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Titolo:
COLORIMETRIC ASSAY FOR RAPID SCREENING OF CORTICOTROPIN-RELEASING FACTOR-RECEPTOR LIGANDS
Autore:
LIAW CW; GRIGORIADIS DE; DESOUZA EB; OLTERSDORF T;
Indirizzi:
NEUROCRINE BIOSCI INC SAN DIEGO CA 92121
Titolo Testata:
Journal of molecular neuroscience
fascicolo: 2, volume: 5, anno: 1994,
pagine: 83 - 92
SICI:
0895-8696(1994)5:2<83:CAFRSO>2.0.ZU;2-M
Fonte:
ISI
Lingua:
ENG
Soggetto:
ADENYLATE-CYCLASE ACTIVITY; FACTOR CRF RECEPTORS; PITUITARY-GLAND; BETA-ENDORPHIN; RAT-BRAIN; SECRETION; HORMONE; STIMULATION; EXPRESSION; ARTHRITIS;
Keywords:
CRF; RECEPTOR; STABLE EXPRESSION; BINDING; CAMP;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Science Citation Index Expanded
Citazioni:
27
Recensione:
Indirizzi per estratti:
Citazione:
C.W. Liaw et al., "COLORIMETRIC ASSAY FOR RAPID SCREENING OF CORTICOTROPIN-RELEASING FACTOR-RECEPTOR LIGANDS", Journal of molecular neuroscience, 5(2), 1994, pp. 83-92

Abstract

The corticotropin releasing factor (CRF) receptor is known to be coupled to G(s) and transduces its signal through stimulation of cyclic AMP (cAMP) production. Here we describe the characterization of several stable CRF receptor-expressing LVIP2.0Zc cell lines that also contain an exogenous cAMP-responsive beta-galactosidase reporter gene construct. The CRF receptor activity was assayed by measuring the induction ofbeta-galactosidase in response to CRF. Rat/human and bovine CRF stimulated beta-galactosidase activity in a dose-dependent manner with EC(50) values of similar to 0.1 nM; the biologically weak deamidated analog of bovine CRF was similar to 500-fold less potent. The CRF receptor antagonist, [d-Phe(12),Nle(21,38),Ala(32)]r/hCRF(12-41) produced a dose-dependent inhibition of CRF-stimulated beta-galactosidase activity, further demonstrating the pharmacological specificity of the interaction. The magnitude of the maximal response to CRF varied among individual cell lines. This variation was independent of the level of CRF receptor expression, but reflected differences in the intrinsic activity of adenylate cyclase. In contrast to most cAMP assay systems, the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine decreased the CRF-induced beta-galactosidase activity when used in the context of the assay regimen described here. Since the assay can be easily performed in a high-throughput 96-well plate format, these cell lines provide an efficient way for the identification of CRF receptor agonists and antagonists.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 27/11/20 alle ore 22:22:57