Inhibition of K(+) channels might mediate renal vasoconstriction. As inhibition of a single type of K(+) channel caused minor or no renal vasoconstriction in vivo in rats, we hypothesized that several classes of K(+) channels must be blocked to elicit renal vasoconstriction. We measured renal blood flow (RBF) in vivo in anesthetized Sprague-Dawley rats. Test agents were infused directly into the renal artery to avoid systemic effects. Inhibition of BK(Ca) and K(ir) channels (with TEA and Ba(2+), respectively) caused small and transient reductions in RBF (to 93¿±¿2% and 95¿±¿1% of baseline, respectively). K(ATP), SK(Ca) or K(v) channel blockade (with glibenclamide, apamin and 4-aminopyridine, respectively) was without effect. However, a cocktail of all blockers caused a massive reduction of RBF (to 15¿±¿10% of baseline). Nifedipine and mibefradil abolished and reduced, respectively, this RBF reduction. The effect of the cocktail of K(+) channel blockers was confirmed in mice using the isolated blood-perfused juxtamedullary nephron preparation. A cocktail of K(+) channel openers (K(+), NS309, NS1619 and pinacidil) had only a minor effect on baseline RBF in vivo in rats, but reduced the vasoconstriction induced by bolus injections of norepinephrine or angiotensin II (by 33¿±¿5% and 60¿±¿5%, respectively). Our results indicate that closure of numerous types of K(+) channels could participate in the mediation of agonist-induced renal vasoconstriction. Our results also suggest that renal vasoconstriction elicited by K(+) channel blockade is mediated by nifedipine-sensitive Ca(2+) channels and partly by mibefradil-sensitive Ca(2+) channels.