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Adrenocorticotropin-Releasing Factor Down-Regulates Glucocorticoid Receptor Expression in Mouse Corticotrope Tumor Cells via an Adenylate Cyclase-Dependent Mechanism^*

Fishberg Research Center in Neurobiology, Mt. Sinai Medical Center New York, New York 10029

Address all correspondence and requests for reprints to: Karen E. Sheppard, Baker Medical Research Institute, P.O. Box 348, Prahran, Victoria, Australia, 3181.

Abstract

Anterior pituitary POMC transcription and peptide release are negatively regulated by glucocorticoids and stimulated by CRF. Although pretreatment of corticotrope cells with CRF markedly inhibits subsequent glucocorticoid effects, the mechanism of this action is unclear. We have thus used a mouse corticotrope tumor (AtT20) cell line, to examine the effects of CRF on glucocorticoid receptor (GR) messenger RNA levels and on GR capacity/nuclear translocation. GR mRNA levels were measured by solution hybridization/S1 nuclease protection, and both total cell binding and nuclear binding were determined with [³H]dexamethasone ([³H]DEX). CRF treatment of AtT20 cells led to a rapid time-dependent decrease in GR mRNA levels which preceded a dose- and time-dependent decrease in GR binding capacity. Scatchard analysis showed a single class of high affinity binding sites (GR) in both control and CRF-treated cultures, and a decrease in the total number of GR after CRF treatment. The relative proportion of nuclear vs. cytoplasmic localized [³H]DEX-bound GR did not differ between control and CRF-treated cultures, indicating that CRF does not interfere with GR nuclear translocation. To investigate whether CRF regulates GR expression through the adenylate cyclase system, as it does POMC, AtT20 cells were treated with either forskolin or 8-bromo-cAMP, and specific nuclear GR binding was determined. Both drugs mimic the CRF-induced decrease in GR binding, and in addition forskolin decreased GR mRNA levels; in contrast, forskolin had no effect on GH₃ cell GR levels. These results suggest that CRF can decrease the cellular concentration of GR, and thus potentially the response to glucocorticoids, through the same mechanism by which it stimulates anterior pituitary POMC expression.

Footnotes

^* This work was supported in part by a Fulbright Postdoctoral Fellowship (to K.E.S.) and NIH Grant DK27484 (to J.L.R. and M.B.).

Received February 19, 1991.

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