TRESK background potassium channel is not gated at the helix bundle crossing near the cytoplasmic end of the pore
Kv1.3 channels were expressed in HEK293T cells. The currents were recorded in inside-out excised patches. Voltage-dependent currents were measured by depolarizing the membrane to +40 mV for 250 ms. The holding potential was -80 mV. Currents were measured at the end of the +40 mV voltage step. Kv1.3 current was calculated by subtracting the current measured in K+-free bath solution from the value measured in the high K+ solution. The currents after the administration of Ba2+ were normalized to the value measured before the application of the blocker (on panel C). A, Representative recording showing the application of Ba2+ (1 mM) to the intracellular surface of the patches while keeping the holding potential at -80 mV (constant hyperpolarization). Afterwards the blocker was removed from the bath solution and the current was measured every 30 s by depolarizing steps to +40 mV. B, Representative recording showing the effect of Ba2+ (1 mM) applied to the intracellular surface of inside-out patches when the membrane was depolarized to +40 mV (transient depolarization). Barium was then removed from the bath solution while the holding potential was -80 mV. Voltage-gated K+ currents were measured every 30 s, as in panel A. C, The normalized currents after the application of Ba2+ are plotted as a function of time. The channels were opened by transient depolarization to +40 mV (black circles) or closed at constant -80 mV (grey squares) in the presence of Ba2+. Data are plotted as scatter plot, the averages of the different groups are plotted as columns. The differences between the groups were significant at all examined time points (p<0.05, statistical analysis was performed using Kruskal-Wallis ANOVA followed by pairwise comparison using the Mann-Whitney U test).