While studying the adult rat skeletal muscle Na+ channel outer vestibule, we found that certain mutations of the lysine residue in the domain III P region at amino acid position 1237 of the α subunit, which is essential for the Na+ selectivity of the channel, produced substantial changes in the inactivation process. When skeletal muscle α subunits (μ1) with K1237 mutated to either serine (K1237S) or glutamic acid (K1237E) were expressed in Xenopus oocytes and depolarized for several minutes, the channels entered a state of inactivation from which recovery was very slow, i.e. the time constants of entry into and exit from this state were in the order of ~100 s. We refer to this process as 'ultra-slow inactivation.' By contrast, wild-type channels and channels with the charge-preserving mutation K1237R largely recovered within ~60 s, with only 20-30% of the current showing ultra-slow recovery. Coexpression of the rat brain β1 subunit along with the K1237E α subunit tended to accelerate the faster components of recovery from inactivation, as has been reported previously of native channels, but had no effect on the mutation-induced ultra-slow inactivation. This implied that ultra-slow inactivation was a distinct process different from normal inactivation. Binding to the pore of a partially blocking peptide reduced the number of channels entering the ultra-slow inactivation state, possibly by interference with a structural rearrangement of the outer vestibule. Thus, ultra-slow inactivation, favored by charge-altering mutations at site 1237 in μ1 Na+ channels, may be analogous to C-type inactivation in Shaker K+ channels.
Bibliographical noteFunding Information:
This work was supported by HL-P01–20592 from the National Institutes of Health and by a grant from the Max Kade Foundation, Inc., New York (to H.T.). Research support was also provided by the Medical Research Council of Canada (MRC), and R.J.F. received salary support as an MRC Distinguished Scientist and a Medical Scientist of the Alberta Heritage Foundation for Medical Research.