Cytoplasmic kinases downstream of GPR30 suppress gonadotropin-releasing hormone (GnRH)-induced luteinizing hormone secretion from bovine anterior pituitary cells

GPR30 is known as a membrane receptor for picomolar concentrations of estradiol. The GPR30-specific agonistG1 causes a rapid, non-genomic suppression of gonadotropin-releasing hormone (GnRH)-induced luteinizinghormone (LH) secretion from bovine anterior pituitary (AP) cells. A few studies have recently clarified thatprotein kinase A (PKA) and phosphorylated extracellular signal-regulated kinase (pERK) might be involved incytoplasmic signaling pathways of GPR30 in other cells. Therefore, we tested the hypothesis that PKA and ERKkinase (MEK) are important cytoplasmic mediators for GPR30-associated non-genomic suppression of GnRH-inducedLH secretion from bovine AP cells. Bovine AP cells (n = 8) were cultured for 3 days under steroid-freeconditions. The AP cells were previously treated for 30 min with one of the following: 5000 nM of PKAinhibitor (H89), 1000 nM of MEK inhibitor (U0126), or a combination of H89 and U0126. Next, the AP cells weretreated with 0.01 nM estradiol for 5 min before GnRH stimulation. Estradiol treatment without inhibitorpretreatment significantly suppressed GnRH-induced LH secretion (P < 0.01). In contrast, estradioltreatment after pretreatment with H89, U0126 or their combination had no suppressive effect on GnRH-induced LHsecretion. The inhibitors also inhibited the G1 suppression of GnRH-induced LH secretion. Therefore, thesedata supported the hypothesis that PKA and MEK (thus, also pERK) are the intracellular mediators downstream ofGPR30 that induce the non-genomic suppression of GnRH-induced LH secretion from bovine AP cells by estradiolor G1.