Activation of meiotic maturation in rat oocytes after treatment with follicular fluid meiosis-activating sterol in vitro and ex vivo

Abstract

Meiosis-activating sterols (MAS) have been found to induce meiotic maturation in mouse oocytes in vitro. In the present study we have extended these observations by investigating the effects of follicular fluid MAS (FF-MAS) on rat oocyte maturation in vitro and ex vivo. Rat oocytes freed from their follicles were cultured with FF-MAS (0 μM, 1 μM, 3 μM, 10 μM, 30 μM) for 22 h in a medium containing the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX; 250 μM). A dose-dependent significant increase in germinal vesicle breakdown (GVB) was observed after adding FF-MAS to the culture medium in both cumulus-enclosed (CEO) and denuded (DO) oocytes. A time course study (0, 3, 8, 14, and 22 h) showed a significant increase in GVB after 14 h when DO and CEO were cultured in the presence of 10 μM FF-MAS + 250 μM IBMX. Furthermore immature rats were primed with eCG (20 IU) and 48 h later perfused ex vivo for 12 h in a recirculating system with either FF-MAS (0 μM, 10 μM, 30 μM, 60 μM), cholesterol (60 μM), or LH (0.2 μg/ml) in the presence of 200 μM IBMX, respectively. In addition, ovarian perfusion was carried out with FF-MAS (30 μM, 60 μM) or 0.2 μg/ml LH in the absence of IBMX. After 12 h, oocytes were freed from the ovaries and checked for GVB. By using the ex vivo perfused rat ovary, we found that FF-MAS, starting at 30 μM, was dose-dependently able to overcome IBMX-induced meiotic arrest leading to a comparable increase in GVB as was observed for LH. Furthermore, it was found that FF-MAS in the absence of IBMX was also able to induce meiotic maturation. Our data are consistent with the notion that the maturation-inducing effects of FF-MAS are mediated by different mechanisms compared to spontaneous maturation.