Figure 1. BK channel activation is strongly inhibited when pH_O is lower than 5.

(A) Macroscopic BK currents recorded from an outside-out patch perfused with extracellular solutions at pH 7 (left), pH 4 (middle), and back to pH 7. Currents were evoked by steps from −160 to +160 mV at pH_O 7 and by steps from −160 mV to +200 mV at pH_O 4 (20 mV increments). The pre- and post-test pulse voltages were −140 and −120 mV, respectively. 300 μM Ca²⁺ was included in pipette (intracellular) solution. The red traces were evoked by +60 mV pulses to highlight the much larger fractional reduction in outward current at this voltage compared to that at +160 mV (blue traces). (B) G-V curves of BK channel generated from tail currents recorded at various pH_Os with values normalized to the maximum value at pH_O 7. Physiologically relevant voltage range (±60 mV) is marked by two vertical dotted lines here and in panel D. Boltzmann fit results (solid lines) are G_max = 1.06 ± 0.01, z = 0.97 ± 0.03 e, Vh = −48.8 ± 1.0 mV (pH 9), G_max = 0.95 ± 0.01, z = 1.00 ± 0.04 e, Vh = −32.4 ± 1.2 mV (pH 7), G_max = 0.80 ± 0.01, z = 0.97 ± 0.06 e, Vh = −11.7 ± 1.8 mV (pH 6), G_max = 0.66 ± 0.01, z = 0.86 ± 0.05 e, Vh =+34.8 ± 2.3 mV (pH 5), G_max = 0.46 ± 0.02, z = 0.90 ± 0.09 e, Vh =+107 ± 3.7 mV (pH 4). The number in parentheses is the number of experiments contributing to each G-V relationship. (C) Voltage-dependent change of BK single channel current induced by extracellular acidification. The filled dots are fractional BK single channel current amplitude determined over the voltage range of ±100 mV when pH_O changed from 7 to 4. The Woodhull model (refer to Materials and methods) fit results (solid line) are: k_d0 = 0.27 ± 0.02 mM (pH 3.6) and δ = 0.17 e ± 0.02. The prediction of fractional BK single channel current amplitude by the Woodhull model is extended to ±200 mV. Note that the residual single channel current at −120 mV at pH_O 4.0 is predicted to be 0.55 of that at pH_O 7.0, which is comparable to the G_max at pH_O 4 in panel B. (D) BK G-V curves as those shown in panel B, but with the values at each pH_O normalized to the maximum value within that pH_O. Boltzmann fit results (solid lines) are G_max = 0.97 ± 0.01, z = 0.98 ± 0.04 e, V_h = −48.3 ± 1.1 mV (pH 9), G_max = 0.98 ± 0.01, z = 1.05 ± 0.04 e, V_h = −30.5 ± 1.1 mV (pH 7), G_max = 0.96 ± 0.01, z = 1.04 ± 0.05 e, V_h = −14.5 ± 1.2 mV (pH 6), G_max = 0.97 ± 0.01, z = 0.86 ± 0.04 e, V_h =+35.0 ± 1.6 mV (pH 5), G_max = 1.01 ± 0.02, z = 0.90 ± 0.04 e, V_h =+107.6 ± 1.8 mV (pH 4). The V_h and z values are virtually identical to those determined from the G-Vs in Figure 1B. (E) Dose-response curve of BK gating shift (ΔV_h) induced by extracellular H⁺. Hill equation (ΔV_h=ΔV_max/(1+(IC₅₀/[H⁺])ⁿ)) fit result (solid line) is: IC₅₀ = 0.17 ± 0.28 mM (pH 3.8), n = 0.41 ± 0.07.

Figure 1—figure supplement 1. BK channel single channel current at various voltages and pH_Os.

(A–B) Representative current traces for determining BK single channel current amplitude at different conditions. These traces were recorded from different outside-out patches due the large change in P_O induced by the change of voltages and pH_Os. [Ca²⁺]_in was 300 μM for all these experiments. Closed level is marked by dotted line. All four traces are displayed at same scale for comparison. Scale bar: 20 pA vertical, 20 ms horizontal. (A) Single channel activity of BK channels at −100 mV at pH_O 7 (left) or 4 (right). (B) Single channel activity of BK channels at +100 mV at pH_O 7 (left) or 4 (right). (C) BK single channel i-V plot at pH_O 7 (black dots) and 4 (red dots) over the voltage range of ±100 mV. The number in parentheses is the number of recordings for each data point and the error bars are smaller than the data points.