THE SCN5A VARIANT C683R RESULTS IN A GAIN-OF-FUNCTION OF THE CARDIAC SODIUM CURRENT (NAV1.5) WITH ADRENERGIC MODULATION

Abstract

Congenital Long QT syndrome (LQTS) is an inherited arrhythmia characterized by prolonged cardiac repolarization that may result in torsades de pointes and sudden cardiac death. Gain-of-function variants of the SCN5A gene, encoding the cardiac sodium channel Nav1.5; INa, underlie nearly 10% of LQTS and cause LQT3. In LQT3 patients, bradycardia and low sympathetic tone (associated with sleep and rest) have been considered to trigger the arrhythmias. Recently, a novel SCN5A variant (C683R) was identified in two unrelated patients with personal/family history of prolonged QT and sudden cardiac arrest. Paradoxically, C683R-associated arrhythmias in these two patients have only been observed during hyperadrenergic states. We therefore conducted a functional study to characterize the electrophysiological effects of the SCN5A C683R variant on Nav1.5 current in a cellular model. The C683R SCN5A variant was engineered into the pcDNA1 vector by using site-directed mutagenesis. Wild-Type (WT) or C683R SCN5A cDNA was transfected into tsA201 cells, along with the human Nav1.5 channel ß1-subunit. Electrophysiological characterization of the C683R variant was performed using the whole cell patch-clamp technique. The selective protein kinase A (PKA) activator 8-CPT-cAMP 50 μM was used in the pipette solution to mimic the effect of epinephrine on either WT or C683R Nav1.5 currents. The C683R variant did not generate any persistent (late) Nav1.5 current when compared to WT. It did not cause any significant change on recovery from inactivation vs WT nor on both steady-state and slow inactivation. However, the C683R variant increased maximal Nav1.5 current density from 341±64 to 766±103 pA/pF when compared to WT (p < 0.05, n≥9) and the midpoint of activation voltage (V½) was negatively shifted from -57.18±1.24 to -63.63±1.40 mV (p < 0.001 vs WT). Moreover, when exposed to the PKA activator 8-CPT-cAMP 50 μM, the C683R variant showed a greater maximal Nav1.5 current density than WT; 993±100 vs 645±137 pA/pF (p < 0.05, n≥6). Our observations suggest a gain-of-function of the Nav1.5 current in the C683R variant compared to the WT protein. This gain of function may be further enhanced upon stimulation with catecholamines. The C683R variant of SCN5A results in a significant gain of function of the Nav1.5 current with increased responsiveness to adrenergic stimulation. Our observations provide a likely explanation for the unusual phenotype in carriers of the C683R variant that is characterized by epinephrine-triggered ventricular arrhythmias. The electrophysiological properties are different from conventional LQT3 and may represent a distinct inherited arrhythmia syndrome.