The chromaffin cells of the adult rat adrenal medulla are essentially growth arrested in situ, but can proliferate in vitro, suggesting the existence of growth inhibitory factors in the adrenal gland. We have investigated whether pituitary adenylate cyclase-activating polypeptide 38 (PACAP38) could be involved in the growth arrest of adrenal chromaffin cells. In adult rat adrenal gland, PACAP38 was detected by radioimmunoassay and high-performance liquid chromatography and its concentration in the medulla was estimated as 24 nmol/kg wet tissue. Immunohistochemistry of the neonatal and adult rat adrenal medulla showed PACAP38 immunoreactivity in a widely distributed network of delicate nerve fibers surrounding the chromaffin cells. In a primary culture system, PACAP38 inhibited growth factor-stimulated DNA synthesis by 90% in neonatal and adult rat chromaffin cells with half-maximal inhibition at 4 and 0.5 nM, respectively, as demonstrated by bromodeoxyuridine pulse-labeling and immunocytochemical staining of cell nuclei. In comparison, corticosterone inhibited neonatal and adult chromaffin cell proliferation by 70% and 95%, respectively, with half-maximal effect at 100 nM. In neonatal chromaffin cells, 100 nM PACAP38 and 1 microM corticosterone added together abolished proliferation completely (99.8% inhibition). Finally, PACAP38 increased cell survival but showed little neurite-promoting activity in the chromaffin cells. Our data suggest that neurally derived PACAP38, in conjunction with glucocorticoids, may override growth factor mitogenic signals, leading to the postmitotic state of chromaffin cells in the adult adrenal medulla.