Because premenopausal women have lower cardiovascular morbidity than postmenopausal women, it has been proposed that estrogen may have a protective role. Estrogen is involved in smooth muscle relaxation both through its specific receptor as well as through calcium channel blockade. This study examined the acute effect of estradiol on invasive cardiovascular hemodynamics in 18 postmenopausal women (age 62.6 ± 7.6 years, means ± SD). The effect of estradiol on left ventricular chamber performance was studied in 9 women using simultaneous left ventricular pressure-volume recordings. In a further group of 9 women, the acute effect of estradiol on arterial function was assessed using input impedance (derived from simultaneous aortic pressure and flow recordings), pressure waveform analysis, and pulse wave velocity. After 2 mg micronized 17-estradiol was administered, serum estradiol levels increased from 50.9 ± 21.9 to 3,190 ± 2,216 pmol/l, P < 0.0001. There was no effect of estradiol on either left ventricular inotropic or lusitropic function. There was no acute effect of estradiol on arterial impedance, reflection coefficient, augmentation index, or pulse wave velocity. There was a trend to decreased heart rate and cardiac output in both groups of 9 women. Because heart rate and cardiac output were common to both hemodynamic data sets, results for these parameters were pooled. Across all 18 women, there was a small but significant decrease in heart rate (69.2 ± 10.4 vs. 67.2 ± 9.9 beats/min, P = 0.02), as well as a significant decrease in cardiac output (4.82 ± 1.77 vs. 4.17 ± 1.56 l/min, P = 0.002). Despite achieving supraphysiological serum levels, this study found no significant effect of acute 17-estradiol on ventricular or large artery function.