Pregnancy is associated with several endocrine and hemodynamic adaptations that result in increased cardiac output (CO) and decreased total peripheral resistance (TPR) and blood pressure (BP). Pregnancy-stimulated elevations in estradiol can (among several vasodilatory mechanisms) attenuate inward Ca2+ currents and contribute to decreased TPR. Conversely, endogenous cardiotonic steroids (CTS) raise blood pressure (BP) via inhibition of α2 Na+ pumps, which in turn, increase Ca2+ influx mediated by vascular sodium calcium exchangers (NCX1.3) and transient receptor potential channels (TRPCs). Several factors (i.e., adrenocorticotropin, angiotensin II, and plasma volume expansion) that stimulate the biosynthesis and secretion of CTS, particularly endogenous ouabain (EO), are elevated in pregnancy. This would suggest that normal pregnancy is associated with elevated EO. Although circulating CTS are super-elevated in pregnancy-induced hypertension (PIH) and in pre-eclampsia (PE), their significance in normal pregnancy, where BP is low, is paradoxical. My hypothesis is that pregnancy-stimulated factors augment EO biosynthesis and secretion, but increased estradiol levels prevent EO-mediated hypertension by downregulating NCX expression, and thereby, attenuating NCX-mediated Ca2+ influx. Combined liquid chromatography-mass spectrometry confirm that pregnancy is per se a state with elevated circulating EO and is associated with two previously unidentified isomers of EO, yet BP is reduced compared to non-pregnant controls. The blunted pressor response to EO is mediated by estradiol, and can be initiated by the activation of a novel G protein coupled estrogen receptor, GPR30. Nonspecific activation of estrogen receptors by estradiol and specific activation of GPR30 by the selective GPR30 agonist, G-1, prevented ouabain-induced hypertension and reduced NCX expression by >30% in the aorta and mesenteric artery in virgin rats. When pregnant rats were administered exogenous ouabain that (super) elevated their circulating ouabain to levels that would be observed in PE, BP declined to the same extent as the pregnant controls. Despite its inability to elevate BP in pregnant rats, ouabain infusion significantly reduced fetal and placental mass and was associated with fetal death and resorption. Taken together, my studies demonstrate that the pregnancy-triggered elevation in estradiol is vasoprotective; it reduces BP by changing the functional relationship between EO, NCX, cell Ca2+ and vascular tone. The key changes involve reduced NCX-mediated Ca2+ entry, and suppressed sensitivity of the contractile machinery to calcium. Abnormalities in the effect of estradiol may contribute to PIH and/or PE.