Follicle-Stimulating Hormone and Estradiol Regulate Antrum-like Reorganization of Granulosa Cells in Rat Preantral Follicle Cultures1

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Abstract
Formation of a fluid-filled antrum results from the actions of FSH and estrogen on preantral ovarian follicles in most mammalian species. To investigate the novel proposal that hormone-regulated cell-cell interactions mediate antrum formation, we isolated preantral follicles from infant (10- or 11-day-old) Wistar rats and cultured them in a substratum-adherent manner in Minimum Essential Medium supplemented with 2 mM hypoxanthine, 3 mg/ml bovine serum albumin, 5 .mu.g/ml insulin, 5 .mu.g/ml transferrin, and 5 ng/ml selenium. Similar cultures were previously shown to support oocyte growth and acquisition of meiotic competence. In the absence of FSH, follicles attached to the plastic surface and granulosa cells spread-out uniformly around granulosa cell-enclosed oocytes. FSH treatment caused certain follicles to show an increase between culture days 3 and 7 in appearance of conspicuous antrum-like reorganization of the granulosa cells, but without forming a completely enclosed fluid-filled cavity. This response was biphasic over 10-500 ng/ml FSH, with an optimal concentration of 50 ng/ml resulting in a mean of 37.8 .+-. 4.7% of follicles showing antrum-like reorganization for 3 similar experiments. Estradiol-17.beta. alone at 10-10-10-8 M was without effect on this response, but at 10-10 and 10 -9 M, it significantly augmented the action of an optimal concentration of FSH by about 2-fold in 4 experiments. In these experiments, the effect of 10-8 estradiol was not significantly different from FSH alone, indicating that the response to estradiol was also biphasic. These results show that FSH and estradiol coordinate an antrum-like reorganization of granulosa cells in cultured preantral follicles that is independent of antral fluid accumulation, suggesting the critical importance of hormone-regulated cellular interactions in development of antral follicles.