A reappraisal of the Ca2+ dependence of fast inactivation of Ca2+ release in frog skeletal muscle

Two procedures to inhibit Ca2+ release designed to differentiate between local and common pool mechanisms for the Ca2+ dependent, fast inactivation of Ca2+ release in skeletal muscle of the frog were compared. Inhibition by voltage dependent inactivation of Ca2+ release, without modification of the single channel current of the Ryanodine Receptor (RyR) and the [Ca2+] close to the open pore, produced a reduction in the rate of inactivation linearly related to the reduction in the peak of Ca2+ release flux. Linear fits in the individual fibers were performed, giving average values (+/- SEM, N = 8) of the best fit parameters of 5.75 x 10(-3) +/- A 7.35 x 10(-4) mu M-1 for the slope and 0.07 +/- A 0.015 ms(-1) for the ordinate intercept. Inhibition of Ca2+ release by reducing the Ca content of the sarcoplasmic reticulum (SR) involves reduction of the Ca2+ current through the single RyR. The reduction in rate of inactivation also followed linearly the reduction in Ca2+ peak release flux. The average values (+/- SEM) of the best fit parameters of linear fits were 14 x 10(-3) +/- A 3.76 x 10(-3) mu M-1 and 0.019 +/- A 0.006 ms(-1) (N = 7) for slope and ordinate intercept respectively. The differences between both parameters were statistically significant (by t test, at P = 0.05). The extent of inactivation, measured by the peak/final Ca2+ release flux ratio, was differentially affected by the two procedures. Inhibition by voltage dependent inactivation, despite slowing down the fast inactivation, increased the peak/final Ca2+ release flux ratio. In contrast, depletion of the SR reticulum reduced it. If the fast inactivation is driven by the high [Ca2+] attained locally, close to the open pore of the RyR, the inhibition of Ca2+ release due to voltage dependent inactivation should not modify the rate of inactivation while inhibition by SR Ca2+ depletion should reduce it. A process driven by [Ca2+] in a common pool should depend on the overall Ca2+ release independently of how it was modified. In this case both inhibitory procedures should reduce the inactivation rate similarly. Our findings are generally consistent with a common pool process. The differences between the two protocols could be understood if the organization of RyR in junctional and parajunctional release units is considered.