Corticotropin-releasing factor (CRF) stimulates 45Ca2+ uptake in the mouse corticotroph cell line AtT-20.
Corticotropin-releasing factor (CRF) stimulates adrenocorticotropin (ACTH) release via the adenylate cyclase/cAMP-dependent protein kinase system. Because calcium is necessary for receptor-mediated release of ACTH, we have examined the effect of CRF on 45Ca2+ uptake in a corticotroph cell line model, AtT-20. Treatment of AtT-20 cells with CRF (10(-9)-10(-6) M) resulted in dose- and time-dependent increases in 45Ca2+ uptake, up to 2.2-fold above control values. The effect was statistically significant at 1 min and persisted for at least 10 min. Treatment with forskolin (1-30 microM), 8-Br-cAMP (0.5 mM), cholera toxin (CT, 100 ng/ml) and K+ (20 mM) also increased cell-associated 45Ca2+. The effect of K+ was completely blocked by nifedipine (100 microM), whereas the effects of CRF (10(-8) M) were only partially inhibited by this calcium channel antagonist. These data suggested a role of voltage-dependent calcium channels in 45Ca2+ uptake. Short term pretreatment (1-2 h) of AtT-20 cells with CRF (10(-8) M) significantly desensitized both CRF-stimulated cAMP accumulation and ACTH release, but did not attenuate CRF-stimulated 45Ca2+ uptake. Pretreatment with CRF (10(-8) M) for 4 h did not alter CT- or forskolin-stimulated cAMP accumulation and ACTH release. This suggests that the molecular mechanisms of desensitization are proximal to adenylate cyclase. Conversely, long term pretreatment (24 h) of AtT-20 cells with CRF (10(-8) M) induced significant desensitization of CRF-stimulated 45Ca2+ uptake. These results indicate that CRF stimulates calcium uptake in AtT-20 cells via cAMP-dependent and cAMP-independent mechanisms, and that the cellular mechanisms involved in desensitization of cAMP accumulation and ACTH release and those involved in desensitization of calcium uptake are qualitatively different.
Endocrine Unit, Massachusetts General Hospital, Harvard Medical School, Boston 02114.
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Abou-Samra
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000324:Adrenocorticotropic Hormone / Q000378:metabolism; D000704:Analysis of Variance; D000818:Animals; D002132:Calcium Radioisotopes / Q000378:metabolism*; D002772:Cholera Toxin / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D003346:Corticotropin-Releasing Hormone / Q000502:physiology*; D000242:Cyclic AMP / Q000378:metabolism; D051379:Mice; D009543:Nifedipine / Q000494:pharmacology; D010903:Pituitary Gland, Anterior / Q000166:cytology / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D002132:Calcium Radioisotopes; D005576:Colforsin; D015124:8-Bromo Cyclic Adenosine Monophosphate; D000324:Adrenocorticotropic Hormone; D002772:Cholera Toxin; D003346:Corticotropin-Releasing Hormone; D000242:Cyclic AMP; D009543:Nifedipine