The elementary events of Ca2+ release elicited by membrane depolarization in mammalian muscle.
Cytosolic [Ca(2+)] transients elicited by voltage clamp depolarization were examined by confocal line scanning of rat skeletal muscle fibres. Ca(2+) sparks were observed in the fibres' membrane-permeabilized ends, but not in responses to voltage in the membrane-intact area. Elementary events of the depolarization-evoked response could be separated either at low voltages (near -50 mV) or at -20 mV in partially inactivated cells. These were of lower amplitude, narrower and of much longer duration than sparks, similar to 'lone embers' observed in the permeabilized segments. Their average amplitude was 0.19 and spatial half-width 1.3 microm. Other parameters depended on voltage. At -50 mV average duration was 111 ms and latency 185 ms. At -20 mV duration was 203 ms and latency 24 ms. Ca(2+) release current, calculated on an average of events, was nearly steady at 0.5-0.6 pA. Accordingly, simulations of the fluorescence event elicited by a subresolution source of 0.5 pA open for 100 ms had morphology similar to the experimental average. Because 0.5 pA is approximately the current measured for single RyR channels in physiological conditions, the elementary fluorescence events in rat muscle probably reflect opening of a single RyR channel. A reconstruction of cell-averaged release flux at -20 mV based on the observed distribution of latencies and calculated elementary release had qualitatively correct but slower kinetics than the release flux in prior whole-cell measurements. The qualitative agreement indicates that global Ca(2+) release flux results from summation of these discrete events. The quantitative discrepancies suggest that the partial inactivation strategy may lead to events of greater duration than those occurring physiologically in fully polarized cells.
Department of Physiology, University of Debrecen, Hungary.
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Csernoch
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Zhou
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Stern
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Brum
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Ríos
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000502:physiology; D002462:Cell Membrane / Q000378:metabolism / Q000502:physiology; D003198:Computer Simulation; D004594:Electrophysiology; D005456:Fluorescent Dyes; D007091:Image Processing, Computer-Assisted; D066298:In Vitro Techniques; D008297:Male; D008564:Membrane Potentials / Q000502:physiology; D018613:Microscopy, Confocal; D018485:Muscle Fibers, Skeletal / Q000502:physiology; D018482:Muscle, Skeletal / Q000378:metabolism* / Q000502:physiology; D018408:Patch-Clamp Techniques; D051381:Rats; D017207:Rats, Sprague-Dawley
D005456:Fluorescent Dyes; D002118:Calcium