Estradiol-mediated ERK phosphorylation and apoptosis in vascular smooth muscle cells requires GPR 30

Abstract

Recent studies suggest that the rapid and nongenomic effects of estradiol may be mediated through the G protein-coupled receptor dubbed GPR30 receptor. The present study examines the role of GPR30 versus a classical estrogen receptor (ERalpha) in mediating the growth regulatory effects of estradiol. GPR30 is readily detectable in freshly isolated vascular tissue but barely detectable in cultured vascular smooth muscle cells (VSMC). In freshly isolated aortic tissue, estradiol stimulated extracellular signal-regulated kinases (ERK) phosphorylation. In contrast, in cultured VSMC, where GPR30 expression is significantly reduced, estradiol inhibits ERK phosphorylation. Transfer of the genes encoding GPR30 led to estradiol stimulation of ERK phosphorylation, which is opposite the effects of estradiol in the primary culture of VSMCs. Transduction of the mineralocorticoid receptor (MR) had no effect on estradiol effects on ERK. Estradiol-mediated stimulation of ERK subsequent to heterologous GPR30 expression was pertussis toxin sensitive and phosphoinositide 3-kinase (PI3 kinase) dependent; under these conditions, estradiol also inhibited protein kinase A (PKA). In contrast, in the absence of GPR30 expression in cultured VSMC, estradiol stimulated PKA activity and inhibited ERK phosphorylation. To determine the functional effect of GPR30 (vs. estrogen receptor expression), we assessed estradiol-mediated apoptosis. In the absence of GPR30 expression, estradiol inhibited apoptosis. This effect was enhanced with ERalpha expression. In contrast, with GPR30 expression, estradiol stimulated apoptosis in an ERK-dependent manner. Thus the effect of estradiol 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 or activity may be an important determinant of the effects of estradiol in the vasculature.

Hetero-oligomeric Complex between the G Protein-coupled Estrogen Receptor 1 and the Plasma Membrane Ca2+-ATPase 4b

“…4C). These results are consistent with previous studies showing that estrogen induced ERK1/2 phosphorylation via GPER/ GPR30 (1,43). The observed effect of G-1 to inhibit PMCA activity via tyrosine phosphorylation of the pump is quite consistent with previous studies demonstrating a clear inhibitory role of phosphorylation of Tyr-1176 on PMCA4b activity (34,44,45).…”

Extent of Vascular Remodeling Is Dependent on the Balance Between Estrogen Receptor α and G-Protein–Coupled Estrogen Receptor

“…30 In contrast, E2 acting via GPER was proapoptotic. However, the in vivo implications of these receptor-specific opposing effects of estradiol on growth regulation have yet to be established.…”

“…This synthetic phenotype is more similar to the phenotype of VSMCs maintained in primary culture, 35 the model in which we have initially characterized the role of GPER versus ERα in regulating growth. 30 Previous studies have suggested an important role of estrogen in the response to vascular injury 36 including inhibition of neointimal proliferation 37,38 and inhibition of the adventitial inflammatory response. 39,40 It has been suggested that the primary estrogen-mediated response occurred via ERα activation.…”

“…Our initial focus on ERα (versus ER β , which is also expressed in rat aortic vascular smooth muscle cells [VSMCs]) was based on our demonstration that in isolated rat VSMCs, estradiol's (E2) antiapoptotic effects were mediated via ERα. 30 In contrast, E2 acting via GPER was proapoptotic. However, the in vivo implications of these receptor-specific opposing effects of estradiol on growth regulation have yet to be established.…”

“…The assessment of the extent of GPER expression in injured versus uninjured vessels at 7 days after the surgery and the validation of expression of flag-tagged adenoviral constructions at 4 days after the surgery were based on immunoblotting using an anti-GPER antibody (MBL, cat#LS-A4272, lot#8970; Woburn, MA) and an antiflag M2 antibody (Sigma-Aldrich, cat#F7425) respectively, as we have previously described. 30 Secondary antibodies were obtained from Sigma-Aldrich (cat#A0545). …”