Figure 3. Gluconate replacement acts inside the cell to reduce Ca²⁺ channel open probability.

CaV1.2 channel openings were recorded using the cell-attached patch configuration. Inward currents into the cell are shown as downward deflections. Panel A shows a sequence of 4 sweeps in control conditions. Panel B shows that that the number of channel openings dropped dramatically when extracellular Cl⁻ was replaced with gluconate (135 mM). Panel C shows that channel openings recovered after washout. The extracellular channel surface was continuously exposed to 164 mM Cl⁻ in the recording pipette suggesting that the reduction in channel opening are due to intracellular effects of gluconate replacement. For illustration, currents were smoothed by Butterworth filtering (8-pole, 800Hz). D. The amplitude of single channel currents was not significantly reduced by gluconate (N = 7, P = 0.38, paired t-test). C. Channel open probability (NP_o) was significantly reduced by gluconate replacement (N = 7, P<0.0001, paired t-test). Control data were analyzed from sweeps obtained at an estimated trans-membrane potential across the patch of ∼−12 mV. Gluconate sweeps were analyzed using a 10 mV more positive test pulse to compensate for the ∼10 mV depolarization of HEK293 cells produced by the gluconate solution.