Project Summary Sensorineural hearing loss, the most common form of hearing loss, is characterized by damage to inner ear sensory hair cells or to the spiral ganglion neurons (SGNs) that conduct auditory information from the hair cells to the brain. SGN death can occur because of direct trauma, e.g., acoustic overexposure, or secondarily after hair cell loss. Degeneration of SGNs can reduce the efficacy of cochlear implants that replace the function of hair cells by electrically stimulating SGNs. The mechanisms that promote SGN death after hair cell loss are unknown, however, mounting evidence suggests a causal role of the immune response. Neonatal rats treated with the ototoxic aminoglycoside antibiotic kanamycin lose all hair cells by 3 weeks of age, rendering them deaf, and subsequently lose more than 80% of SGNs by 17 weeks of age. Gene expression profiling analyses from the Green lab indicate that post-deafening, there is a significant increase in the expression of immune response related genes in the spiral ganglion. This is accompanied by a significant increase in macrophage number and activation up to two weeks prior to the start of significant post-deafening SGN death. In addition, lymphocytes (CD4+ helper T cells, CD8+ cytotoxic T cells, and CD161+ NK cells) increase in number within the spiral ganglion concurrent with the first indication of significant post-deafening SGN death. This increase in immune cell abundance and activation persists throughout the neurodegeneration period, indicating a robust and prolonged immune responses occurs in the cochlea after kanamycin-induced hair cell loss. I hypothesize that this immune response is causal to SGN death after hair cell loss. In this proposal, I will elucidate the function of macrophages and lymphocytes, independently, to determine their involvement in SGN degeneration after hair cell loss. First, I will test the hypothesis that macrophages are required for SGN after hair cell loss by evaluating three primary criteria. Using immunohistochemistry, I will assess 1) SGN survival and 2) recruitment of lymphocytes (CD45+/IBA1- cells) in deafened PLX5622-fed macrophage depleted rats. These experiments will indicate the extent to which macrophages are required for SGN death and lymphocyte recruitment, respectively. In addition, I will use 3) qPCR to measure gene expression of key chemokines/cytokines in the spiral ganglion to determine the extent to which macrophages may be creating an environment neurotoxic to SGNs. Secondly, I will test the hypothesis that lymphocytes are causal to SGN death after hair cell loss using the T, B, and NK cell deficient SRG rat strain. Together, the results of these experiments will determine the extent to which macrophages and/or lymphocytes contribute to SGN death after hair cell loss. These results will help guide the development of future therapeutics aimed at preventing SGN degeneration after hair cell loss, ultimately improving the efficacy of cochlear ...