Gender differences in the incidence of stroke and migraine appear to be related to circulating levels of estrogen; however, the underlying mechanisms are not yet understood. Using resistance-sized arteries pressurized in vitro, we have found that myogenic tone of rat cerebral arteries differs between males and females. This difference appears to result from estrogen enhancement of endothelial nitric oxide (NO) production. Luminal diameter was measured in middle cerebral artery segments from males and from females that were either untreated, ovariectomized (Ovx), or ovariectomized with estrogen replacement (Ovx + Est). The maximal passive diameters (0 Ca2+ + 1 mM EDTA) of arteries from all four groups were identical. In response to a series of 10-mmHg step increases in transmural pressure (20-80 mmHg), myogenic tone was greater and vascular distensibility less in arteries from males and Ovx females compared with arteries from either untreated or Ovx + Est females. In the presence of NG-nitro-L-arginine methyl ester (L-NAME; 1 microM), an NO synthase inhibitor, myogenic tone was increased in all arteries, but the differences among arteries from the various groups were abolished. Addition of L-arginine (1 mM) in the presence of L-NAME restored the differences in myogenic tone, suggesting that estrogen works through an NO-dependent mechanism in cerebral arteries. To determine the target of NO-dependent modulation of myogenic tone, we used tetraethylammonium (TEA; 1 mM) to inhibit large-conductance, calcium-activated K+ (BKCa) channels. In the presence of TEA, the myogenic tone of arteries from all groups increased significantly; however, myogenic tone in arteries from males and Ovx females remained significantly greater than in arteries from either untreated or Ovx + Est females. This suggests that activity of BKCa channels influences myogenic tone but does not directly mediate the effects of estrogen. Estrogen appears to alter myogenic tone by increasing cerebrovascular NO production and/or action.