Epitope-dependent localization of estrogen receptorα, but not -β, in en face arterial endothelium

Rapid, nongenomic effects of 17beta-estradiol (E-2) in endothelial cells are postulated to arise from membrane-associated estrogen receptors (ERs), which have not been visualized in vascular tissue. To identify membrane ERs, we used multiple site-directed ERalpha or ERbeta antibodies to label en face rat cerebral and coronary arterial endothelia. Western blots revealed a novel 55-kDa ERalpha isoform. Three-dimensional images of cells labeled with these antibodies and markers for the nucleus and caveolin-1 were acquired with a wide-field microscope, deconvolved, and numerically analyzed. We found ERalpha in the nucleus and cell periphery, where one-third colocalized with caveolin-1. The receptor location was dependent on the epitope of the antibody. Human ovarian surface epithelium produced similar results; but in rat myometrium, the distribution was epitope independent and nuclear. ERbeta distribution was predominately intranuclear and epitope independent. A small amount of ERalpha colocalized with ERbeta within the nucleus. The results were identical in both arterial preparations and insensitive to E-2. We postulate that the different ERalpha conformations at the membrane, in the nucleus, and between different cell types allow E-2 to trigger cell- and location-specific signaling cascades.