Paramecium Na+ channels activated by Ca2+-calmodulin: Calmodulin is the Ca2+ sensor in the channel gating mechanism |
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Authors: | Y Saimi K -Y Ling |
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Institution: | (1) Laboratory of Molecular Biology, University of Wisconsin-Madison, 53706 Madison, WI |
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Abstract: | Paramecium Na+ channels, which were Ca2+-calmodulin activated, were studied in the inside-out mode of patch clamp. After excision of the membrane patch, they were active in the presence of 10–5 to 10–3
m Ca2+ in the bath. They became much less active in the presence of 10–6
m Ca2+, and their activity subsided completely at 10–8
m Ca2+. A Hill plot showed a dissociation constant of 6 m for Ca2+ binding. This dissociation constant shifted to a submicromolar range in the presence of 1 mm Mg2+. The channels also exhibited a mild voltage dependence. When exposed to 10–8
m Ca2+ for an extended period of 2–4 min, channels were further inactivated even after bath Ca2+ was restored to 10–4
m. Whereas neither high voltage (+100 mV) nor high Ca2+ (10–3
m) was effective in reactivation of the inactive channels, addition of Paramecium wild-type calmodulin together with high Ca2+ to the bath restored channel activity without a requirement of additional Mg2+ and metabolites such as ATP. The channels reactivated by calmodulin had the same ion conductance, ion selectivity and Ca2+ sensitivity as those prior to inactivation. These inactivation and reactivation of the channels could be repeated, indicating that the direct calmodulin effect on the Na+ channel was reversible. Thus, calmodulin is a physiological factor critically required for Na+ channel activation, and is the Ca2+ sensor of the Na+-channel gating machinery.We thank C. Kung for his kind support, and A. Boileau for critical reading. Supported by grants from National Institutes of Health GM 22714-20 and 36386-09. |
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Keywords: | Paramecium Patch clamp Calmodulin Ion channels Channel regulation Ca2+ sensor |
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