Background and Purpose-It has been reported that estrogens modulate peripheral vascular synthesis of vasodilatory hormones, including prostacyclin. If this occurs in the cerebral circulation, it could have important consequences in the modulation of cerebral hemodynamic function and improvement of stroke outcome. We investigated the hypothesis that in vivo 17␤-estradiol treatment of ovariectomized rats increases cerebrovascular prostacyclin production via elevation of the enzymes responsible for prostacyclin synthesis. Methods-Cerebral blood vessels from 17␤-estradiol-treated and nontreated ovariectomized rats were isolated and examined for prostacyclin synthesis by enzyme-linked immunosorbent assay or for protein levels of cyclooxygenase-1, prostacyclin-synthase, and cytosolic phospholipase A 2 by immunoblot analysis. Results-We report that chronic in vivo 17␤-estradiol treatment significantly enhanced basal prostacyclin synthesis in rat cerebral blood vessels by 2.6-fold over control. 17␤-Estradiol treatment also resulted in a 5.1-fold increase of cyclooxygenase-1 protein and a 6.7-fold increase of prostacyclin-synthase protein in the cerebral vasculature. There was no effect of estrogen on levels of cytosolic phospholipase A 2 . Conclusions-Our findings suggest that estrogen influences the biosynthesis of prostacyclin, which may be important in the regulation of cerebral blood flow and thrombosis. This finding may shed light on the mechanisms that govern sex-based differences in cerebrovascular disease. (Stroke. 2002;33:600-605.)

We have previously shown that estrogen treatment increases cerebrovascular cyclooxygenase-1, prostacyclin synthase, and production of prostacyclin. Therefore, vascular tone and prostanoid production were measured to investigate functional consequences of estrogen exposure. Middle cerebral arteries were isolated from ovariectomized female Fischer-344 rats with or without chronic in vivo 17beta-estradiol treatment. In vivo 17beta-estradiol treatment increased cerebral artery diameter; functional endothelium was required for expression of these differences. The nonspecific cyclooxygenase inhibitor indomethacin constricted, whereas arachidonic acid dilated, cerebral arteries from estrogen-treated animals. Estrogen exposure increased production of prostacyclin by cerebral arteries. Conversely, in estrogen-deficient animals, indomethacin dilated and arachidonic acid constricted cerebral blood vessels. This correlated with vasorelaxation following inhibition of the thromboxane-endoperoxide receptor with SQ-29548 but not after selective blockade of thromboxane synthase with furegrelate, suggesting prostaglandin endoperoxide (i.e., PGH2) activity. Removal of the endothelium or selective blockade of cyclooxygenase-1 with SC-560 abolished estrogen-mediated differences in the effects of arachidonate on vessel diameter and on prostacyclin production by cerebral arteries. These data suggest 17beta-estradiol decreases cerebrovascular tone by shifting the primary end product of the endothelial cyclooxygenase-1 pathway from the constrictor prostaglandin PGH2 to the vasodilator prostacyclin. These effects of estrogen may contribute to the heightened thromboresistance and enhanced cerebral blood flow documented in pre-versus postmenopausal women.

