The favorable effects of estrogen on cardiovascular diseases can be explained by several mechanisms such as changes in serum lipid profiles and thrombogenecity. Estrogen also affects the vascular tone, but there has been no report in which the effect of estrogen was tested comprehensively for several vasoactive substances, especially after long-term administration. Two weeks after bilateral ovariectomy in 8-week old female Sprague-Dawley rats, placebo or 17 β-estradiol (E₂) pellets (0.5 mg; released over 3 weeks) were implanted subcutaneously. Two weeks after pellet implantation, organ chamber experiments were performed using aortae. Compared with control, E₂-treated vessels showed impaired endothelium-dependent relaxation to acetylcholine. E₂ enhanced the contraction to norepinephrine and U46619 and had no effect on endothelin-1-induced contraction. In contrast, the contraction to angiotensin (AT)-II was inhibited by E₂. Northern blot analysis for AT₁ receptor expression using cultured aortic smooth muscle cells showed no difference between control and E₂-treated cells, suggesting that AT₁ receptor downregulation is not the likely mechanism. These results suggest that E₂ affects the vascular tone variably according to vasoactive substances.