The majority of hearing and balance disorders occur as the result of death to mechanosensory hair cells of the inner ear. A number of environmental factors contribute to hair cell death in the human population, including age, noise exposure history, and exposure to ototoxic drugs. There persist fundamental gaps in our understanding of the intracellular mechanisms governing these processes. The long term objectives of the proposed research are two-fold: 1) to obtain a mechanistic understanding of the events underlying hair cell death due to aminoglycoside (AG) antibiotic exposure, and 2) to identify therapeutic approaches to prevent it. This project focuses on hair cell death caused by exposure to AG antibiotics, effective against a number of systemic infections yet responsible for hair cell death within a significant portion of individuals receiving them. I recently identified that exposue to AG antibiotics induces disruption of calcium homeostasis between the two largest calcium stores within the cell - the endoplasmic reticulum (ER) and mitochondria - which transfer calcium between each other to regulate cellular bioenergetics. My preliminary data in zebrafish have identified an upstream factor responsible for regulating this process - the Sigma Receptor (SigR1) - during AG-induced hair cell death. FDA-approved compounds that regulate SigR1 protect zebrafish hair cells against AG toxicity. This research plan will test the central hypothess that SigR1 facilitates the disruption of calcium homeostasis in the ER and mitochondria during AG toxicity. Guided by preliminary data, this hypothesis will be tested by pursuing two specific aims: 1) determine the proteins with which SigR1 associates during AG-induced hair cell death and how association with these proteins affects ER-mitochondrial calcium transfer; and 2) assess the protective value of SigR1 modulation on mammalian ototoxicity. To address these aims, I will combine genetic, pharmacological, and imaging techniques to address how SigR1 modulates ER and mitochondrial calcium homeostasis during AG-induced hair cell death, and seek to evaluate the feasibility of SigR1 as a therapeutic target intended to ameliorate or entirel prevent AG ototoxicity. The approach seeks to delve upstream of calcium homeostasis to understand how its disruption comes about in the zebrafish model system, and will explore the importance of this pathway in mammals. I will evaluate the effectiveness of FDA-approved SigR1 modulators at mitigating toxic effects of AG exposure in mammals, potentially establishing their use to stem hearing loss and balance disruption in the human population.