The voltage- and Ca2+-dependent gating mechanism of large-conductance Ca2+-activated K+ (BK) channels from cultured rat skeletal muscle was studied using single-channel analysis. Channel open probability (P(o)) increased with depolarization, as determined by limiting slope measurements (11 mV per e-fold change in P(o); effective gating charge, q(eff), of 2.3 ± 0.6 e(o)). Estimates of q(eff) were little changed for intracellular Ca2+ (Ca2+(i)) ranging from 0.0003 to 1,024 μM. Increasing Ca2+(i) from 0.03 to 1,024 μM shifted the voltage for half maximal activation (V1/2) 175 mV in the hyperpolarizing direction. V(1/2) was independent of Ca2+(i) for Ca2+(i) ≤ 0.03 μM, indicating that the channel can be activated in the absence of Ca2+(i). Open and closed dwell-time distributions for data obtained at different Ca2+(i) and voltage, but at the same P(o), were different, indicating that the major action of voltage is not through concentrating Ca2+ at the binding sites. The voltage dependence of P(o) arose from a decrease in the mean closing rate with depolarization (q(eff) = -0.5 e(o)) and an increase in the mean opening rate (q(eff) = 1.8 e(o)), consistent with voltage-dependent steps in both the activation and deactivation pathways. A 50-state two-tiered model with separate voltage- and Ca2+-dependent steps was consistent with the major features of the voltage and Ca2+ dependence of the single-channel kinetics over wide ranges of Ca2+(i) (~0 through 1,024 μM), voltage (+80 to -80 mV), and P(o) (10-4 to 0.96). In the model, the voltage dependence of the gating arises mainly from voltage-dependent transitions between closed (C-C) and open (O-O) states, with less voltage dependence for transitions between open and closed states (C-O), and with no voltage dependence for Ca2+-binding and unbinding. The two-tiered model can serve as a working hypothesis for the Ca2+- and voltage-dependent gating of the BK channel.
- K(Ca) channel
- Large-conductance Ca-activated K channel
- Monod- Wyman-Changeux
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