We associate altered ventricular myocardial CV potentially resulting in arrhythmogenic substrate with arrhythmic properties associated with genetic RyR2 alterations for the first time.

Catecholaminergic polymorphic ventricular tachycardia (CPVT) predisposes to ventricular arrhythmia due to altered Ca2+ homeostasis and can arise from ryanodine receptor (RyR2) mutations including RyR2-P2328S. Previous reports established that homozygotic murine RyR2-P2328S (RyR2S/S) hearts show an atrial arrhythmic phenotype associated with reduced action potential (AP) conduction velocity and sodium channel (Nav1.5) expression. We now relate ventricular arrhythmogenicity and slowed AP conduction in RyR2S/S hearts to connexin-43 (Cx43) and Nav1.5 expression and Na+ current (INa). Stimulation protocols applying extrasystolic S2 stimulation following 8 Hz S1 pacing at progressively decremented S1S2 intervals confirmed an arrhythmic tendency despite unchanged ventricular effective refractory periods (VERPs) in Langendorff-perfused RyR2S/S hearts. Dynamic pacing imposing S1 stimuli then demonstrated that progressive reductions of basic cycle lengths (BCLs) produced greater reductions in conduction velocity at equivalent BCLs and diastolic intervals in RyR2S/S than WT, but comparable changes in AP durations (APD90) and their alternans. Western blot analyses demonstrated that Cx43 protein expression in whole ventricles was similar, but Nav1.5 expression in both whole tissue and membrane fractions were significantly reduced in RyR2S/S compared to wild-type (WT). Loose patch-clamp studies similarly demonstrated reduced INa in RyR2S/S ventricles. We thus attribute arrhythmogenesis in RyR2S/S ventricles resulting from arrhythmic substrate produced by reduced conduction velocity to downregulated Nav1.5 reducing INa, despite normal determinants of repolarization and passive conduction. The measured changes were quantitatively compatible with earlier predictions of linear relationships between conduction velocity and the peak INa of the AP but nonlinear relationships between peak INa and maximum Na+ permeability.

We explored for relationships between SCN5A haploinsufficiency, implicated in clinical arrhythmogenicity, and right ventricular (RV) conduction disorders in Langendorff-perfused, male and female, and young (3 months) and old (>12 month old) Scn5a+/− and wild type (WT) hearts. The investigated conditions of genotype, age, and sex affected latencies but not repolarization time courses of RV monophasic action potentials. This prompted examination of the patterns of RV epicardial activation, its dispersion, and their interrelationships as possible arrhythmic mechanisms using a 64-channel, multi-electrode array. Mean ventricular activation times (T*MEAN), spatial dispersions (D*S) between recording channels/cardiac cycle, and maximum activation times (T*MAX) representing the slowest possible conduction in any given heart were all higher in old male Scn5a+/− compared with young male and old female Scn5a+/− and old male WT. Temporal dispersions (D*T) of recording channels were similarly higher in old male Scn5a+/− compared with old male WT. All groupings of D*T, D*S, and T*MAX nevertheless linearly correlated with T*MEAN, with indistinguishable slopes. The variates explored thus influence D*T, D*S, and T*MAX through actions on T*MEAN. These findings in turn correlated with increased levels of fibrosis in young male, young female, and old male Scn5a+/− compared with the corresponding WTs. We thus demonstrate for the first time independent and interacting effects of genotype, age, and sex on epicardial conduction and its dispersions at least partially attributable to fibrotic change, resulting in the greatest effects in old male Scn5a+/− in an absence of alterations in repolarization time courses. This directly implicates altered depolarization in the clinical arrhythmogenicity associated with Scn5a+/−.