has been made on understanding regeneration in the lateral line system, the series of mechanosensory hair cells on the surface of the body used detect water flow that share many properties with hair cells of the inner ear. By contrast there have been very few studies of hair cell regeneration in the zebrafish inner ear, mainly because there has been no reliable method for causing rapid and reproducible damage. In addition little is known about supporting cell types in the zebrafish inner ear. Overcoming both of these obstacles would establish the zebrafish inner ear as a new model for hair cell regeneration. We propose to develop tools for rapid, reproducible damage and regeneration in the zebrafish inner ear. We also propose to identify inner ear supporting cell types, including potential hair cell precursors. Successful completion of these aims will provide important tools for the community to begin new studies in zebrafish inner ear regeneration.