This study was to determine whether prostacyclin (prostaglandin I₂; PGI₂) evokes mouse renal vasoconstriction, if so, the underlying mechanism(s) and how its synthesis via cysclooxygenase-1 (COX-1) influences local vasomotor reaction. Experiments were performed on vessels from C57BL/6 mice and/or those with COX-1 deficiency (COX-1^-/-). Results showed that in renal arteries, PGI₂ evoked contraction more potently than in carotid arteries, where COX-1 is suggested to mediate prominent endothelium-dependent contraction. A similar result was observed with the thromboxane-prostanoid (TP) receptor agonist U46619. However, in renal arteries TP receptor antagonism, which inhibited the contraction, did not result in any relaxation in response to PGI₂. Moreover, we noted that the endothelial muscarinic receptor agonist ACh evoked an increase in the production of the PGI₂ metabolite 6-keto-PGF_1α, which was prevented by endothelial denudation or COX-1^-/-. Interestingly, COX-1^-/- was further found to abolish a force development that was sensitive to TP receptor antagonism and result in enhanced relaxation evoked by ACh following NO synthase inhibition. Also, in renal arteries the COX substrate arachidonic acid evoked a vasoconstrictor response, which was again abolished by COX-1^-/-. Meanwhile, non-selective COX inhibition did not show any effect in vessels from COX-1^-/- mice. Thus, in mouse renal arteries high expression of TP receptors together with little functional involvement from the vasodilator PGI₂ receptors results in a potent vasoconstrictor effect evoked by PGI₂. Also, our data imply that endogenous COX-1-mediated PGI₂ synthesis leads to vasoconstrictor activity and this could be an integral part of endothelium-derived mechanisms in regulating local renal vascular function.