Abstract
Flecainide blocks ryanodine receptor type 2 (RyR2) channels in the open state, suppresses arrhythmogenic Ca2+ waves and prevents catecholaminergic polymorphic ventricular tachycardia (CPVT) in mice and humans. We hypothesized that differences in RyR2 activity induced by CPVT mutations determines the potency of open-state RyR2 blockers like flecainide (FLEC) and R-propafenone (RPROP) against Ca2+ waves in cardiomyocytes. Using confocal microscopy, we studied Ca2+ sparks and waves in isolated saponin-permeabilized ventricular myocytes from two CPVT mouse models (Casq2-/-, RyR2-R4496C+/-), wild-type (c57bl/6, WT) mice, and WT rabbits (New Zealand white rabbits). Consistent with increased RyR2 activity, Ca2+ spark and wave frequencies were significantly higher in CPVT compared to WT mouse myocytes. We next obtained concentration-response curves of Ca2+ wave inhibition for FLEC, RPROP (another open-state RyR2 blocker), and tetracaine (TET) (a state-independent RyR2 blocker). Both FLEC and RPROP inhibited Ca2+ waves with significantly higher potency (lower IC50) and efficacy in CPVT compared to WT. In contrast, TET had similar potency in all groups studied. Increasing RyR2 activity of permeabilized WT myocytes by exposure to caffeine (150 µM) increased the potency of FLEC and RPROP but not of TET. RPROP and FLEC were also significantly more potent in rabbit ventricular myocytes that intrinsically exhibit higher Ca2+ spark rates than WT mouse ventricular myocytes. In conclusion, RyR2 activity determines the potency of open-state blockers FLEC and RPROP for suppressing arrhythmogenic Ca2+ waves in cardiomyocytes, a mechanism likely relevant to antiarrhythmic drug efficacy in CPVT.
Highlights
Type 2 ryanodine receptors (RyR2) are intracellular Ca2+ release channels located on the surface of the sarcoplasmic reticulum (SR) in cardiac muscle cells [1,2,3]
We find that ryanodine receptor type 2 (RyR2) activity determines the potency of open-state blockers FLEC and R-propafenone [RPROP] for suppressing arrhythmogenic Ca2+ waves in cardiomyocytes, a mechanism likely relevant to antiarrhythmic drug efficacy in catecholaminergic polymorphic ventricular tachycardia (CPVT)
Consistent with an increased RyR2 channel activity, both R4496C+/- and Casq2-/- myocytes exhibited increased spark frequency compared to WT myocytes (Fig 1A and 1B)
Summary
Type 2 ryanodine receptors (RyR2) are intracellular Ca2+ release channels located on the surface of the sarcoplasmic reticulum (SR) in cardiac muscle cells [1,2,3]. Together with L-type Ca2+ channels located on the sarcolemma, RyR2 channels are responsible for the triggering arm of the Ca2+-induced Ca2+ release mechanism in the ventricle and excitation contraction coupling in cardiac cells [4,5,6,7]. Regardless of the specific molecular mechanism, CPVT mutations induce a common dysregulation of intracellular Ca2+ release that is characterized by an increased open probability of RyR2 Ca2+ release channels and results in the generation of arrhythmogenic Ca2+ waves [18,19,20]
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