Channels that activate in the presence of paxilline gate with normal voltage dependence and kinetics. (A) G-V curves were normalized to the maximal conductance observed in control saline. Currents were activated with 10 µM cytosolic Ca²⁺ from a holding potential of 0 mV. For prepaxilline, V_h = 16.31 ± 0.15 mV (z = 0.87 ± 0.01 e); in 100 nM paxilline, V_h = 24.75 ± 0.13 mV (z = 0.86 ± 0.01 e); and, after washout of paxilline, V_h = 28.0 ± 0.19 mV (z = 0.88 ± 0.01 e). (B) G-V curves from A were normalized to the fitted maximum for each condition to highlight the lack of G-V shift. (C) G-V curves were generated in the presence and absence of paxilline, but with all solutions containing 10 mM DTT (n = 7 patches). For control saline, V_h = 25.04 ± 1.2 mV (z = 0.86 ± 0.06 e); with 50 nM paxilline, V_h = 21.21 ± 1.85 (z = 0.79 ± 0.07 e); and after wash, V_h = 28.65 ± 9.0 (z =1.13 ± 0.05 e). (D) The same G-V curves in C are normalized to the maximal conductance in each case to highlight their similarity. (E) 2-ms steps to 160 mV were applied at 0.2 Hz to monitor BK activation with 300 µM Ca²⁺, while holding the patch at −80 mV. The application of 100 nM paxilline results in slow diminution of BK current with a time constant (fitted red line) of 9.6 ± 1.1 s. (F) For the same patches as in E, steps to 160 mV with 300 µM Ca²⁺ were used to monitor recovery from paxilline inhibition, with recovery at 300 µM Ca²⁺ at −80 mV with the indicated recovery time constant (red line). (G) For a single patch, BK current activation (closed circles) was monitored at 160 mV and deactivation (open circles) at −80 mV with 300 µM Ca²⁺ before and during the application of 20 nM paxilline, showing that paxilline does not alter the kinetics of channels that open and close in the presence of paxilline. (H) A patch was held at 0 mV and 500 nM paxilline was applied at either of two different Ca²⁺ concentrations (10 or 100 µM). The onset of paxilline block is more rapid with 10 µM Ca²⁺.