FSH receptor binding inhibitor influences estrogen production, receptor expression and signal pathway during in vitro maturation of sheep COCs.

Follicle-stimulating hormone (FSH) promotes secretion of follicle fluid and follicle development. FSH acts via cognate FSH receptor (FSHR). It remains unknown whether the supplement of FSH-receptor binding inhibitor (FRBI) into the in vitro maturation (IVM)medium influence the estrogen receptor expression and signal pathway of oocytes in sheep. The present study aimed to investigate FRBI effects on inositol trisphosphate (IP3) of oocytes and protein kinase A (PKA) of sheep granulosa cells, further to elucidate the signal pathway of FRBI effects. Cumulus-oocyte complexes (COCs) were recovered from antral follicles. COCs were cultured for 24 h in the IVM medium supplemented with varying concentrations of FRBI (0, 10, 20, 30 and 40 μg/mL) and FSH (10IU/mL). ELISA was used to measure the concentrations of estradiol (E2) and IP3 in the IVM medium. Western blotting was utilized to detect protein expression of ERβ of COCs and protein kinase A (PKA) of granulosa cells. The results showed IP3 concentrations of FRBI-3 and FRBI-4 groups were less than that of CG and FSH groups at 22 h and 24 h (P < 0.05). PKA levels of FRBI-3 and FRBI-4 groups were significantly less than that of CG and FSH group (P < 0.05 or P < 0.01). Expression levels of ERβ mRNA and protein of FRBI-treated groups were gradually decreased in comparison to CG and FSH group. The minimum value was detected in the FRBI-4 group. ERβ protein level of the FRBI-4 group was significantly less than that of FSH group (P < 0.05). E2 concentrations of FRBI-treated groups were elevated as compared to CG, with the highest increment of FRBI-2 group (P < 0.05). Our results revealed a higher dose of FRBI reduced IP3 production. FRBI could suppress slightly expression levels of ERβ mRNA and protein of COCs and PKA of granulosa cells, additionally increased E2 production of sheep COCs.