In the normal anterior pituitary, intercellular interactions are important for the expression of hormones including adrenocorticotropin (ACTH). Leukemia inhibitory factor (LIF) is secreted by anterior pituitary cells and stimulates both basal and corticotropin-releasing hormone (CRH)-stimulated ACTH secretion in AtT20 cells. To determine the effects of LIF on normal pituitary cells, we measured the effects of LIF and an immunoneutralizing antiserum against LIF on ACTH secretion by cultured sheep anterior pituitary cells. In intact populations of anterior pituitary cells, LIF (10 nM) stimulated ACTH secretion (from 0.30 +/- 0.06 to 0.77 +/- 0.01 ng/well per 3 h) and antiserum to LIF (by itself) had no effect (0.29 +/- 0.05 ng/well per 3 h). In marked contrast, following pharmacological elimination of CRH-target cells, a condition known to disinhibit ACTH secretion, basal ACTH secretion was elevated (0.74 +/- 0.13 ng/well per 3 h); LIF produced no further stimulation (0.73 +/- 0.22 ng/well per 3 h) but immunoneutralization of LIF significantly reduced secretion to 0.50 +/- 0.10 ng/well per 3 h. Medium, conditioned by exposure to CRH-target-depleted cultures of anterior pituitary cells, increased net ACTH secretion (from 0.29 +/- 0.03 to 6.54 +/- 0.71 ng/well per 3 h), when added as a challenge to naive, cultured anterior pituitary cells. Inclusion of antiserum to LIF significantly attenuated (5.29 +/- 0.62 ng/well per 3 h) this response. The presence in and secretion of LIF by normal individual pituitary cells was detected using immunocytochemical methods. Seven to 8% of all cells stained positively for LIF, with 66 +/- 11% of those secreting detectable amounts of LIF under unstimulated conditions. LIF colocalized with TSH and LH in pituitary cells. Taken together, these data suggest that LIF can stimulate ACTH secretion by normal anterior pituitary cells, and potentially plays a role as an intrapituitary stimulator of ACTH secretion under certain conditions.