In collaboration with the Laboratory of Molecular Biology, BNP, DIR, NINDS we have continued a study of the molecular biology of genes involved in the regulation and synthesis of putative auditory nerve and auditory hair cell neurotransmitters, i.e. the excitatory amino acids glutamic and aspartate. We have focused on the enzymes glutamate dehydrogenase and glutaminase and have as reported previously characterized a cDNA molecule which partially encodes glutaminase. Our goal has been to identify and sequence additional cDNAs to obtain a complete sequence for glutaminase. We have initiated a study of the hereditary deafness in a strain of mouse, the Bronx Waltzer, with the mutant gene bv that is autosomal, recessive, with full penetrance. The mutant gene bv will be mapped, partly with the use of a mouse linkage testing stock, in collaboration with Dr. B. A. Taylor, The Jackson Laboratory, Bar Harbor, Maine. The deafness mouse, with the mutant gene dn, has recently been brought to the NIH, also for molecular genetic experiments in this Laboratory. As an additional step to define biochemically active molecules in the inner ear we are making monoclonal antibodies to the chick cochlea. Hybridomas were successfully generated using an in vitro immunization procedure. Supernatants were screened using a combination of ELISA assays and immunocytochemistry. Preliminary findings indicate that the one of our hybridomas secretes an antibody which labels the surface of a sub-population of cells in the sensory auditory organ in the chick. Studies of this antibody and other monoclonal antibodies are in progress. Our immunocytochemical studies of neurotransmitters, neuromodulators, receptor and other markers in the cochlea are being continued. We have examined the calcitonin-gene related peptide (CGRP)-like immunoreactivity in the guinea pig organ of Corti at both the light and electron microscope level using immunofluorescence and immunoperoxidase techniques. Our findings were that the neuropeptide, CGRP is present in both the lateral and the medial efferent system. Moreover, the immunoreactivity was localized in large dense core vesicles.
