Abstract Corticotropin releasing factor (CRF) was tested for its ability to stimulate ACTH and β-endorphin secretion from clonal AtT-20 D16-16 mouse pituitary tumor cells. Release of both hormones was stimulated 4 to 5-fold over the basal release at nanomolar concentrations of synthetic CRF. CRF analogues stimulated ACTH β-endorphin release with the same order of potency in the tumor cells as in primary cultures of anterior pituitary cells. A 90-min exposure to CRF elicited a 29–35% increase in total ACTH and β-endorphin immunoreactivity in tumor cell cultures. Dexamethasone markedly inhibited CRF-stimulated and basal ACTH and β-endorphin release. AtT-20 D16-16 cells may serve as a good model system for studying the biochemistry of CRF receptor-mediated events involved in ACTH β-endorphin release and synthesis.