Female Wistar rats, displaying a regular 4-day oestrus cycle, were killed in succession every 2 or 3 h on the day of pro-oestrus until ovulation. The population of preovulatory ovarian follicles was isolated and cultured for 24 h in Eagle's medium or in this medium supplemented with testosterone, or progesterone or aromatase inhibitor (CGS 16949A, Ciba-Geigy). In collected media the released steroids were estimated. The follicles isolated in the morning and afternoon secreted predominantly oestradiol and androgens. In the evening a fall in oestradiol and androgen levels was observed, whereas progesterone production rose, reaching a peak value at 20.00 h. Addition of progesterone suppressed oestradiol release at 18.00 h. CGS 16949A inhibited oestradiol secretion in all the cultures investigated and in some of them also affected progesterone secretion. The presence of testosterone in control cultures changed the progesterone release and stimulated oestradiol production at 20.00 h and 22.00 h, i.e. during oestradiol decline. These results suggest that the fall in oestradiol production is initiated by the lack of aromatizable androgens and this is followed by a suppression of aromatase activity. The role of progesterone as an inhibitor of the aromatase enzyme system is very probable.