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Utilizing paper and gas—liquid chromatography, the pattern of steroid secretion by cultures of equine granulosa, thecal and stromal cells has been studied.

Granulosa cell cultures secreted primarily progesterone and 20-hydroxypregn-4-en-3-one. A granulosa cell culture of 700,000 cells harvested from two large vascular follicles of a mare in oestrus secreted the following amounts of steroid from days 0 to 6 of culture: 561 µg. progesterone, 68 µg. 20-hydroxypregn-4-en-3-one, 5·3 µg. 17-hydroxyprogesterone, 1 µg. androstenedione, 0·12 µg. dehydroepiandrosterone and 2·7 µg. oestradiol. Thecal tissue harvested from the same follicles secreted 29, 3·0, 1·8, 1·4, 1·1 and 0·99 µg., respectively, of the same steroids. All these steroids originated from endogenous precursors or from acetate or cholesterol in the tissue culture medium which contained 15% horse serum, 30% medium '199' and 55% Hanks's solution.

Cultures of granulosa cells harvested from two large vascular follicles of a mare in early oestrus secreted progesterone at a constant rate of 35 pg./cell/day between days 6 and 34 of culture, thus demonstrating that these cells do not dedifferentiate in culture. The secretion of 20-hydroxypregn-4-en-3-one relative to progesterone increased from days 3 to 6 and remained constant from days 6 to 15.