Ca2(+)-activated K+ current involvement in neuronal function revealed by in situ single-channel analysis in Helix neurones.
1. The properties of single calcium-activated potassium channels (or C-channels) were studied in cell-attached patches using the patch-clamp technique. Experiments were performed on identified Ca2(+)-dependent U cells in juvenile specimens (1-2 months old) of Helix aspersa. 2. The criteria used to identify C-channels were based on comparison between macroscopic C-currents and currents reconstructed from unitary recordings. Both currents had a slow activation rate at large positive potentials which turned into fast activation after large Ca2+ entries. Both currents were blocked by intracellularly injected EGTA. 3. The unitary conductance in normal (5 mM) or reduced (0.5 mM) [K+]o ranged from 24 to 65 pS (mean +/- S.D., 48 +/- 13; n = 64). With 85-110 mM [K+]o, which is approximately equal to the internal [K+], the conductance was 64 pS and the reversal potential was approximately 0 mV. 4. C-channels in U cells were distributed in clusters of three to ten channels (mean 5.05 channels in seventy-five patches). Calcium channels were present in patches containing clustered C-channels. C-channels within clusters behaved independently. 5. With patch electrode containing 8 mM-calcium, C-channels opened transiently upon patch depolarization. Reopenings in quiescent depolarized patches were induced by whole-cell spikes triggered by current pulses applied to an intracellular electrode. Apparent inactivation of C-channels in depolarized patches was in fact due to a decrease in [Ca2+]i resulting from inactivation of Ca2+ channels. 6. Calcium-free saline solutions in the patch electrodes prevented C-channels from opening upon patch depolarization. Entry of calcium through the surrounding membrane induced delayed openings in the patch. Peak opening probability Po occurred 330 +/- 30 ms after a brief Ca2+ entry with a lag period of 50-80 ms. With patch electrodes filled with Ca2(+)-containing saline solutions and under conditions which maximized C-channel opening, peak Po was reached in 20-50 ms. The same value was observed for the whole-cell C-current. 7. The peak Po at a given patch potential and in response to a whole-cell spike was not altered by a previous long-lasting patch depolarization, or by producing several successive Ca2+ entries. Thus, C-channels did not appear to be inactivated by depolarization or increase in [Ca2+]i. 8. C-channels were found to be relatively highly voltage dependent, with an e-fold increase in Po per 14.9 mV increase in potential.(ABSTRACT TRUNCATED AT 400 WORDS)
Gola
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M
Laboratoire de Neurobiologie, CNRS, Marseille, France.
Ducreux
C
C
Chagneux
H
H
D000200:Action Potentials / Q000187:drug effects; D000818:Animals; D002118:Calcium / Q000502:physiology*; D004533:Egtazic Acid / Q000494:pharmacology; D006372:Helix, Snails / Q000502:physiology*; D066298:In Vitro Techniques; D008564:Membrane Potentials / Q000187:drug effects; D008954:Models, Biological; D009474:Neurons / Q000502:physiology; D015221:Potassium Channels / Q000502:physiology*; D013997:Time Factors
D015221:Potassium Channels; D004533:Egtazic Acid; D002118:Calcium