Ospina, Jose A., Holly N. Brevig, Diana N. Krause, and Sue P. Duckles. Estrogen suppresses IL-1␤-mediated induction of COX-2 pathway in rat cerebral blood vessels. Am J Physiol Heart Circ Physiol 286: H2010-H2019, 2004. First published December 18, 2003 10.1152/ajpheart.00481.2003.-Interleukin (IL)-1␤ is a potent inducer of inflammatory prostaglandins, which are important mediators of vascular response to cerebral injury, whereas estrogen reduces brain injury in models of ischemic stroke. Thus we examined the effects of in vivo IL-1␤ exposure on cerebrovascular cyclooxygenase (COX)-2 expression and function in an animal model of chronic estrogen replacement. Estrogen-treated and nontreated ovariectomized female rats received IL-1␤ injections (10 g/kg ip), and then cerebral vessels were isolated for biochemical and contractile measurements. In estrogen-deficient rats, IL-1␤ induced cerebrovascular COX-2 protein expression; a peak response occurred 3 h after injection. COX-2 was localized to arterial endothelium using confocal microscopy. IL-1␤ increased PGE2 but not PGI2 production and decreased vascular tone as measured in isolated cerebral arteries; the latter effect was partially reversed by treatment with the selective COX-2 inhibitor NS-398 (10 mol/l). In contrast, in animals treated with estrogen, IL-1␤ had no significant effect on COX-2 protein levels, PGE2 production, or vascular tone. Combined treatment with 17␤-estradiol and medroxyprogesterone acetate also prevented increases in PGE2 production after IL-1␤ treatment, but treatment with 17␣-estradiol had no effect. IL-1␤ induction of COX-2 protein was prevented by treatment with the nuclear factor-B inhibitor caffeic acid phenethyl ester (20 mg/kg ip), and estrogen treatment reduced cerebrovascular nuclear factor-B activity. Estrogen thus has potent anti-inflammatory effects with respect to cerebral vascular responses to IL-1␤. These effects may have important implications for the incidence and severity of cerebrovascular disease.17␤-estradiol; cyclooxygenase-2; prostaglandin E 2; nuclear factor-B; interleukin-1␤

Estrogen alters reactivity of cerebral arteries by modifying production of endothelium-dependent vasodilators. Estrogen receptors (ER) are thought to be involved, but the responsible ER subtype is unknown. ER-alpha knockout (alphaERKO) mice were used to test whether estrogen acts via ER-alpha. Mice were ovariectomized, with or without estrogen replacement, and cerebral blood vessels were isolated 1 mo later. Estrogen increased levels of endothelial nitric oxide synthase and cyclooxygenase-1 in vessels from wild-type mice but was ineffective in alphaERKO mice. Endothelium-denuded middle cerebral artery segments from all animals constricted when pressurized. In denuded arteries from alphaERKO but not wild-type mice, estrogen treatment enhanced constriction. In endothelium-intact, pressurized arteries from wild-type estrogen-treated mice, diameters were larger compared with arteries from untreated wild-type mice. In addition, contractile responses to indomethacin were greater in arteries from wild-type estrogen-treated mice compared with arteries from untreated wild-type mice. In contrast, estrogen treatment of alphaERKO mice had no effect on diameter or indomethacin responses of endothelium-intact arteries. Thus ER-alpha regulation of endothelial nitric oxide synthase and cyclooxygenase-1 pathways appears to contribute to effects of estrogen on cerebral artery reactivity.

Abstract:Quantitative autoradiography was used to compare the binding properties of a7-type nicotinic acetylcholine receptors in fetal and adult rat hippocampus. Whereas there were high levels of 125l-a-bungarotoxin (1251-a-BTX) binding throughout fetal hippocampal field CAl, there was a significant decrease in binding site density in the adult. The affinity of 1251-a-BTX binding, as well as a-cobratoxin and nicotine potency to displace 1251-a-BTX, did not change with age. Addition of Ca2~to the assay buffer did not alter 1251-a-BTX binding, or a-cobratoxin inhibition of 125l-ct-BT)( binding, although it significantly increased nicotine affinity at both ages. The effect of Ca2õn agonist affinity was dose-dependent, with an EC 50 value of 0.25-0.5 mM. Ca 2~also significantly increased the cooperativity of nicotine displacement curves in stratum oriens of the adult, but not in the fetus. These findings indicate that the properties of hippocampal 1251-a-BTX binding sites are largely similar across age. Ca2~selectively enhances the affinity of agonist binding, with no change in antagonist binding. This ionic effect may result from potentiation of agonist binding to a desensitized state of the a7 nicotinic acetylcholine receptor and may represent an important neuroprotective mechanism. Key Words: Autoradiography-a-BungarotoxinDevelopment-Nicotine.