Estrogen (E) has nitric oxide (NO)-mediated effects in certain vascular beds, and fetal E levels rise acutely with parturition, suggesting that E may be involved in NO-mediated pulmonary vasodilation at birth. We tested the hypothesis that E acutely stimulates NO synthase (NOS) activity in ovine fetal pulmonary artery endothelial cells (PAEC) by measuring L-[3H]arginine conversion to L-[3H]citrulline in intact cells. NOS activity in the presence of 17 beta-estradiol (E2 beta) rose in a dose-dependent manner, increasing 70-100%, with a threshold concentration of 10(-10) M. This effect was detectable within 5 min of E2 beta exposure, and the maximal response was comparable to that obtained with acetylcholine, which had a threshold concentration of 10(-8) M. Ca2+ removal completely inhibited E2 beta-stimulated NOS activity, and activity with E2 beta and the Ca2+ ionophore A-23187 was not additive. In addition, the expression of the endothelial isoform of NOS (eNOS) was not altered, and the inducible and neuronal NOS isoforms were not detected by immunoblot analysis. These findings indicate that E2 beta acutely stimulates eNOS by Ca2+ influx. Furthermore, E2 beta-stimulated NOS activity was fully inhibited by the E receptor (ER) antagonists tamoxifen and ICI-182,780, and ER mRNA expression was evident in reverse transcription-polymerase chain reaction assays. Thus E acutely stimulates eNOS activity in fetal PAEC via the activation of endothelial ER and increases in intracellular Ca2+.
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