Paradoxical effect of 3-isobutyl-1-methylxanthine on cytochrome P450 cholesterol side-chain cleavage mRNA accumulation in porcine granulosa cells

Earlier studies in immature porcine granulosa cells cultured in serum-free medium showed dual actions of the protein kinase C activator 12-0-tetradecanoylphorbol-13-acetate (TPA). In cells incubated for 24 h, TPA inhibited follicle-stimulating hormone (FSH)-stimulated cytochrome P450 cholesterol side-chain cleavage (P450(scc)) mRNA accumulation. In contrast, at 4 h, TPA increased P450(scc) mRNA concentration in the absence and presence of FSH or 8-bromo-cAMP; in addition, TPA augmented FSH-stimulated cAMP accumulation. The actions of TPA were then examined in the presence of the phosphodiesterase (PDE) inhibitor, 3-isobutyl-1-methylxanthine (IBMX). With IBMX present, TPA caused a smaller relative augmentation of cAMP accumulation during a 4-h incubation period, suggesting that TPA may both increase cAMP synthesis and inhibit its degradation. The stimulatory effect of FSH or 8-bromo-cAMP on P450(scc) mRNA concentration was not modified by IBMX. However, TPA no longer augmented the FSH- or 8-bromo-cAMP-stimulated P450(scc) mRNA accumulation when IBMX was present. In cells treated with FSH for 24 h, IBMX augmented progesterone production, but paradoxically accentuated the inhibitory effect of TPA on steroidogenesis. These results indicate that IBMX converts TPA from a stimulatory into an inhibitory agent by an action unrelated to cAMP, and points to the need for caution in interpreting experiments with this drug.