Reply to Eisner et al.: CaMKII phosphorylation of RyR2 increases cardiac contractility
Proceedings of the National Academy of Sciences of the United States of America
The ryanodine receptor/calcium-release channel (RyR2) on the sarcoplasmic reticulum (SR) is the source of Ca 2+ required for myocardial excitation-contraction (EC) coupling. During stress (i.e., exercise), contractility of the cardiac muscle is increased largely because of phosphorylation and activation of key proteins that regulate SR Ca 2+ release. These include the voltage-gated calcium channel (Cav1.2) on the plasma membrane through which Ca 2+ enters the cardiomyocyte, the
... he sarco/endoplasmic reticulum calcium ATPase (SERCA2a)/phospholamban complex that pumps Ca 2+ into the SR, and the RyR2 channel that releases Ca 2+ from the SR, all of which are activated by phosphorylation. For the past 10 y, Eisner et al. (1) have advanced the idea that activation of the RyR2 channel (e.g., by phosphorylation) cannot play a role in regulating systolic Ca 2+ release and cardiac contractility. They base their position on an experiment in which they used caffeine to activate the RyR2 channel and showed that Ca 2+ release was increased but after a few beats, returned to baseline (1). However, their experiment is not a good model for the physiological response to stress in which the three key regulators of EC coupling are all activated by the same signal (i.e., phosphorylation) such that there is increased Ca 2+ influx, increased SR Ca 2+ uptake, and increased SR Ca 2+ release. In the Eisner caffeine experiment, RyR2 was activated, but the Cav1.2 and SERCA2a were not. Selective activation of RyR2 is not physiological, and the outcome of their experiment was predictable. Caffeine-induced activation of RyR2 resulted in a transient increase in SR Ca 2+ release, but because there was no concomitant increase in Ca 2+ influx or SR Ca 2+ uptake, the increase in SR Ca 2+ release could not be sustained. However, on the basis of this experiment, Eisner et al. (1) concluded that activation of RyR2 plays no role in stress-induced increased cardiac contractility. We have shown that, during stress, the increased heart rate results in a rate-dependent activation of CaMKII that phosphorylates and activates RyR2. We showed the essential role of this rate-dependent activation of RyR2 by CaMKII by showing that genetically engineered mice, lacking the CaMKII phosphorylation site on RyR2 (RyR2-S2814A), exhibit blunted increases in systolic Ca 2+ -transient amplitudes and contractile responses as heart rate increases (2). We also showed that a reduction in the amount of CaMKII in the RyR2 complex in failing hearts results in defective regulation of the channel, which could explain the loss of the rate-dependent increase in contractility in heart failure. Eisner et al. (3) challenge all of our findings based on their caffeine experiment. However, our experiments have been conducted under physiological conditions in which all three components involved in Ca 2+ signaling during muscle contraction are activated, not just one. The only perturbation that we have introduced is to ablate the CaMKII phosphorylation site on RyR2 using a single amino acid substitution. This results in a blunted contractile response, leading us to conclude that CaMKII phosphorylation of RyR2 does indeed play a key role in enhancing contractility as the heart rate increases. 1. Trafford AW, Díaz ME, Sibbring GC, Eisner DA (2000) Modulation of CICR has no maintained effect on systolic Ca2+: Simultaneous measurements of sarcoplasmic reticulum and sarcolemmal Ca2+ fluxes in rat ventricular myocytes. J Physiol 522:259-270. 2. Kushnir A, Shan J, Betzenhauser MJ, Reiken S, Marks AR (2010) Role of CaMKIIδ phosphorylation of the cardiac ryanodine receptor in the force frequency relationship and heart failure.