Cold-induced defects of sodium channel gating in atypical periodic paralysis plus myotonia

Abstract

Background: Missense mutations of the skeletal muscle voltage-gated sodium channel (NaV1.4) are an established cause of several clinically distinct forms of periodic paralysis and myotonia. The mechanistic basis for the phenotypic variability of these allelic disorders of muscle excitability remains unknown. An atypical phenotype with cold-induced hypokalemic paralysis and myotonia at warm temperatures was reported to segregate with the P1158S mutation.

Objective: This study extends the functional characterization of the P1158S mutation and tests the specific hypothesis that impairment of Na channel slow inactivation is a common feature of periodic paralysis.

Methods: Mutant NaV1.4 channels (P1158S) were transiently expressed in human embryonic kidney cells and characterized by voltage-clamp studies of Na currents.

Results: Wild-type and P1158S channels displayed comparable behavior at 37 degrees C, but upon cooling to 25 degrees C, mutant channels activated at more negative potentials and slow inactivation was destabilized.

Conclusions: Consistent with other NaV1.4 mutations associated with a paralytic phenotype, the P1158S mutation disrupts slow inactivation. The unique temperature sensitivity of the channel defect may contribute to the unusual clinical phenotype.

Figure 1

Kinetics of sodium channel slow inactivation. (A) Entry kinetics of slow inactivation were measured by applying a conditioning pulse to 0 mV of varying duration, followed by a brief hyperpolarization to −120 mV for 20 msec to allow recovery from fast inactivation, and then application of a test depolarization to measure the fractional recovery (pulse protocol, top). Sodium currents (traces at top) show the maximum current elicited in the absence of a conditioning depolarization (left), and a superposition of traces recorded after progressively longer conditioning pulses, which results in smaller peak currents. Plots show the decrement in relative current as slow inactivation ensues with longer duration conditioning pulses (logarithmic scale). Entry to slow inactivation is comparable for P1158S and WT channels at 37 °C, but is less complete for P1158S mutants at 25 °C. (B) Recovery from slow inactivation was monitored by brief test depolarizations after a prolonged conditioning pulse (30 s at 25 °C or 2 sec at 37 °C) to maximally slow inactivate channels (top). At 25 °C, recovery of P1158S channels precedes that of WT, primarily because the extent of slow inactivation was less complete for P1158S (larger fractional recovery at the shortest recovery times).

Figure 2

Voltage dependence of steady-state slow inactivation. Relative available current decreased with progressively more positive conditioning pulses (inset at top) that caused a larger fraction of channels to become slow inactivated. At 25 °C, the voltage dependence of slow inactivation is shifted toward more positive potentials and the extent of slow inactivation at strongly depolarized potentials is reduced (larger available relative current).

Figure 3

Membrane-spanning model for NaV1.4 topology and location of missense mutations associated with hyperkalemic periodic paralysis (HyperPP), paramyotonia congenita (PMC), potassium aggravated myotonia (PAM), hypokalemic periodic paralysis type 2 (HypoPP-2), or congenital myasthenic syndrome (CMS). Mutations previously known to disrupt slow inactivation are highlighted and are all associated with HyperPP.