PT  - JOURNAL ARTICLE
AU  - D. KIRK WAYS
AU  - DAVID A. ONTJES
TI  - Reversal of Persistently Stimulated Steroidogenesis by GTP and an Inhibitory Adrenocorticotropin Analogue in Adrenal Cells Pretreated with Adrenocorticotropin
DP  - 1979 Mar 01
TA  - Molecular Pharmacology
PG  - 271--286
VI  - 15
IP  - 2
4099  - http://molpharm.aspetjournals.org/content/15/2/271.short
4100  - http://molpharm.aspetjournals.org/content/15/2/271.full
SO  - Mol Pharmacol1979 Mar 01; 15
AB  - A single addition of ACTH1-24 at concentrations of 0.1 to 10 nM to an in vitro suspension of isolated rat adrenal cells elicits a dose related steroidogenic response lasting from 150 to 240 min. The addition of fresh ACTH can initiate a new response after the spontaneous termination of the first, but persistent steroidogenesis does not require the continued presence of active hormone in the incubation medium. Biologically active hormone disappears from the extracellular medium within the first 90 min of incubation, while steroidogenesis can continue at an undiminished rate for an additional 120 min or more. Cells treated for 1 min with 100 nM ACTH, then rinsed and resuspended in hormone-free medium, produce as much cyclic AMP and corticosterone as do cells continually exposed to the same concentration of hormone. The persistent steroidogenesis seen in the ACTH-pretreated cells may be rapidly terminated by exposure of the cells to GTP or to the ACTH antagonist, ACTH6-39. These agents can also greatly accelerate the slow dissociation of bound 125I-ACTH1-24 from the cells. These findings suggest that the persistence of steroidogenesis depends upon the presence of persistently bound biologically active hormone on the exterior surface of the adrenocortical cell. The rapid displacement of bound hormone by the competitive antagonist, ACTH6-39, supports the hypothesis that there is negative cooperativity among ACTH receptors. The mechanism by which GTP increases the dissociation of bound hormone is unclear. Under the experimental conditions used, the addition of GTP to the medium does not appear to stimulate the production of cyclic AMP. GTP enhances the dissociation of the radiolabeled antagonist, 125I-ACTH6-39, as well as the dissociation of labeled agonist hormone. Thus it appears that GTP increases the dissociation of bound ACTH independently of adenylate cyclase activity, probably by interacting with sites on the cell surface and altering the ability of the receptors to bind and retain hormone. ACKNOWLEDGMENTS We are grateful to Dr. Alton L. Steiner for his gift of cyclic AMP antisera and radiolabeled cyclic AMP. We would also like to thank Dr. John Perkins for his helpful suggestions concerning this manuscript and Ms. Celeste Layton for her assistance in the preparation of the manuscript.