Enhanced closed-state inactivation of mutant cardiac sodium channels (SCN5A N1541D and R1632C) through different mechanisms.

BACKGROUND

SCN5A variants can be associated with overlapping phenotypes such as Brugada syndrome (BrS), sinus node dysfunction and supraventricular tachyarrhythmias. Our genetic screening of SCN5A in 65 consecutive BrS probands revealed two patients with overlapping phenotypes: one carried an SCN5A R1632C (in domain IV-segment 4), which we have previously reported, the other carried a novel SCN5A N1541D (in domain IV-segment 1).

OBJECTIVE

We sought to reveal whether or not these variants are associated with the same biophysical defects.

METHODS

Wild-type (WT) or mutant SCN5A was expressed in tsA201-cells, and whole-cell sodium currents (hNa1.5/I) were recorded using patch-clamp techniques.

RESULTS

The N1541D-I density, when assessed from a holding potential of -150 mV, was not different from WT-I as with R1632C-I, indicating that SCN5A N1541D did not cause trafficking defects. The steady-state inactivation curve of N1541D-I was markedly shifted to hyperpolarizing potentials in comparison to WT-I (V-WT: -82.3 ± 0.9 mV, n = 15; N1541D: -108.8 ± 1.6 mV, n = 26, P < .01) as with R1632C-I. Closed-state inactivation (CSI) was evaluated using prepulses of -90 mV for 1460 ms. Residual N1541D-I and R1632C-I were markedly reduced in comparison to WT-I (WT: 63.8 ± 4.6%, n = 18; N1541D: 15.1 ± 2.3%, n = 19, P < .01 vs WT; R1632C: 5.3 ± 0.5%, n = 15, P < .01 vs WT). Entry into CSI of N1541D-I was markedly accelerated, and that of R1632C-I was weakly accelerated in comparison to WT-I (tau-WT: 65.8 ± 7.4 ms, n = 18; N1541D: 13.7 ± 1.1 ms, n = 19, P < .01 vs WT; R1632C: 39.5 ± 2.9 ms, n = 15, P < .01 vs WT and N1541D). Although N1541D-I recovered from closed-state fast inactivation at the same rate as WT-I, R1632C-I recovered very slowly (tau-WT: 1.90 ± 0.16 ms, n = 10; N1541D: 1.72 ± 0.12 ms, n = 10, P = .41 vs WT; R1632C: 53.0 ± 2.5 ms, n = 14, P < .01 vs WT and N1541D).

CONCLUSIONS

Both N1541D-I and R1632C-I exhibited marked enhancement of CSI, but through different mechanisms. The data provided a novel understanding of the mechanisms of CSI of I. Clinically, the enhanced CSI of N1541D-I leads to a severe loss-of-function of I at voltages near the physiological resting membrane potential (~-90 mV) of cardiac myocytes; this can be attributable to the patient's phenotypic manifestations.