Somatomedin-C as an amplifier of follicle-stimulating hormone action: enhanced accumulation of adenosine 3',5'-monophosphate

Somatomedin-C (Sm-C) has recently been found to amplify the FSH-mediated acquisition of granulosa cell progestin biosynthetic capacity, aromatase activity, and LH receptors, an effect distinct from its established replicative property. To further characterize the cellular mechanism(s) underlying the synergistic interaction of Sm-C with FSH, we have set out to evaluate the intermediary role of cAMP in this regard. Isolated granulosa cells from immature hypophysectomized diethylstilbestrol-treated rats were cultured for up to 3 days under serum-free conditions. The basal extracellular accumulation of cAMP remained unchanged in response to treatment with highly purified Sm-C (50 ng/ml). However, concurrent treatment with increasing concentrations (0.3-50 ng/ml) of Sm-C, produced dose- and time-dependent increments in the FSH-stimulated accumulation of cAMP, with an apparent median effective dose (ED50; mean +/- SE) of 5.1 +/- 0.6 ng/ml, a maximal response 8.8-fold greater than that induced by FSH alone, and a minimal time requirement of 1-2 days. Given increasing concentrations of FSH, treatment with a constant concentration (50 ng/ml) of Sm-C resulted in 1.7-, 5.8-, and 4.3-fold increases in cAMP accumulation for 10, 30, and 100 ng/ml FSH, respectively. The ability of Sm-C to augment FSH-stimulated cAMP accumulation was evident and, in fact, enhanced by ZK62711 (Rolipram; 3 X 10(-6) M)-induced blockade of cAMP-phosphodiesterase activity. Decreasing dilutions (1:64,000 to 1:1,000) of a monoclonal antibody raised against Sm-C (sm 1.2) produced progressive and complete immunoneutralization of the synergistic interaction of Sm-C with FSH, suggesting specificity of action. Taken together, these findings suggest that Sm-C, acting at nanomolar concentrations compatible with its granulosa cell receptor binding affinity (0.6-2.0 nM), is capable of amplifying FSH-stimulated cAMP accumulation in a time- and dose-dependent manner. These observations suggest that the synergistic action of Sm-C is exerted, at least in part, at a site(s) proximal to cAMP generation.