Patch-clamp recording of K+ currents

Whole-cell K⁺ currents were measured in freshly dissociated smooth muscle cells using the standard (ruptured-patch) or perforated patch-clamp method as previously described.^{15, 21} The pipette solution contained (in mmol/L) 90 potassium aspartate, 30 KCl, 20 NaCl, 1 MgCl₂, 1 Mg-ATP, 1 EGTA, and 10 HEPES (pH 7.2 with KOH). The bath solution was composed of (in mmol/L) 140 NaCl, 5 KCl, 0.1 CaCl₂, 1 MgCl₂, 5 glucose, and 10 HEPES (pH 7.4 with NaOH). Paxilline (100 nmol/L) was added in the bath solution to further minimize BK_Ca currents and thus allow relative isolation of K_V currents. Unless otherwise stated, all chemicals were applied to the bath through perfusion. Experiments were performed at room temperature.