Termination of cardiac Ca2+ sparks: Role of intra-SR [Ca2+], release flux, and intra-SR Ca2+ diffusion

Ca release from cardiac sarcoplasmic reticulum (SR) via ryanodine receptors (RyRs) is regulated by dyadic cleft [Ca] and intra-SR free [Ca] ([Ca]SR). Robust SR Ca release termination is important for stable excitation-contraction coupling, and partial [Ca]SR depletion may contribute to release termination. Here, we investigated the regulation of SR Ca release termination of spontaneous local SR Ca release events (Ca sparks) by [Ca]SR, release flux, and intra-SR Ca diffusion. We simultaneously measured Ca sparks and Ca blinks (localized elementary [Ca]SR depletions) in permeabilized ventricular cardiomyocytes over a wide range of SR Ca loads and release fluxes. Sparks terminated via a [Ca]SR-dependent mechanism at a fixed [Ca]SR depletion threshold independent of the initial [Ca]SR and release flux. Ca blink recovery depended mainly on intra-SR Ca diffusion rather than SR Ca uptake. Therefore, the large variation in Ca blink recovery rates at different release sites occurred because of differences in the degree of release site interconnection within the SR network. When SR release flux was greatly reduced, long-lasting release events occurred from well-connected junctions. These junctions could sustain release because local SR Ca release and [Ca]SR refilling reached a balance, preventing [Ca]SR from depleting to the termination threshold. Prolonged release events eventually terminated at a steady [Ca]SR, indicative of a slower, [Ca]SR-independent termination mechanism. These results demonstrate that there is high variability in local SR connectivity but that SR Ca release terminates at a fixed [Ca]SR termination threshold. Thus, reliable SR Ca release termination depends on tight RyR regulation by [Ca]SR.