N-type calcium channel inactivation probed by gating-current analysis

N-type calcium channels inactivate most rapidly in response to moderate, no t extreme depolarization. This behavior reflects an inactivation rate that bears a U-shaped dependence on voltage. Despite this apparent similarity to calcium-dependent inactivation, N-type channel inactivation is insensitive to the identity of divalent charge carrier and, in some reports, to the le vel of internal buffering of divalent cations. Hence, the inactivation of N -type channels fits poorly with the "classic" profile for either voltage-de pendent or calcium-dependent inactivation. To investigate this unusual inac tivation behavior, we expressed recombinant N-type calcium channels in mamm alian HEK 293 cells, permitting in-depth correlation of ionic current inact ivation with potential alterations of gating current properties. Such corre lative measurements have been particularly useful in distinguishing among v arious inactivation mechanisms in other voltage-gated channels. Our main re sults are the following: 1) The degree of gating charge immobilization was unchanged by the block of ionic current and precisely matched by the extent of ionic current inactivation. These results argue for a purely voltage-de pendent mechanism of inactivation. 2) The inactivation rate was fastest at a voltage where only similar to 1/3 of the total gating charge had moved. T his unusual experimental finding implies that inactivation occurs most rapi dly from intermediate closed conformations along the activation pathway, as we demonstrate with novel analytic arguments applied to coupled-inactivati on schemes. These results provide strong, complementary support for a "pref erential closed-state" inactivation mechanism, recently proposed on the bas is of ionic current measurements of recombinant N-type channels (Patil et a l., 1998. Neuron. 20:1027-1038).