Single Ca2+-activated Cl− channel currents recorded from toad olfactory cilia

Ricardo Delgado, Casilda V. Mura, Juan Bacigalupo

Odor transduction, occurring in the chemosensory cilia of vertebrate olfactory sensory neurons, is triggered by guanosine triphosphate-coupled odor receptors and mediated by a cyclic adenosine monophosphate (cAMP) signaling cascade, where cAMP opens cationic non-selective cyclic nucleotide-gated (CNG) channels. Calcium enters through CNG gates Ca(2+)-activated Cl(-) channels, allowing a Cl(-) inward current that enhances the depolarization initiated by the CNG-dependent inward current. The anoctamin channel 2, ANO2, is considered the main Ca(2+)-activated Cl(-) channel of olfactory transduction. Although Ca(2+)-activated Cl(-) channel-dependent currents in olfactory sensory neurons were reported to be suppressed in ANO2-knockout mice, field potentials from their olfactory epithelium were only modestly diminished and their smell-dependent behavior was unaffected, suggesting the participation of additional Ca(2+)-activated Cl(-) channel types. The Bestrophin channel 2, Best2, was also detected in mouse olfactory cilia and ClCa4l, belonging to the ClCa family of Ca(2+)-activated Cl(-) channels, were found in rat cilia. Best2 knock-out mice present no electrophysiological or behavioral impairment, while the ClCa channels have not been functionally studied; therefore, the overall participation of all these channels in olfactory transduction remains unresolved. We explored the presence of detectable Ca(2+)-activated Cl(-) channels in toad olfactory cilia by recording from inside-out membrane patches excised from individual cilia and detected unitary Cl(-) current events with a pronounced Ca(2+) dependence, corresponding to 12 and 24 pS conductances, over tenfold higher than the aforementioned channels, and a approx. fivefold higher Ca(2+) affinity (K0.5 = 0.38 µM). Remarkably, we observed immunoreactivity to anti-ClCa and anti-ANO2 antibodies in the olfactory cilia, suggesting a possible cooperative function of both channel type in chemotransduction. These results are consistent with a novel olfactory cilia channel, which might play a role in odor transduction.