Synthesis of the allylic regulatory steroid, 3α-hydroxy-4-pregnen-20- one, by rat granulosa cells and its regulation by gonadotropins

The production of the allylic regulatory steroid 3α-hydroxy-4-pregnen- 20-one (3αHP) in the rat ovary was examined and compared to progesterone levels through use of specific RIAs that had been validated by capillary gas chromatography-mass spectrometry (GC/MS). Results showed that serum levels of 3αHP are comparable to levels of progesterone at all ages examined. In the 4-day cycling rat, serum levels of 3αHP were highest during diestrus and lowest during proestrus and estrus, while serum FSH levels were highest during proestrus/estrus and lowest during diestrus. Hypophysectomy resulted in decreases in ovarian and serum 3αHP. Treatment of hypophysectomized rats with eCG, but not hCG, increased ovarian and serum 3αHP, while serum progesterone was elevated by treatment with hCG. Ovariectomy resulted in a 55-60% reduction in serum 3αHP, indicating that ovaries are a substantial, but not exclusive, source of 3αHP in serum. As further evidence, cultures of preparations consisting primarily of either granulosa cells or granulosa/theca 'shells' produced 3αHP in time-dependent amounts comparable to those of progesterone. Granulosa cells in culture showed a significant increase in accumulation of 3αHP (and progesterone) due to treatment with FSH, but not LH. In contrast to the granulosa-only cell cultures, follicle shells consisting of theca and granulosa cells responded to either LH or FSH treatment with marked increases in 3αHP; increases resulting from combined treatment (FSH + LH) were significantly greater than those due to each hormone alone, but the increases were not additive. Studies of the metabolism of [14C]progesterone by cultured granulosa cells indicated not only that 3αHP is a direct product of progesterone but also that it can serve as a precursor for 4-ene steroids (such as 4-pregnen-20α-ol-3-one) as well as for the 5α-reduced steroid, 5α-pregnan-3α-ol-20-one. A 4-ene 21-carbon steroid cycle is proposed for ovarian steroidogenesis that includes 3αHP as a pivotal member and suggests an additional new pathway for steroid interconversion in the ovary. Since there is rapid interconversion between progesterone and 3αHP, it is possible that some of the actions (intraovarian and otherwise) previously ascribed to progesterone may be due, wholly or in part, to the actions of 3αHP.