We hypothesized that voltage-gated K+ (K-v) currents regulate the resting membrane potential (E-m), and that serotonin (5-HT) causes E-m depolarization by reducing K-v currents in rat mesenteric artery smooth muscle cells (MASMCs). The resting E-m was about -40 mV in the nystatin-perforated patch configuration, and the inhibition of K-v currents by 4-aminopyridine caused marked E-m depolarization. The inhibition of Ca2+-activated K+ (K-Ca) currents had no effect on E-m. 5-HT (I mu M) depolarized E-m by similar to 11 mV and reduced the K-v currents to similar to 63% of the control at -20 mV. Similar 5-HT effects were observed with the conventional whole-cell configuration with a weak Ca2+ buffer in the pipette solution, but not with a strong Ca2+ buffer. In the presence of tetraethylammonium (I mM), 5-HT caused E-m depolarization similar to the control condition. These results indicate that the resting E-m is largely under the regulation of K-v currents in rat MASMCs, and that 5-HT depolarizes E-m by reducing K-v currents in a [Ca2+](i)-dependent manner. (c) 2006 Elsevier Inc. All rights reserved.