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

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Abstract
Rapid, nongenomic effects of 17β-estradiol (E2) 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 ERα or ERβ antibodies to label en face rat cerebral and coronary arterial endothelia. Western blots revealed a novel 55-kDa ERα 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 ERα 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. ERβ distribution was predominately intranuclear and epitope independent. A small amount of ERα colocalized with ERβ within the nucleus. The results were identical in both arterial preparations and insensitive to E2. We postulate that the different ERα conformations at the membrane, in the nucleus, and between different cell types allow E2 to trigger cell- and location-specific signaling cascades.