Abstract
Signal transduction depends critically on the spatial localization of protein constituents. A key question in odor transduction is whether chemotransduction proteins organize into discrete molecular complexes throughout olfactory cilia or distribute homogeneously along the ciliary membrane. Our recordings of Ca(2+) changes in individual cilia with unprecedented spatial and temporal resolution, by the use of two-photon microscopy, provide solid evidence for Ca(2+) microdomains (transducisomes). Dissociated frog olfactory neurons were preloaded with caged-cAMP and fluo-4 acetoxymethyl ester probe Ca(2+) indicator. Ca(2+) influx through cyclic nucleotide-gated (CNG) channels was evoked by uniformly photoreleasing cAMP, while ciliary Ca(2+) was measured. Discrete fluorescence events were clearly resolved. Events were missing in the absence of external Ca(2+) , consistent with the absence of internal Ca(2+) sources. Fluorescence events at individual microdomains resembled single-CNG channel fluctuations in shape, mean duration and kinetics, indicating that transducisomes typically contain one to three CNG channels. Inhibiting the Na(+) /Ca(2+) exchanger or the Ca(2+) -ATPase prolonged the decay of evoked intraciliary Ca(2+) transients, supporting the participation of both transporters in ciliary Ca(2+) clearance, and suggesting that both molecules localize close to the CNG channel. Chemosensory transducisomes provide a physical basis for the low amplification and for the linearity of odor responses at low odor concentrations.