The goal of this application is to develop Dr. Marlan Hansen into an independent physician/scientist. Drs. Steve Green and Jack Lilien will serve as mentors to guide his development. Drs. Hanna Sobkowicz and Allen Ryan will serve as independent outside reviewers to ensure the progress of Dr. Hansen's development. Formal coursework in confocal microscopy techniques and application, developmental neurobiology, and research career survival skills will increase Dr. Hansen's knowledge and skills to develop into an independent investigator. The principal focus of this proposal is an intensive laboratory experience aimed at determining the effects of electrical activity on spiral ganglion neuron (SGN) neurite growth and on the innervation of the cochlea. This includes determining the necessity of activity dependent signals to mediate the effects of electrical activity. Most types of sensorineural hearing loss result from damage to the hair cells accompanied by primary or secondary damage to the SGNs. Cochlear implants rehabilitate the most severe forms of sensorineural hearing loss by providing direct electrical stimulation to the SGNs. Electrical activity regulates neurite growth and innervation patterns in many neurons. Thus, the consequences of electrical stimulation on SGN neurite growth carry important implications for cochlear development, regeneration, and implantation. We find that membrane depolarization, Ca2+/calmodulin dependent kinase II (CaMKII) activity, and protein kinase A (PKA) activity inhibit SGN neurite growth. We hypothesize that electrical stimulation inhibits SGN neurite growth by activating CaMKII and/or PKA.
The first aim of this proposal is to define the effects of physiological patterns of electrical activity on SGN neurite growth by providing different rates of electrical stimulation to SGNs in dissociated cultures and in cochlear slices. Using transgenes that encode highly specific CaMKII and PKA peptide inhibitors, we will then determine the requirement of CaMKII and PKA activity for the inhibition of neurite growth by electrical activity. Constitutively active CaMKII and PKA mutants will be introduced into SGNs in cochlear slices to test the role of these molecules in the maintenance of afferent cochlear innervation. The results of these studies will provide critical data on the consequences of electrical activity on SGN neurite growth and cochlear innervation.
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