Intra-sarcoplasmic reticulum Ca2+ oscillations are driven by dynamic regulation of ryanodine receptor function by luminal Ca2+ in cardiomyocytes

During the cardiac cycle, the release of Ca2+ from the sarcoplasmic reticulum (SR) through the ryanodine receptor (RyR2) channel complex is controlled by the levels of cytosolic and luminal Ca2+ and alterations in these regulatory processes have been implicated in cardiac disease including arrhythmia. To better understand the mechanisms of regulation of SR Ca2+ release by Ca2+ on both sides of the SR membrane, we investigated SR Ca2+ release in a wide range of cytosolic Ca2+ concentrations ([Ca2+](cyt); 1-100 mu M) in permeabilized canine ventricular myocytes by monitoring [Ca2+] inside the SR ([Ca2+](SR)). Exposing myocytes to activating [Ca2+](cyt) resulted in spontaneous oscillations of [Ca2+](SR) due to periodic opening and closing of the RyR2s. Elevating [Ca2+](cyt) (up to 10 mu M) increased the frequency of [Ca2+](SR) oscillations; however at higher [Ca2+](cyt) (> 50 mu M) the oscillations diminished due to RyR2s staying perpetually open, resulting in depleted SR. Ablation of cardiac calsequestrin (CASQ2) altered the [Ca2+](cyt) dependence of Ca2+ release oscillations such that oscillations were highly frequent at low [Ca2+](cyt) (100 nM) but became diminished at moderate [Ca2+](cyt) (10 mu M), as determined in myocytes from calsequestrin-null versus wild-type mice. Our results suggest that under conditions of continuous activation by cytosolic Ca2+, RyR2s can periodically cycle between open and deactivated states due to effects of luminal Ca2+. Deactivation at reduced [Ca2+](SR) appears to involve reduction of sensitivity to cytosolic Ca2+ and might be mediated by CASQ2. Inactivation by cytosolic Ca2+ plays no detectable role in controlling SR Ca2+ release.