In vitro steroid-induced meiosis in Rhinella arenarum oocytes: role of pre-MPF activation.

In this work we showed the relationship between seasonal periods and the response of R. arenarum follicles and oocytes to different steroids. Using in vitro germinal vesicle breakdown (GVBD) assays, we demonstrated that P 4 is the main steroid capable of inducing maturation in R. arenarum oocytes and follicles. In the second part of this work we showed that androgens can activate pre-maturation promoting factors (pre-MPFs) such as P 4 , by cytoplasm microinjection experiments. The results indicated that the steroids assayed induced oocyte and follicle maturation in a dose- and time-dependent manner. In oocytes, P 4 was the most efficient steroid as a maturation inducer (EC 50 of the reproductive period, 6 nM, EC 50 of the non-reproductive period ≅ 30 nM). Androgens (DHEA, dehydroepiandrosterone; T, testosterone; and AD, androstenedione) were less efficient maturation inducers than P 4 (EC 50 reproductive period ≅ 50, 120 and 600 nM respectively). Similar results were obtained with intact follicles in both seasonal periods. Although the response of follicles to the different androgens was variable, in no case was it above the above the response induced by P 4 . Independently of the season, oocytes and follicles incubated in P 4 , P 5 and T underwent GVBD after 6–10 h while oocytes and follicles incubated in DHEA and AD matured more slowly. Furthermore, we demonstrated that microinjection of mature cytoplasm from androgen-treated oocytes is sufficient to promote GVBD in immature recipient oocytes (DHEA, 57 ± 12%; AD, 60 ± 8%; T, 56 ± 13%). Thus, androgens such as DHEA, T and AD are as competent as P 4 to activate pre-MPF.