The regulation by calcium of the ryanodine receptor/SR calcium release channel (RyR) from rat skeletal muscle was studied under isolated conditions and in situ. RyRs were either solubilized and incorporated into lipid bilayers or single fibres were mounted into a Vaseline gap voltage clamp. Single channel data were compared to parameters determined from the calculated calcium release flux. With K+ (250 mM) being the charge carrier the single channel conductance was 529 pS at 50 μM Ca2+ cis and trans, and decreased with increasing cis [Ca2+]. Open probability showed a bell shaped calcium dependence revealing an activatory and an inhibitory Ca2+ binding site (Hill coefficients of 1.18 and 1.28, respectively) with half activatory and inhibitory concentrations of 9.4 and 298 μM. The parameters of the inhibitory site agreed with the calcium dependence of channel inactivation deduced from the decline in SR calcium release in isolated fibres. Mean open time showed slight [Ca2+] dependence following a single exponential at every Ca2+ concentration tested. Closed time histograms, at high [Ca2+], were fitted with three exponentials, from which the longest was calcium independent, and resembled the recovery time constant of SR inactivation (115±15 ms) obtained in isolated fibres. The data are in agreement with a model where calcium binding to the inhibitory site on RyR would be responsible for the calcium dependent inactivation in situ.
ASJC Scopus subject areas
- Cell Biology