Interaction between permeant ions and voltage sensor during inactivation of N-type Ca2+ channels

1. Inactivation of neuronal N-type Ca2+ channels transiently expressed in human kidney tSA-201 cells was studied at the level of whole-cell Ca2+ current and intramembrane charge movement. 2. Prolonged (5 s) depolarization to 40 mV shifted the voltage distribution of intramembrane charge movement from a transition potential (mid-point voltage) of 9.5 +/- 3.8 mV to -55.4 +/- 8.2 mV. Because of the large negative shift, it was possible to record intramembrane charge movement from unblocked inactivated channels and determine the effect of Ca2+ influx on inactivation of intramembrane charge movement. 3. In unblocked channels, the rate of inactivation of charge movement (21 +/- 3 s(-1) at 0 mV) was close to that of Ca2+ current decay during the conditioning pulse. However, in blocked channels inactivation was significantly slower (4 +/- 1 s(-1) at 9 mV). In unblocked channels, the availability of Ca2+ current was minimal and charge movement from inactivated channels was maximal after conditioning to about 10 mV. After the block of ionic current, inactivation of charge movement gradually increased with voltage. 4. Although the rate of Ca2+ current run-down was not affected by 10-15 mu M free Ca2+ in the pipette solution, inactivation of Ca2+ currents during depolarization was about two times faster in high intracellular Ca2+ 5. The present results favour the current-dependent mechanism of inactivation of N-type channels. They also suggest that Ca2+ acting in the permeation pathway and transmembrane voltage are the proximate causes of the same inactivation transitions of voltage sensing moieties in these channels.