Functional eukaryotic voltage-gated Na+ (NaV) channels comprise four domains (DI-DIV),
each containing six membrane-spanning segments (S1-S6). Voltage sensing is accomplished
by the first four membrane-spanning segments (S1-S4), which together form a voltage-sensing
domain (VSD). A critical NaV channel gating process, inactivation, has previously
been linked to activation of the VSDs in DIII and DIV. Here, we probe this interaction
by using voltage-clamp fluorometry to observe VSD kinetics in the presence of mutations
at locations that have been shown to impair NaV channel inactivation. These locations
include the DIII-DIV linker, the DIII S4-S5 linker, and the DIV S4-S5 linker. Our
results show that, within the 10-ms timeframe of fast inactivation, the DIV-VSD is
the primary regulator of inactivation. However, after longer 100-ms pulses, the DIII-DIV
linker slows DIII-VSD deactivation, and the rate of DIII deactivation correlates strongly
with the rate of recovery from inactivation. Our results imply that, over the course
of an action potential, DIV-VSDs regulate the onset of fast inactivation while DIII-VSDs
determine its recovery.
