Abstract—Pregnancy is associated with a significant decrease in uterine vascular tone and an increase in uterine blood flow. The present study tested the hypothesis that estrogen and progesterone differentially regulate the extracellular signal–regulated kinase (ERK)1/2 and protein kinase C (PKC) signaling pathways in vascular smooth muscle, resulting in a decrease in uterine vascular myogenic tone in pregnancy. Uterine arteries were isolated from nonpregnant and near-term pregnant sheep. Chronic treatment (48 hours) of nonpregnant uterine arteries with 17-estradiol and progesterone caused a significant decrease in PKC-mediated contractions and pressure-induced myogenic tone. In accordance, treatment of near-term pregnant uterine arteries for 48 hours with ICI 182780 and RU 486 significantly increased PKC-induced contractions and myogenic tone. In contrast, acute treatment for 30 minutes had no effect on uterine artery contractility. An ERK1/2 inhibitor, PD098059, restored the chronic effect of steroids on PKC-mediated contractions in nonpregnant sheep. ERK1/2 protein and mRNA levels were greater in near-term pregnant as compared with nonpregnant uterine arteries. 17-Estradiol and progesterone increased ERK1/2 protein in nonpregnant sheep. In agreement, ICI 182780 and RU 486 caused significant decreases in ERK1/2 protein in near-term pregnant sheep. Western blot showed 6 PKC isozymes, , _I, _II, , , and , in the uterine arteries. 17-Estradiol and progesterone decreased the particulate:cytosolic ratios of PKC, , and , respectively, in nonpregnant sheep. ICI 182780 and RU 486 increased the ratios in near-term pregnant sheep. The results indicate a direct chronic effect of the steroid hormones in the upregulation of ERK1/2 expression and downregulation of the PKC signaling pathway, resulting in attenuated myogenic tone of the uterine artery in pregnancy.