The effects of human chorionic gonadotrophin (HCG) and prostaglandin F(2alpha) (PGF(2alpha)) on regulation of human granulosa-luteal cell (GLC) function at different stages of differentiation (day 2 versus day 8 of culture) were studied. Expression of LH receptor mRNA and biosynthesis of progesterone were HCG dependent in human GLC at all stages (n = 6, P < 0.05). Steady-state concentrations of mRNA encoding for FP (a specific high-affinity plasma membrane receptor for PGF(2alpha)) were not dependent on, but were stimulated by, addition of HCG (10 IU/ml) or 8-bromo-cAMP (0.5 mmol/l) (n = 6, P < 0.05). Treatment with PGF(2alpha) (100 nmol/l) decreased FP mRNA concentration, but had no effect on LH receptor and cyclo oxygenase-2 (COX-2) expression on day 2 of cultured GLC (n = 8). As a result, the progesterone biosynthesis by GLC was not affected. On day 8, PGF(2alpha) induced FP and PGHS-2 expression and at the same time decreased LH receptor expression, resulting in inhibition of progesterone output by GLC. Our data demonstrated that early stage GLC (day 2 of culture) are resistant to PGF(2alpha)-induced inhibition of progesterone synthesis but underwent further differentiation and acquired luteolytic capacity after 8 days culture in vitro. We conclude that, via distinct gene regulation at different stages of differentiation, human GLC may become resistant or susceptible to PGF(2alpha)-induced luteolysis.