Activation of a 4-AP–sensitive K⁺ channel by EGF stimulation. (A) Effect of EGF on the 4-AP–sensitive K⁺ current. The membrane potential was depolarized from a holding potential of −60 mV to +80 mV at 20-mV increments with a pulse protocol showing in the top panel. Whole-cell currents were recorded from (1) ML-1 cells in the absence (control) and presence of 2 mmol/L 4-AP (as indicated); and (2) ML-1 cells simulated with 10 ng/mL EGF in the absence (EGF) and presence of 4-AP (EGF + 4-AP). (B) Current-voltage relationship of the 4-AP–sensitive K⁺current activated by EGF in the absence and presence of 4-AP. (C) Time course of EGF-activated K⁺ current in the absence and presence of 4-AP. Currents were normalized as I_EGF/I_C , where I_EGF represents amplitudes of the EGF-induced K⁺ current and I_C represents the K⁺ current measured from control ML-1 cells. (D) Single-channel recording of K⁺ channel in ML-1 cells. Inward current recorded as a downward deflection was obtained from cell-attached patches at a membrane potential of −60 mV in the symmetrical 140/140 mmol/L KCl condition. EGF (50 ng/mL) was directly applied to the patch chamber to activate K⁺ channels in the same patch. (E) Current trace demonstrates that application of 100 μmol/L 4-AP in the patch pipette prevented the EGF-induced increase of K⁺-channel activity. Channel activity (NPo) was plotted as a function of time in the lower portion of D and E. (F) Statistics of K⁺-channel activity stimulated by EGF and FBS in the absence and presence of 100 μmol/L 4-AP. Vertical bars represent mean K⁺-channel activity (horizontal bars represent standard error of the mean [SE]). *Significant difference (statistical tests: ANOVA and Tukey, P < .001). Data were collected from five independent experiments.