Objectives: Sensorineural hearing loss (SNHL) is strongly associated with many aspects of military service including blast injury. The overall objectives of this proposal are to improve the prevention and treatment of SNHL in Veterans. Research Design: We previously used in vitro screening to identify novel compounds that can protect hair cells (HCs) from ototoxic damage. We tested them in vivo against noise-induced hearing loss (NIHL) and found partial protection. We also used high-resolution proteomics to identify additional processes involved in NIHL. Overall, our results suggest that many cellular processes contribute to HC damage. In this application we propose to screen compound combinations targeting diverse HC damage and survival processes, to identify the most effective combinations. This will allow us to identify optimal strategies for further development as pharmacological interventions in humans, using compound combinations and/or multi-acting compounds. Methodology: Studies will be performed in vitro using aminoglycoside damage or in vivo using noise damage to the cochlea. For in vivo studies, intracochlear or intratympanic delivery of HC protectants will begin immediately after exposure. We will use a well-established mouse model of noise damage, chronic delivery of compounds to cochlear perilymph or the round window with osmotic minipumps, serial auditory brainstem response (ABR) and distortion product otoacoustic emission (DPOAE) audiometry, and morphological evaluation of cochlear HCs and afferent nerve endings. Progress over the past period of funding: During the past period of funding, using the mammalian organ of Corti, the only tissue containing the damage-sensitive mammalian outer HCs, we screened several compound libraries. We identified a broad range of compounds targeting different cellular processes, that were effective in protecting HCs. We then transferred selected compounds to an in vivo model of noise-induced hearing loss (NIHL), where we also noted protection. We also completed the first-ever high-resolution proteomics study of NIHL, which identified many proteins and processes not previously known to be involved in noise damage, and which are potential targets for pharmacotherapy. We also performed the first high-resolution proteomic study of isolated HCs. In addition, we completed studies of gene regulation relevant to HC regeneration, and found that gene therapy with espin1 dramatically enhanced stereocilia formation on regenerating HCs. We discovered a novel protective HC pathway mediated by ATP receptors that reduce activity in HCs at high stimulus levels, protecting them from noise damage. Our studies resulted in 18 peer-reviewed publications to date. Clinical Relationship: The prevention and treatment of SNHL is of great importance to Veterans and the VA. The effects of SNHL on Veterans’ quality of life are substantial. SNHL and tinnitus also account for more disability compensation in the VA than any other d...