Peroxynitrite reversibly inhibits Ca(2+)-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 Ca(2+)-activated K(+) (BK) channels, and ONOO(-) (10-100 microM) were determined. At a pipette potential of +50 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 Ca(2+) 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.