Project Summary A goal of the cochlear physiology laboratory is to understand how the components of the organ of Corti tune the sound induced vibration of the organ of Corti. A process known as cochlear amplification (CA), now the subject of intense work around the world, has critical components not yet studied. Two questions of broad interest that this proposal address are; 1) how does the fundamental hydrodynamic viscosity contribute to the unique frequency analysis capacity of the cochlear apex where speech frequencies are processed and 2) does the tectorial membrane in have a central and biochemical role in regulating the calcium ion concentration that is so critical to hair cell function. There are three Aims. New and innovative experimental approaches are needed to address these questions. For the measurement of output variables, we continue to use the optical coherence tomography (OCT) method, that we pioneered, to record inner ear tissue vibration. We use state of the art confocal imaging methods applied to whole organ explant systems and measure calcium ion concentrations in quiescent and stimulated inner ears. In Aim 1, about question 1, we also propose to determine if perilymph macroscopic viscosity is a crucial parameter of apical frequency tuning. As well as whether the tuning is dependent upon the process of cochlear amplification within the traveling wave as it propagates to the apex. To manipulate viscosity, normal perilymph is replaced with altered viscosity perilymph via a real time perfusion system. Aims 2 and 3 are about question 2 where we seek to understand how and with what consequence is calcium stored by the tectorial membrane. Involved is the use of mutant mural models of defective tectorial membrane structural proteins and quantitative fluorescent determination of calcium concentrations in endolymph and tectorial membrane. Additionally, in Aim 3, we explore how age might factor into the tectorial membrane calcium sequestration via two models that manipulate the physiology of the stria vascularis a known target of age degeneration. Model 1 is the chronic application of furosemide, an agent to suppress endocochlear potential. Model 2 is the genetically targeted chemical alteration of stria vascularis blood flow). Taken together the work will significantly advance not only fundamental knowledge of organ of Corti function but open a path to pharmacological interventions to treat tectorial membrane calcium pathology.