Isolated Cardiac Ryanodine Receptor Function Varies Between Mammals.

Concerted robust opening of cardiac ryanodine receptors' (RyR2) Ca2+ release 1oplasmic reticulum (SR) is fundamental for normal systolic cardiac function. During diastole, infrequent spontaneous RyR2 openings mediate the SR Ca2+ leak that normally constrains SR Ca2+ load. Abnormal large diastolic RyR2-mediated Ca2+ leak events can cause delayed after depolarizations (DADs) and arrhythmias. The RyR2-associated mechanisms underlying these processes are being extensively studied at multiple levels utilizing various model animals. Since there are well-described species-specific differences in cardiac intracellular Ca2+ handing in situ, we tested whether or not single RyR2 function in vitro retains this species specificity. We isolated RyR2-rich heavy SR microsomes from mouse, rat, rabbit, and human ventricular muscle and quantified RyR2 function using identical solutions and methods. The single RyR2 cytosolic Ca2+ sensitivity was similar across these species. However, there were significant species differences in single RyR2 mean open times in both systole and diastole-like solutions. In diastole-like solutions, single rat/mouse RyR2 open probability and frequency of long openings (> 6 ms) were similar, but these values were significantly greater than those of either single rabbit or human RyR2s. We propose these in vitro single RyR2 functional differences across species stem from the species-specific RyR2 regulatory environment present in the source tissue. Our results show the single rabbit RyR2 functional attributes, particularly in diastole-like conditions, replicate those of single human RyR2 best among the species tested.

Diminished inhibition and facilitated activation of RyR2-mediated Ca2+ release is a common defect of arrhythmogenic calmodulin mutations.

A number of calmodulin (CaM) mutations cause severe cardiac arrhythmias, but their arrhythmogenic mechanisms are unclear. While some of the arrhythmogenic CaM mutations have been shown to impair CaM-dependent inhibition of intracellular Ca2+ release through the ryanodine receptor type 2 (RyR2), the impact of a majority of these mutations on RyR2 function is unknown. Here, we investigated the effect of 14 arrhythmogenic CaM mutations on the CaM-dependent RyR2 inhibition. We found that all the arrhythmogenic CaM mutations tested diminished CaM-dependent inhibition of RyR2-mediated Ca2+ release and increased store-overload induced Ca2+ release (SOICR) in HEK293 cells. Moreover, all the arrhythmogenic CaM mutations tested either failed to inhibit or even promoted RyR2-mediated Ca2+ release in permeabilized HEK293 cells with elevated cytosolic Ca2+ , which was markedly different from the inhibitory action of CaM wild-type. The CaM mutations also altered the Ca2+ -dependency of CaM binding to the RyR2 CaM-binding domain. These results demonstrate that diminished inhibition, and even facilitated activation, of RyR2-mediated Ca2+ release is a common defect of arrhythmogenic CaM mutations.