Major contribution of sarcoplasmic reticulum Ca2+ depletion during long-lasting activation of skeletal muscle

Depolarization of skeletal muscle fibers induces sarcoplasmic reticulum (SR) Ca(2+) release and contraction that progressively decline while depolarization is maintained. Voltage-dependent inactivation of SR Ca(2+) release channels and SR Ca(2+) depletion are the two processes proposed to explain the decline of SR Ca(2+) release during long-lasting depolarizations. However, the relative contribution of these processes, especially under physiological conditions of activation, is not clearly established. Using Fura-2 and Fluo-5N to monitor cytosolic and SR Ca(2+) changes, respectively, in voltage-controlled mouse muscle fibers, we show that 2-min conditioning depolarizations reduce voltage-activated cytosolic Ca(2+) signals with a V1/2 of -53 mV but also induce SR Ca(2+) depletion that decreased the releasable pool of Ca(2+) with the same voltage sensitivity. In contrast, measurement of SR Ca(2+) changes indicated that SR Ca(2+) release channels were inactivated after SR had been depleted and in response to much higher depolarizations with a V1/2 of -13 mV. In response to trains of action potentials, cytosolic Ca(2+) signals decayed with time, whereas SR Ca(2+) changes remained stable over 1-min stimulation, demonstrating that SR Ca(2+) depletion is exclusively responsible for the decline of SR Ca(2+) release under physiological conditions of excitation. These results suggest that previous studies using steady-state inactivation protocols to investigate the voltage dependence of Ca(2+) release inactivation in fact probed the voltage dependence of SR Ca(2+) depletion, and that SR Ca(2+) depletion is the only process that leads to Ca(2+) release decline during continuous stimulation of skeletal muscle.