The alpha-subunit cDNAs encoding voltage-sensitive sodium channels of human heart (hH1) and rat skeletal muscle (rSkM1) have been expressed in the tsA201 mammalian cell line, in which inactivation properties appear to be normal in contrast to Xenopus oocytes. A series of rSkM1/hH1 chimeric sodium channels has been evaluated to identify the domains of the alpha-subunits that are responsible for a set of electrophysiological differences between hH1 and rSkM1, namely, midpoints and slope factors of steady-state activation and inactivation, inactivation kinetics and recovery from inactivation kinetics and their voltage-dependence. The phenotype of chimeric channels in which each hH1 domain was successively introduced into a rSkM1 alpha-subunit framework confirmed the following conclusions. (i) The D4 and or/C-ter. are responsible for the slow inactivation of hH1 sodium channels. (ii) Concerning the other differences between rSkM1 and hH1: steady-state activation and inactivation, kinetics of recovery from inactivation, the phenotypes are determined probably by more than one domain of the alpha-subunit.