Movement of voltage sensor S4 in domain 4 is tightly coupled to sodium channel fast inactivation and gating charge immobilization

The highly charged transmembrane segments in each of the four homologous do mains (S4D1-S4D4) represent the principal voltage sensors for sodium channe l gating. Hitherto, the existence of a functional specialization of the fou r voltage sensors with regard to the control of the different gating modes, i.e., activation, deactivation, and inactivation, is problematic, most lik ely due to a functional coupling between the different domains. However, re cent experimental data indicate that the voltage sensor in domain 4 (S4D4) plays a unique role in sodium channel fast inactivation. The correlation of fast inactivation and the movement of the S4D4 voltage sensor in rat brain IIA sodium channels was examined by site-directed mutagenesis of the centr al arginine residues to histidine and by analysis of both ionic and gating currents using a high expression system in Xenopus oocytes and an optimized two-electrode voltage clamp. Mutation R1635H shifts the steady state inact ivation to more hyperpolarizing potentials and drastically increases the re covery time constant, thereby indicating a stabilized inactivated state. In contrast, R1638H shifts the steady state inactivation to more depolarizing potentials and strongly increases the inactivation time constant, thereby suggesting a preferred open state occupancy. The double mutant R1635/1638H shows intermediate effects on inactivation. Tn contrast, the activation kin etics are not significantly influenced by any of the mutations. Gating curr ent immobilization is markedly decreased in R1635H and R1635/1638H but only moderately in R1638H. The time courses of recovery from inactivation and i mmobilization correlate well in wild-type and mutant channels, suggesting a n intimate coupling of these two processes that is maintained in the mutati ons. These results demonstrate that S4D4 is one of the immobilized voltage sensors during the manifestation of the inactivated state. Moreover, the pr esented data strongly suggest that S4D4 is involved in the control of fast inactivation.