Ephedrine is the preferred vasoconstrictor for the treatment of hypotension after epidural and spinal anesthesia in obstetrics because it preserves uterine perfusion better than pure alpha-adrenergic agonists. Previous studies of uterine vascular rings in vitro suggested that direct uterine vasoconstriction from ephedrine is reduced during pregnancy. This study examined the hypothesis that nitric oxide synthase (NOS) is up-regulated in uterine arteries during pregnancy, and that ephedrine stimulates NOS to release nitric oxide (NO) and diminish direct vasoconstriction. Uterine arterial vessels were obtained from 12 pregnant and 9 nonpregnant ewes, and vessel tension was monitored in vitro in response to escalating concentrations of ephedrine or metaraminol. In some experiments, vascular endothelium was mechanically removed, while in others antagonists of NO synthesis (N omega-nitro-L-arginine methyl ester [L-NAME], NO diffusion (hemoglobin [Hgb]), or guanylate cyclase (methylene blue [MB]) were included. In other experiments, solutions containing ephedrine were superfused over uterine arteries from pregnant ewes onto uterine arteries from nonpregnant ewes. Finally, NOS activity, determined by 14C-citrulline generation, was determined in uterine arteries from pregnant and nonpregnant ewes. Both ephedrine and metaraminol caused concentration-dependent constriction of uterine arterial rings from pregnant and nonpregnant animals. Pregnancy reduced maximum constriction from ephedrine more than metaraminol. Similarly, ephedrine-induced constriction was increased more than that of metaraminol in uterine arteries from pregnant animals treated to diminish the effects of nitric oxide (L-NAME, Hgb, MB, endothelium removal). Ephedrine's constriction of uterine arteries from nonpregnant animals was reduced when it was superfused over uterine arteries from pregnant animals. NOS activity was increased in uterine arteries from pregnant compared to nonpregnant animals. These studies confirm decreased direct uterine arterial vasoconstriction during pregnancy from ephedrine and support the hypothesis of increased release of an endogenous vasodilator (NO), either from the vascular endothelium or the vessel wall, as the cause for this decreased vasoconstriction.