Flecainide induces a sustained countercurrent dependent effect on RyR2 in permeabilized WT ventricular myocytes but not in intact cells

Background and purpose: While flecainide is now an accepted treatment for arrhythmias associated with catecholaminergic polymorphic ventricular tachycardia (CPVT), its mechanism of action remains controversial. In studies on myocytes from CPVT mice, inhibition of proarrhythmic Ca 2+ waves was initially attributed to a novel action on the type-2 ryanodine receptor (RyR2). However, subsequent work on wild type (WT) myocytes questioned the conclusion that flecainide has a direct action on RyR2. In the present study, the effects of flecainide were compared in intact and permeabilized WT myocytes. Experimental approach: Intracellular Ca 2+ was measured using confocal microscopy in intact or saponin permeabilized adult rat ventricular myocytes (ARVM). In some experiments on permeabilized cells, flecainide was studied following partial inhibition of the sarcoplasmic reticulum (SR) counter-current. Key results: Flecainide induced sustained changes Ca 2+ sparks and waves in permeabilized ARVM, which were comparable to those reported in intact or permeabilized myocytes from CPVT mice. However, a relatively high level of flecainide (25 μM) was required to induce these effects. Inhibition of the SR counter-current potentiated the effects of flecainide on SR Ca 2+ waves. In intact field stimulated ARVM, prolonged exposure to 15 μM flecainide decreased wave frequency but RyR2 dependent effects on Ca 2+ sparks were absent; higher drug concentrations blocked field stimulation, consistent with inhibition of Nav1.5. Conclusions and implications: In intact ARVM, the absence of effects on Ca 2+ sparks suggests that the intracellular flecainide concentration was insufficient to influence RyR2. Wave inhibition in intact ARVM may reflect secondary effects of Nav1.5 inhibition. Potentiation of flecainide’s action by counter-current inhibition can be explained if transient polarization of the SR membrane during SR Ca 2+ release facilitates its action on RyR2.