There are two targets for efferent fibers in the adult cochlea: The outer hair cells (OHCs) and the dendrites of the type I spiral ganglion cells. The medial olivocochlear axons also send collaterals to the ventral cochlear nucleus (VCN). Both the medial and the lateral systems are largely cholinergic, although the receptors in neither system are well-characterized. In this laboratory, we have studied the mRNA expression of all of the nAChR subunits. We have proposed that the NACHR on OHCs may consist of alpha7 in addition to alpha9 subunit, and that the lateral efferent axodendritic receptor may consist of the alpha6alpha7beta2 subunits. In developing rat, the axodendritic receptor may also possess the alpha3 and alpha4 subunits, suggesting a developmental switch in receptor subunits, and a receptor with unique physiological or pharmacological properties. In this application we will test the hypothesis that the axodendritic NACHR consists of alpha6alpha7beta2 by using immunoprecipitation methods and by characterizing the physiology and pharmacology of the receptor using the Xenopus oocyte expression system. We will also extend our previous observations by performing experiments to quantify, with quantitative RT-PCR, and to localize, with in situ-PCR, the alpha7 subunit in the organ of Corti. In addition, we propose to study, using immunoprecipitation methods and the oocyte expression system, the alpha9 homomer and the putative alpha7alpha9 heteromeric receptor. Lastly, we propose to identify the nAChRs in the VCN, the target of the olivocochlear collaterals. Specifically, we will determine if the alpha4 and beta2 subunits are expressed in the same cells using in situ hybridization on adjacent sections from adult rat brain. Recent data from this laboratory indicates that alpha3 decreases in expression and alpha7 increases in expression in the VCN during the early postnatal period, at about the time of synaptogenesis. We hypothesize that alpha3 subunit is replaced by alpha7 and propose determine this with in situ hybridization on adjacent sections from postnatal VCN at ages PN3-21.
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