Ion permeation and block of the gating pore in the voltage sensor of Na(V)1.4 channels with hypokalemic periodic paralysis mutations

Hypokalemic periodic paralysis and normokalemic periodic paralysis are caused by mutations of the gating charge–carrying arginine residues in skeletal muscle Na(V)1.4 channels, which induce gating pore current through the mutant voltage sensor domains. Inward sodium currents through the gating pore of mutant R666G are only ∼1% of central pore current, but substitution of guanidine for sodium in the extracellular solution increases their size by 13- ± 2-fold. Ethylguanidine is permeant through the R666G gating pore at physiological membrane potentials but blocks the gating pore at hyperpolarized potentials. Guanidine is also highly permeant through the proton-selective gating pore formed by the mutant R666H. Gating pore current conducted by the R666G mutant is blocked by divalent cations such as Ba(2+) and Zn(2+) in a voltage-dependent manner. The affinity for voltage-dependent block of gating pore current by Ba(2+) and Zn(2+) is increased at more negative holding potentials. The apparent dissociation constant (K(d)) values for Zn(2+) block for test pulses to −160 mV are 650 ± 150 µM, 360 ± 70 µM, and 95.6 ± 11 µM at holding potentials of 0 mV, −80 mV, and −120 mV, respectively. Gating pore current is blocked by trivalent cations, but in a nearly voltage-independent manner, with an apparent K(d) for Gd(3+) of 238 ± 14 µM at −80 mV. To test whether these periodic paralyses might be treated by blocking gating pore current, we screened several aromatic and aliphatic guanidine derivatives and found that 1-(2,4-xylyl)guanidinium can block gating pore current in the millimolar concentration range without affecting normal Na(V)1.4 channel function. Together, our results demonstrate unique permeability of guanidine through Na(V)1.4 gating pores, define voltage-dependent and voltage-independent block by divalent and trivalent cations, respectively, and provide initial support for the concept that guanidine-based gating pore blockers could be therapeutically useful.