Stimulus activation of olfactory neurons is initiated by reversible interactions of odorants with receptor proteins in the dendritic ciliary membrane. Stimulus binding to receptor subsequently leads to membrane depolarization, action potential generation, and synaptic transmission. However, the sequence of molecular events between receptor activation and membrane depolarization remains incompletely characterized. Recent bio-chemical and electrophysiological evidence in several vertebrate species has indicated that G-protein-linked second messenger systems may be involved in olfactory stimulus response coupling. The major goal of the proposed research is to identify stimulus regulated second messengers in the olfactory system of the channel catfish and to determine the role of second messengers in the ionic events underlying membrane depolarization. Biochemical, neurophysiological, and biophysical methods will be used to characterize the molecular and cellular mechanisms that link stimulus-receptor interaction and subsequent ionic events. Biochemical experiments will be used to determine the relationships between second messenger metabolism and receptor specificity and occupancy. The role of G-proteins in mediating second messenger metabolism will be studied biochemically in isolated cilia using established G-protein effectors. These experiments, in combination with electrophysiological and biophysical studies in reconstituted membranes and dissociated receptor cells, will be used to determine the roles of second messengers and G-protein in regulation of ion channels and intracellular ion concentrations. Application of these techniques to a model system in which stimulus receptor interaction is well characterized, will provide a comprehensive and integrated description of the molecular and cellular events underlying olfactory signal transduction.
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