The adrenal medulla of several species and some human pheochromocytomas contain CRH. The first aim of the present work was to find out whether normal rat adrenal chromaffin cells and the PC12 rat pheochromocytoma cell line produce CRH in vitro and what regulates its production. CRH was measured and characterized in the media of both types of chromaffin cells under basal conditions and after exposure to K+, nicotine, interleukin-1 beta, and nerve growth factor (NGF). The second aim was to examine the biological effect of exogenous CRH (and of its antagonist) on the production of catecholamines from these two types of cells. Our results are as follows: 1) Both types of chromaffin cells contained and secreted comparable amounts of immunoreactive-CRH under basal conditions and after K(+)-induced depolarization, nicotine, and interleukin-1 beta; 2) the physicochemical characteristics of the immunoreactive-CRH in the cells and the media were identical to the putative CRH peptide on both sieve chromatography and RP-HPLC; 3) synthetic CRH induced the production of catecholamines from both cell types in a dose- and time-dependent manner; this effect was abolished by the antagonist, alpha helical CRH; 4) exposure of PC12 cells to NGF (for 1 week) resulted in their neuronal differentiation and the stimulation of their production of CRH by 30 times and of dopamine by 10 times, compared with parallel controls; this effect of NGF was abolished by alpha helical CRH. In conclusion, our data suggest that the production of CRH by PC12 cells represents the preservation of a normal chromaffin cell characteristic rather than a tumor-induced ectopic phenomenon.