Abstract: The peripheral auditory system is buried deep within the temporal bone with a cochlea surrounded by bone, pressurized and exquisitely sensitive to mechanical stimulation. These properties have made in vivo imaging of multicellular function difficult. We have developed a surgical approach that allows access to the cochlea and we have developed a chemo-mechanical approach to create imaging windows with the cochlea without loss of hearing function. We have progressed to the point where we need to survey existing and novel genetically encoded sensors to monitor cell function. We have partnered with an expert in genetically encoded sensors, Dr. Michael Lin to develop strategies to target hair cells and spiral ganglia neurons with ASAP4 voltage sensors. The goal of this proposal is to characterize sensor properties in spiral ganglia neurons in vitro and in vivo to identify the best sensor and the best means to target the sensor to soma and synapses. We further are characterizing calcium sensors like GCaMP 6s and f in both inner and outer hair cells as these exist in cre activated forms that can be selectively activated in hair cells for in vivo imaging. We will also characterize voltage sensors in hair cells in vitro and move to in vivo if these sensors prove better than the GCaMPs. This preparation along with the sensor characterization will open a new era in multicellular functional imaging within the cochlea.