Conformations of voltage-sensing domain iii differentially define na_v channel closed-and open-state inactivation

Voltage-gated Na⁺ (Na_V) channels underlie the initiation and propagation of action potentials (APs). Rapid inactivation after Na_V channel opening, known as open-state inactivation, plays a critical role in limiting the AP duration. However, Na_V channel inactivation can also occur before opening, namely closed-state inactivation, to tune the cellular excitability. The voltage-sensing domain (VSD) within repeat IV (VSD-IV) of the pseudotetrameric Na_V channel α-subunit is known to be a critical regulator of Na_V channel inactivation. Yet, the two processes of open-and closed-state inactivation predominate at different voltage ranges and feature distinct kinetics. How inactivation occurs over these different ranges to give rise to the complexity of Na_V channel dynamics is unclear. Past functional studies and recent cryo-electron microscopy structures, however, reveal significant inactivation regulation from other Na_V channel components. In this Hypothesis paper, we propose that the VSD of Na_V repeat III (VSD-III), together with VSD-IV, orchestrates the inactivation-state occupancy of Na_V channels by modulating the affinity of the intracellular binding site of the IFMT motif on the III-IV linker. We review and outline substantial evidence that VSD-III activates in two distinct steps, with the intermediate and fully activated conformation regulating closed-and open-state inactivation state occupancy by altering the formation and affinity of the IFMT crevice. A role of VSD-III in determining inactivation-state occupancy and recovery from inactivation suggests a regulatory mechanism for the state-dependent block by small-molecule anti-arrhythmic and anesthetic therapies.