title: Direct measurement of single-channel Ca(2+) currents in bullfrog hair cells reveals two distinct channel subtypes.
abstract: 1. To confer their acute sensitivity to mechanical stimuli, hair cells employ Ca(2+) ions to mediate sharp electrical tuning and neurotransmitter release. We examined the diversity and properties of voltage-gated Ca(2+) channels in bullfrog saccular hair cells by means of perforated and cell-attached patch-clamp techniques. Whole-cell Ca(2+) current records provided hints that hair cells express L-type as well as dihydropyridine-insensitive Ca(2+) currents. 2. Single Ca(2+) channel records confirmed the presence of L-type channels, and a distinct Ca(2+) channel, which has sensitivity towards omega-conotoxin GVIA. Despite its sensitivity towards omega-conotoxin GVIA, the non-L-type channel cannot necessarily be considered as an N-type channel because of its distinct voltage-dependent gating properties. 3. Using 65 mM Ca(2+) as the charge carrier, the L-type channels were recruited at about -40 mV and showed a single-channel conductance of 13 pS. Under similar recording conditions, the non-L-type channels were activated at approximately -60 mV and had a single-channel conductance of approximately 16 pS. 4. The non-L-type channel exhibited at least two fast open time constants (tau(o) = 0.2 and 5 ms). In contrast, the L-type channels showed long openings (tau(o) = approximately 23 ms) that were enhanced by Bay K 8644, in addition to the brief openings (tau(o) = 0.3 and 10 ms). 5. The number of functional channels observed in patches of similar sizes suggests that Ca(2+) channels are expressed singly, in low-density clusters (2-15 channels) and in high-density clusters (20-80 channels). Co-localization of the two channel subtypes was observed in patches containing low-density clusters, but was rare in patches containing high-density clusters. 6. Finally, we confirmed the existence of two distinct Ca(2+) channel subtypes by using immunoblot and immunohistochemical techniques.
authors.affiliation: University of California-Davis, Center for Neuroscience, School of Medicine, Department of Otolaryngology, 1544 Newton Court, Davis, CA 95616, USA.
authors.forename: A
authors.initials: A
authors.lastname: Rodriguez-Contreras
authors.forename: E N
authors.initials: EN
authors.lastname: Yamoah
mesh_terms: D000818:Animals; D001671:Biochemistry / Q000379:methods; D015220:Calcium Channels / Q000502:physiology*; D020746:Calcium Channels, L-Type / Q000502:physiology; D004553:Electric Conductivity; D006198:Hair Cells, Auditory / Q000378:metabolism*; D007150:Immunohistochemistry; D007700:Kinetics; D020033:Protein Isoforms; D011892:Rana catesbeiana; D014018:Tissue Distribution
chemical_list: D015220:Calcium Channels; D020746:Calcium Channels, L-Type; D020033:Protein Isoforms