Paramecium Na+ channels activated by Ca(2+)-calmodulin: calmodulin is the Ca2+ sensor in the channel gating mechanism.
Paramecium Na+ channels, which were Ca(2+)-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 Ca+ 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 microM 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.
Saimi
Y
Y
Laboratory of Molecular Biology, University of Wisconsin-Madison, 53706, USA.
Ling
K Y
KY
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism / Q000502:physiology*; D004305:Dose-Response Relationship, Drug; D015640:Ion Channel Gating; D008274:Magnesium / Q000494:pharmacology; D008564:Membrane Potentials; D008954:Models, Biological; D010247:Paramecium; D011188:Potassium / Q000494:pharmacology; D015398:Signal Transduction; D015222:Sodium Channels / Q000378:metabolism*; D013268:Stimulation, Chemical
D002147:Calmodulin; D015222:Sodium Channels; D008274:Magnesium; D011188:Potassium; D002118:Calcium