Enhanced Cytosolic Ca2+ Activation Underlies a Common Defect of Central Domain Cardiac Ryanodine Receptor Mutations Linked to Arrhythmias

Recent three-dimensional structural studies reveal that the central domain of ryanodine receptor (RyR) serves as a transducer that converts long-range conformational changes into the gating of the channel pore. Interestingly, the central domain encompasses one of the mutation hotspots (corresponding to amino acid residues 3778-4201) that contains a number of cardiac RyR (RyR2) mutations associated with catecholaminergic polymorphic ventricular tachycardia (CPVT) and atrial fibrillation (AF). However, the functional consequences of these central domain RyR2 mutations are not well understood. To gain insights into the impact of the mutation and the role of the central domain in channel function, we generated and characterized eight disease-associated RyR2 mutations in the central domain. We found that all eight central domain RyR2 mutations enhanced the CA²⁺-dependent activation of [³H]ryanodine binding, increased cytosolic CA²⁺-induced fractional CA²⁺ release, and reduced the activation and termination thresholds for spontaneous CA²⁺ release in HEK293 cells. We also showed that racemic carvedilol and the non-beta-blocking carvedilol enantiomer, (R)-carvedilol, suppressed spontaneous CA²⁺ oscillations in HEK293 cells expressing the central domain RyR2 mutations associated with CPVT and AF. These data indicate that the central domain is an important determinant of cytosolic CA²⁺ activation of RyR2. These results also suggest that altered cytosolic CA²⁺ activation of RyR2 represents a common defect of RyR2 mutations associated with CPVT and AF, which could potentially be suppressed by carvedilol or (R)-carvedilol.

Ca2+-dependent activation; atrial fibrillation; calcium; calcium channel; calcium imaging; calcium intracellular release; cardiac ryanodine receptor; carvedilol; carvedilol enantiomer; disease mutations; endoplasmic reticulum (ER); ryanodine receptor; sarcoplasmic reticulum (SR); spontaneous Ca2+ release; ventricular tachyarrhythmia.