PROTEIN-KINASE-C SIGNAL-TRANSDUCTION PATHWAY IN ACTH-INDUCED GROWTH EFFECT OF RAT ADRENOCORTICAL-CELLS IN PRIMARY CULTURE

ACTH exerts a biphasic effect on the growth of fetal rat adrenocortical cells in primary culture when bromodeoxyuridine (BrdU) incorporation is used as an indicator of proliferation. The immediate inhibitory effect during the first 24 h of ACTH stimulation is not dependent on cyclic AMP (cAMP). Protein kinase C (PKC) inhibitors H-7 and staurosporine blocked this inhibitory effect of ACTH, whereas 12-0-tetradecanoyl phorbol-13-acetate (TPA; a PKC activator) mimicked the ACTH-induced antimitogenic effect. The stimulatory growth effect of ACTH appears after 72 h of treatment. A similar mitogenic effect is also achieved with cAMP derivative 8-bromo cAMP (8-Br cAMP). However, both ACTH- and 8-Br cAMP-induced proliferations could be reduced with H-7. ACTH-induced corticosterone secretion was inhibited 50% with H-7 after 24 h, but 8-Br cAMP-induced secretion was unaffected. However, if the treatments were continued for 72 h, H-7 no longer reduced the steroid secretions. Reduction (50-75%) of cholesterol side-chain cleavage enzyme (P450scc) mRNA expression was also noted with H-7 in ACTH-treated cultures after 6 and 24 h. In contrast, TPA doubled the corticosterone secretion induced by 8-Br cAMP, but did not further increase the ACTH-induced secretion after 24 h. TPA alone, however, was not able to induce steroid secretion or P450scc mRNA expression. The morphological differentiation of fetal rat adrenocortical cells with ACTH or 8-Br cAMP from zona glomerulosa-like cells into zona fasciculata-like cells was not disturbed by H-7 nor was it induced by TPA alone. These results therefore suggest that PKC- and cAMP-dependent signal transductions are involved in the ACTH-induced biphasic growth effect of fetal rat adrenocortical cells. PKC plays a role in the inhibitory growth effect, and both PKC and cAMP are involved in the stimulatory growth phase of ACTH. Both PKC and cAMP are air, involved in the steroid secretion of zona glomerulosa-type cells, but differentiation into zona fasciculata-type cells and their steroid production is transduced through cAMP.