Abstract 5681: GPR30 is Required for the Pro-apoptotic Effects of Estrogen in Vascular Smooth Muscle Cells

Abstract

Rapid and non-genomic effects of estradiol (EST) may be mediated through the G-Protein-coupled receptor dubbed GPR30 receptor. The present studies tested the hypothesis that GPR30 expression, which we demonstrate is readily detectable in freshly isolated vascular tissue but is decreasingly detectable in cultured VSMCs, mediates the effects of EST to alter ERK phosphorylation status and apoptosis. In freshly isolated aortic tissue, EST-stimulated ERK phosphorylation whereas in cultured VSMCs, EST inhibits ERK phosphorylation in a dose-dependent manner. However, gene transfer of GPR30 resulted in EST-mediated stimulation of ERK phosphorylation (138±4%, n=10), which is in contrast to the effects of EST in GFP-expressing primary culture of VSMCs (68±3%, n=14). EST-mediated stimulation of ERK subsequent to heterologous GPR30 expression was pertussis toxin-sensitive and PI3 kinase-dependent; under these conditions, EST also inhibited PKA activation. In contrast, in the absence of GPR30 expression in cultured VSMC, EST stimulated PKA activity and inhibited ERK phosphorylation. We also assessed the impact of GPR30 expression on EST-mediated regulation of VSMCs apoptosis. In cultured VSMC transduced with GFP vector (control conditions), EST (10 nM) mediated a decrease in VSMC apoptosis (80±2%, n=7), an effect that was marginally but significantly enhanced by gene transfer of ER α (71±3%, n=7). The anti-apoptotic effect of EST was attenuated by co-incubation with the PKA inhibitor Rp-cAMP. However, PI3 kinase inhibition did not affect estradiol’s anti-apoptotic actions. In contrast, gene transfer of GPR30 into the VSMC resulted in enhancement of apoptotic rates following EST treatment (169±2%, n=7), and this GPR30-mediated pro-apoptotic effect of EST was attenuated by the PI3 kinase inhibitor, but not by Rp-cAMP. Thus, the effect of EST on vascular smooth muscle cell apoptosis is likely dependent on the balance between ER-mediated PKA activation and GPR30-mediated PKA inhibition and PI3 kinase activation. Taken together, we postulate that modulation of GPR30 expression and/or activity may be an important determinant of the effects of estradiol in the vasculature.