Activation of estrogen receptor beta is a prerequisite for estrogen-dependent upregulation of nitric oxide synthases in neonatal rat cardiac myocytes

Physiological effects of estrogen on myocardium are mediated by two intrace llular estrogen receptors, ER alpha and ER beta, that regulate transcriptio n of target genes through binding to specific DNA target sequences. To defi ne the role of ERP in the transcriptional activation of both endothelial (e NOS) and inducible nitric oxide synthase (MOS) in cardiac myocytes, we used the complete ERP-specific antagonist R,R-tetrahydrochrysene (R,R-THC). R,R -THC inhibited activation of iNOS/eNOS promoter-luciferase reporter constru cts (iNOS/eNOS-Luc) in a dose-dependent fashion in COS7 cells selectively t ransfected with ER beta, but failed to influence ER alpha -mediated increas e of iNOS/ eNOS-Luc. In neonatal rat cardiomyocytes transfected with eNOS-L uc or iNOS-Luc, incubation with 17 beta -estradiol (E2, 10(-8) M) for 24 h stimulated expression of eNOS and iNOS. R,R-THC (10(-5) M) completely inhib ited this effect. Furthermore, eNOS and iNOS protein expression in cardiac myocytes induced by E2 was completely blocked by R,R-THC as shown by immuno blot analysis. Taken together, these results show that FRO mediates transcr iptional activation of eNOS and iNOS by E2. (C) 2001 Federation of European Biochemical Societies. Published by Elsevier Science B.V. All rights reser ved.