Roles of gonadotropins and meiosis-activating sterols in meiotic resumption of cultured follicle-enclosed mouse oocytes

This study describes a model for short-term culture of intact mouse follicles under serum-free conditions. Follicles were either obtained from immature mice receiving no ovarian stimulation (i.e. no eCG-primed protocol, group I) or from mice undergoing ovarian stimulation (i.e. eCG-primed protocol, group II). Follicles were grouped according to size (100-170, 180-200, 210-250, 260-350 and 360-400 microm, respectively) and cultured for 24h (group I) or for only 6h (group II). Induced meiotic resumption of follicle-enclosed oocytes were evaluated following stimulation with gonadotropins (i.e. FSH and hCG), AY9944-A-7, an inhibitor of Delta14-reductase, and RS-21745, an inhibitor of lanosterol 14alpha-demethylase; both enzymes affect synthesis of the meiosis activating sterols (MAS) that induce oocyte maturation. The frequency of oocyte degeneration was also recorded. In group I, FSH (10-200 IUl-1) and AY9944-A-7 (5, 25 and 50 microM) separately induced resumption of meiosis in oocytes derived from follicles with a diameter of 180-400 microm. hCG (1.0 and 10 IUml-1) exhibited a similar but weaker effect on oocytes present in follicles with a diameter of 260-400 microm. Irrespective of follicular diameter oocytes obtained from follicles in group II responded to hCG and FSH by resuming meiosis. FSH (50 IUl-1) alone or hCG (10 IUml-1) alone both increased the GVBD percentage of oocytes enclosed in follicles with a diameter 260-400 microm, but the response to hCG was not significant compared to control. FSH (50 IUl-1) combination with hCG (10 IUml-1) showed an additive effect raising the rate of GVBD after 6h culture. Addition of 50 or 100 microM RS-21745 was able to attenuate gonadotropins-induced resumption of meiosis to below background levels. In conclusion, the ability of FSH to induce meiotic resumption of follicle-enclosed mouse oocytes is correlated to follicle size, being most pronounced in larger follicles. hCG caused a similar but less pronounced effect. The ability of RS-21745 to inhibit and the ability of AY9944-A-7 to enhance oocyte maturation of follicle-enclosed oocytes support the concept of FSH employing MAS as a downstream signal transduction molecule for initiation of oocyte maturation in mice.