Effects of Purified Hypothalamic Corticotropin-Releasing Factor and Other Substances on the Secretion of Adrenocorticotropin and β-Endorphin-Like Immunoactivities in Vitro*

Abstract

The effects of various substances on the secretion of β-endorphin-like immunoactivities (β- END-LI) or ACTH-like immunoactivities (ACTH-LI) by primary anterior pituitary cell cultures have been studied. Culture medium levels of /J-END-LI and ACTH-LI were estimated by RIAs directed against the LeuH-His27 region of β-endorphin and the N-terminal region of ACTH (ACTH antiserum courtesy of D. Orth). General secretagogues, such as 8Br-AMP, 3-isobutyl methyl xanthine, and elevated medium [K+], stimulate the secretion of both /3-END-LI and ACTH-LI. More specific ACTH secretagogues, such as norepinephrine, vasopressin, and a highly purified preparation of ovine hypothalamic corticotropin-releasing factor (HYPCRF), increase the secretory rates of 8-END-LI as wellas ACTH-LI. The glucocorticoid, dexamethasone-21-PO4, progesterone; and prostaglandin, PGE2, inhibit theCRF-mediated secretion of 0-END-LI and ACTH-LI. Multiple peaks of 0-END-LI and ACTH-LI were resolved by gel filtration of cu ture fluids on Biogel P-60 under denaturing conditions. In media from nonstimulated cells, β-END-LI peaks were observed which coeluted with 31K ACTH, /Mipotropin, and /8-endorphin (β-LPH6i-9i), while peaks of ACTH were seen which coeluted with 31K ACTH and with ACTH. Media from cultured cells stimulated by HYP-CRF or 8Br-AMP had much larger peaks of immunoreactivities coeluting with /Mipotropin, β-endorphin, (ACTH. Fluids from stimulated cells contained an additional peak of ACTH-LI, which eluted as a ca. 6.5 dalton moiety. In summary, purified HYP-CRF and other putative regulatory agents have similar effects on the secretions of /J-END-LI and ACTH-LI in vitro.