MODIFICATION OF INACTIVATION IN CARDIAC SODIUM-CHANNELS - IONIC CURRENT STUDIES WITH ANTHOPLEURIN-A TOXIN

The site 3 toxin, Anthopleurin-A (Ap-A), was used to modify inactivati on of sodium channels in voltage-clamped single canine cardiac Purkinj e cells at similar to 12 degrees C. Although Ap-A toxin markedly prolo nged decay of sodium current (I-Na) in response to step depolarization s, there was only a minor hyperpolarizing shift by 2.5 +/- 1.7 mV (n = 13) of the half-point of the peak conductance-voltage relationship wi th a slight steepening of the relationship from -8.2 +/- 0.8 mV to -7. 2 +/- 0.8 mV (n = 13). Increases in G(max) were dependent on the choic e of cation used as a Na substitute intracellularly and ranged between 26 +/- 15% (Cs, n = 5) to 77 +/- 19% (TMA, n = 8). Associated with Ap A toxin modification time to peak I-Na occurred later, but analysis of the time course I-Na at multiple potentials showed that the largest e ffects were on inactivation with only a small effect on activation. Co nsistent with little change in Na channel activation by ApA toxin, I-N a tail current relaxations at very negative potentials, where the domi nant process of current relaxation is deactivation, were similar in co ntrol and after toxin modification. The time course of the development of inactivation after Ap-A toxin modification was dramatically prolon ged at positive potentials where Na channels open, However, it was not prolonged after Ap-A toxin at negative potentials, where channels pre dominately inactivate directly from dosed states. Steady state voltage -dependent availability (h(infinity) or steady state inactivation), wh ich predominately reflects the voltage dependence of closed-closed tra nsitions equilibrating with closed-inactivated transitions was shifted in the depolarizing direction by only 1.9 +/- 0.8 mV (n = 8) after to xin modification. The slope factor changed from 7.2 +/- 0.8 to 9.9 +/- 0.9 mV (n = 8), consistent with a prolongation of inactivation from t he open state of Ap-A toxin modified channels at more depolarized pote ntials. We conclude that Ap-A selectively modifies Na channel inactiva tion from the open state with little effect on channel activation or o n inactivation from closed state(s).