Hormone-independent activation of adenylate cyclase in large steroidogenic ovine luteal cells does not result in increased progesterone secretion.

Abstract

The role of cAMP in controlling steroidogenesis in small and large ovine luteal cells was examined. Corpora lutea collected from superovulated ewes (9-11 days postovulation) were dissociated, and the two cell types were purified by elutriation. Both cell types were incubated for 0.5, 1, 2, and 4 h at 37 C with ovine LH (100 ng/ml), cholera toxin (100 ng/ml), or forskolin (50 microM). At each time point, progesterone levels were measured in the medium. Adenylate cyclase activity and cAMP concentrations in the cells and incubation medium were also determined. Progesterone secretion by small cells was significantly stimulated by ovine LH (up to 7.3-fold), cholera toxin (up to 4.2-fold), and forskolin (up to 4.5-fold) during the 4-h incubation. Intracellular levels of cAMP were significantly elevated in the small cells by ovine LH (up to 2.5-fold) and forskolin (up to 5.6-fold). Accumulation of cAMP in medium after incubation of small cells was also significantly stimulated by ovine LH (up to 215-fold), cholera toxin (up to 93-fold), and forskolin (up to 1105-fold). Adenylate cyclase activity, however, was only significantly stimulated by cholera toxin (2.6-fold) and forskolin (3.8-fold). None of the treatments stimulated progesterone secretion by large cells at any time (less than 1.6-fold). Intracellular levels of cAMP in the large cells were not elevated after treatment with ovine LH, but were elevated in cells treated with cholera toxin (up to 2.8-fold) and forskolin (up to 2.6-fold). Accumulation of cAMP in the medium was markedly increased with forskolin treatment (106-fold). Adenylate cyclase activity was found to be significantly stimulated by cholera toxin (2.2-fold) and forskolin (up to 5.1-fold), but not by ovine LH (less than 1.1-fold). Steroid secretion in the small cells appears to be enhanced by elevated intracellular cAMP levels. However, treatments that result in dramatic increases in intracellular levels of cAMP failed to influence the secretion of progesterone in large cells.