Slow inactivation determines the availability of voltage-gated sodium channels during prolonged depolarization. Slow inactivation in hNav1.4 channels occurs with a higher probability than hNav1.5 sodium channels; however, the precise molecular mechanism for this difference remains unclear. Using the macropatch technique we show that the Dll S5-S6 p-region uniquely confers the probability of slow inactivation from parental hNav1.5 and hNav1.4 channels into chimerical constructs expressed in Xenopus oocytes. Site-directed mutagenesis was used to test whether a specific region within Dll S5-S6 controls the probability of slow inactivation. We found that substituting V754 in hNav1.4 with isoleucine from the corresponding position (891)in hNav1.5 produced steady-state slow inactivation statistically indistinguishable from that in wild-type hNav1.5 channels, whereas other mutations have little or no effect on slow inactivation. This result indicates that residues V754 in hNav1.4 and 1891in hNav1.5 are unique in determining the probability of slow inactivation characteristic of these isoforms. Exchanging S5-S6 linkers between hNav1.4 and hNav1.5 channels had no consistent effect on the voltage-dependent slow time inactivation constants [τ(V)]. This suggests that the molecular structures regulating rates of entry into and exit from the slow inactivated state are different from those controlling the steady-state probability and reside outside the p-regions.
Vilin, Y. Y., Fujimoto, E., & Ruben, P. C. (2001). A single residue differentiates between human cardiac and skeletal muscle Na+ channel slow inactivation. Biophysical Journal, 80(5), 2221–2230. https://doi.org/10.1016/S0006-3495(01)76195-4