Arrhythmogenic mechanisms in a mouse model of catecholaminergic polymorphic ventricular tachycardia

Abstract

Catecholaminergic polymorphic ventricular tachycardia (VT) is a lethal familial disease characterized by bidirectional VT, polymorphic VT, and ventricular fibrillation. Catecholaminergic polymorphic VT is caused by enhanced Ca2+ release through defective ryanodine receptor (RyR2) channels. We used epicardial and endocardial optical mapping, chemical subendocardial ablation with Lugol's solution, and patch clamping in a knockin (RyR2/RyR2(R4496C)) mouse model to investigate the arrhythmogenic mechanisms in catecholaminergic polymorphic VT. In isolated hearts, spontaneous ventricular arrhythmias occurred in 54% of 13 RyR2/RyR2(R4496C) and in 9% of 11 wild-type (P=0.03) littermates perfused with Ca2+and isoproterenol; 66% of 12 RyR2/RyR2(R4496C) and 20% of 10 wild-type hearts perfused with caffeine and epinephrine showed arrhythmias (P=0.04). Epicardial mapping showed that monomorphic VT, bidirectional VT, and polymorphic VT manifested as concentric epicardial breakthrough patterns, suggesting a focal origin in the His-Purkinje networks of either or both ventricles. Monomorphic VT was clearly unifocal, whereas bidirectional VT was bifocal. Polymorphic VT was initially multifocal but eventually became reentrant and degenerated into ventricular fibrillation. Endocardial mapping confirmed the Purkinje fiber origin of the focal arrhythmias. Chemical ablation of the right ventricular endocardial cavity with Lugol's solution induced complete right bundle branch block and converted the bidirectional VT into monomorphic VT in 4 anesthetized RyR2/RyR2(R4496C) mice. Under current clamp, single Purkinje cells from RyR2/RyR2(R4496C) mouse hearts generated delayed afterdepolarization-induced triggered activity at lower frequencies and level of adrenergic stimulation than wild-type. Overall, the data demonstrate that the His-Purkinje system is an important source of focal arrhythmias in catecholaminergic polymorphic VT.

Figure 1

Ventricular epicardial activation during SR. A, Activation map and ECG from a representative WT heart. B, Map and ECG from a RyR2/RyR2^R4496C heart. C, Mean activation map of 4 WT hearts. D, Mean activation map of 5 RyR2/RyR2^R4496C hearts. Maps are superimposed on the image of a mouse heart. Asterisks indicate sites of initial breakthrough; AAo, aorta; RA, right atrium; LA, left atrium; PA, pulmonary artery.

Figure 2

Epicardial (A and B) and endocardial (C and D) activation during SR and MVT in RyR2/RyR2^R4496C hearts. A, Epicardial activation map in SR, with corresponding OSPR (red) and ECG. B, Epicardial activation map during MVT, with corresponding OSPR and ECG. C, RV endocardial activation map in SR showing activation sequence in the RBB. OSPR and ECG are shown below. D, RV endocardial activation map during MVT with corresponding OSPR and ECG. White scale bars, 1 mm; black scale bars, 100 ms. Red squares indicate time of OPSR upstroke with respect to ECG; white lines on color maps, 0.5-ms isochrones; Epi, epicardium; Endo, endocardium. See Figure 1, for the remaining abbreviations.

Figure 3

BVT in a RyR2/RyR2^R4496C heart. A, Heart image. B, Epicardial activation map and ECG in SR. C, Epicardial activation maps of 2 consecutive ventricular beats with an origin that changed from RV (beat 1) to LV (beat 2). Bottom, ECG in BVT (2.7 mmol/L Ca²⁺; 100 nmol/L isoproterenol). LAD indicates left anterior descendent coronary; LA, left atrium.

Figure 4

PVT in RyR2/RyR2^R4496C heart. A, Location of each of 8 consecutive ectopic discharges during PVT superimposed on a high-resolution image of RV endocardium. Each color corresponds to a different discharge and complex number on ECG. B, Magnification of 2 boxed areas in A highlights Purkinje fibers. Each number and color indicates the origin of each discharge and complex number on ECG. Beat 5 originated outside the field of view and is not represented on the map.

Figure 5

Left, PVT-to-VF transition in RyR2/RyR2^R4496C heart (1.8 mmol/L Ca²⁺; 1 mmol/L caffeine). Left, successive epicardial phase maps (10-ms snapshots). Top, Three maps showing a focal discharge during PVT. Middle, Transition from focal discharge (leftmost) to reentry (rightmost) by formation of a wave break (center). Bottom, Drifting rotor maintained VF. Right, ECG (top) and color code for action potential phases.

Figure 6

Effects of chemical ablation of RV endocardium in anesthetized WT and RyR2/RyR2^R4496C mice. A, ECG during SR in WT mouse (top) and ECG during SR after Lugol’s solution injection (bottom). Note RBB block pattern. B, ECG during SR in RyR2/RyR2^R4496C mouse (top), BVT after administration of 120 mg/kg caffeine and 2 mg/kg epinephrine (middle), and Lugol’s solution injection in RV cavity converts BVT to MVT with RBB block (bottom). C, Morphology and width of the QRS complex in baseline (left) and after Lugol (right) in WT and RyR2/RyR2^R4496C mice.

Figure 7

Frequency dependence of DADs and TA. A, WT Purkinje cell driven by trains of 20 pulses at 1, 5, 10 Hz at baseline (A1 to A3) and during 30 nmol/L isoproterenol superfusion (A4 to A6). B, Two different RyR2/RyR2^R4496C Purkinje cells (B1 and B2; B3 and B4) subjected to the same protocol. DADs and TA were elicited in RyR2/RyR2^R4496C already at baseline (B1 to B3). In the presence of isoproterenol (30 nmol/L), trains of stimuli (5 Hz) generated long episodes of sustained TA.

Figure 8

DAD amplitude vs CI in Purkinje cells from RyR2/RyR2^R4496C (○) (5 mice) and WT (●) (5 mice) at 10 Hz. The RyR2/RyR2^R4496C data are in the absence of isoproterenol. The WT data are in the presence of 30 nmol/L isoproterenol.