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 adrenocortica l cells in primary culture when bromodeoxyuridine (BrdU) incorporation is used as an indicator of proliferation. The immediate inhibitory ef fect during the first 24 h of ACTH stimulation is not dependent on cyc lic AMP (cAMP). Protein kinase C (PKC) inhibitors H-7 and staurosporin e blocked this inhibitory effect of ACTH, whereas 12-0-tetradecanoyl p horbol-13-acetate (TPA; a PKC activator) mimicked the ACTH-induced ant imitogenic effect. The stimulatory growth effect of ACTH appears after 72 h of treatment. A similar mitogenic effect is also achieved with c AMP derivative 8-bromo cAMP (8-Br cAMP). However, both ACTH- and 8-Br cAMP-induced proliferations could be reduced with H-7. ACTH-induced co rticosterone secretion was inhibited 50% with H-7 after 24 h, but 8-Br cAMP-induced secretion was unaffected. However, if the treatments wer e continued for 72 h, H-7 no longer reduced the steroid secretions. Re duction (50-75%) of cholesterol side-chain cleavage enzyme (P450scc) m RNA expression was also noted with H-7 in ACTH-treated cultures after 6 and 24 h. In contrast, TPA doubled the corticosterone secretion indu ced by 8-Br cAMP, but did not further increase the ACTH-induced secret ion after 24 h. TPA alone, however, was not able to induce steroid sec retion or P450scc mRNA expression. The morphological differentiation o f fetal rat adrenocortical cells with ACTH or 8-Br cAMP from zona glom erulosa-like cells into zona fasciculata-like cells was not disturbed by H-7 nor was it induced by TPA alone. These results therefore sugges t that PKC- and cAMP-dependent signal transductions are involved in th e ACTH-induced biphasic growth effect of fetal rat adrenocortical cell s. 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 an d cAMP are air, involved in the steroid secretion of zona glomerulosa- type cells, but differentiation into zona fasciculata-type cells and t heir steroid production is transduced through cAMP.