A novel mechanism for LQT3 with 2:1 block: a pore-lining mutation in Nav1.5 significantly affects voltage-dependence of activation.

BACKGROUND: SCN5A mutations that cause a gain of function in the cardiac voltage-gated sodium channel (Nav1.5) lead to long QT syndrome and a higher risk for sudden cardiac death. OBJECTIVE: Here we functionally characterize the biophysical properties of the LQT3 variant, V411M, found in a newborn with a QT interval of 640 ms and 2:1 atrioventricular block. METHODS: Whole cell patch clamp was performed on wild-type and V411M Nav1.5 channels stably expressed in human embryonic kidney cells. RESULTS: V411M channels showed hyperpolarizing shifts in both the conductance-voltage (V(1/2) = -48.5 ± 2.2 mV vs. -40.4 ± 1.6 mV for wild-type) and inactivation-voltage (-95.6 ± 1.9 mV vs. -87.7 ± 1.7 mV) relationships, and a two-fold increase in late (sustained) sodium current during voltage ramp repolarizations. While neither mexiletine nor lidocaine exhibited potency differences between WT and V411M, or shortened the QTc in vivo, increased mutant block was observed with 10 µM flecainide (71.4 ± 3.0% vs. 60.3 ± 2.8%), in a voltage-dependent manner. Incorporation of V411M kinetics into atrial and ventricular action potential models reproduced prolonged action potential repolarization. CONCLUSIONS: Our data suggest a novel mechanism for LQT3, a result of a hyperpolarizing shift in the steady state activation relationship and re-activation of Nav1.5 towards a higher open probability during repolarization of the cardiac action potential. This results in an increased number of open-activated sodium channels, and so drugs that bind this state preferentially are expected to shorten the QTc more than those that favour the inactivated state.

The molecular basis for the actions of KVbeta1.2 on the opening and closing of the KV1.2 delayed rectifier channel.

Cytosolic K(V)beta1 subunits co-assemble with transmembrane K(V)1 channel alpha-subunits and have complex effects on channel function. Fast inactivation, the most obvious effect conferred, is due to fast open channel block resulting from the binding of the N-terminus within the inner mouth of the pore. K(V)beta1 subunits also slow current deactivation, enhance slow inactivation and shift channel activation to more negative voltages, but the mechanisms underlying these actions are not known. Here we use voltage clamp fluorimetry at sites near the extracellular end of the S4 helix, the channel's primary voltage sensor, in combination with voltage clamp electrophysiology, to independently track the movement of the S4 helix along with ionic current, and thus identify the structural and mechanistic means by which the K(V)beta1.2 subunit confers its actions on the K(V)1.2 channel. We show that the negative shift in current activation is not due to direct actions of K(V)beta1.2 on the S4 segment. Instead, this shift results from an apparent saturation of channel activation at depolarized potentials as the extent of open channel block by the K(V)beta1.2 N-terminus progressively increases. The return of fluorescence to baseline is slowed along with current deactivation. According to our data, this is due to an inability of the activation gate to close while the K(V)beta1.2 N-terminus occupies the pore and strong coupling of the gate with the S4 segment. Together with data from previous studies, our findings provide a complete and coherent picture of the functional and structural interactions between K(V)beta1.2 and K(V)1.2.