Peroxynitrite reversibly inhibits Ca2+-activated K+ channels in rat cerebral artery smooth muscle cells

Peroxynitrite (ONOO-) is a contractile agonist of rat middle cerebral arteries. To determine the mechanism responsible for this component of ONOO- bioactivity, the present study examined the effect of ONOO- on ionic current and channel activity in rat cerebral arteries. Whole cell recordings of voltage-clamped cells were made under conditions designed to optimize K+ current. The effects of iberiotoxin, a selective inhibitor of large-conductance Ca2+-activated K+ (BK) channels, and ONOO- (10-100 mu M) were determined. At a pipette potential of 150 mV, ONOO- inhibited 39% of iberiotoxin-sensitive current. ONOO- was selective for iberiotoxin-sensitive current, whereas decomposed ONOO- had no effect. In excised, inside-out membrane patches, channel activity was recorded using symmetrical K+ solutions. Unitary currents were sensitive to increases in internal Ca2+ concentration, consistent with activity due to BK channels. Internal ONOO- dose dependently inhibited channel activity by decreasing open probability and mean open times. The inhibitory effect of ONOO- could be overcome by reduced glutathione. Glutathione, added after ONOO-, restored whole cell current amplitude to control levels and reverted single-channel gating to control behavior. The inhibitory effect of ONOO- on membrane K+ current is consistent with its contractile effects in isolated cerebral arteries and single myocytes. Taken together, our data suggest that ONOO- has the potential to alter cerebral vascular tone by inhibiting BK channel activity.