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Agonist and antagonist modulation of [35S]-GTP gamma S binding in transfected CHO cells expressing the neurotensin receptor

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Hermans, Emmanuel [UCL] Geurts, Muriel [UCL] Maloteaux, Jean-Marie [UCL]

1. The functional interaction of the cloned rat neurotensin receptor with intracellular G-proteins was investigated by studying the binding of the radiolabelled guanylyl nucleotide analogue [35S]-GTP gamma S induced by neurotensin to membranes prepared from transfected Chinese hamster ovary (CHO) cells. 2. The agonist-induced binding of [35S]-GTP gamma S was only detected in the presence of NaCl in the incubation buffer. However, it was also demonstrated that the binding of [3H]-neurotensin to its receptor was inhibited by NaCl. In the presence of 50 mM NaCl, the binding of the labelled nucleotide was about 2 fold increased by stimulation with saturating concentrations of neurotensin (EC50 value of 2.3 +/- 0.9 nM). 3. The stimulation of [35S]-GTP gamma S binding by neurotensin was mimicked by the stable analogue of neurotensin, JMV-449 (EC50 value of 1.7 +/- 0.4 nM) and the neurotensin related peptide neuromedin N (EC50 value of 21 +/- 6 nM). 4. The NT-induced [35S]-GTP gamma S binding was competitively inhibited by SR48692 (pA2 value of 9.55 +/- 0.28), a non-peptide neurotensin receptor antagonist. SR48692 alone had no effect on the specific binding of [35S]-GTP gamma S. 5. The response to neurotensin was found to be inhibited by the aminosteroid U-73122, a putative inhibitor of phospholipase C-dependent processes, indicating that this drug may act at the G-protein level. 6. Taken together, these results constitute the first characterization of the exchange of guanylyl nucleotides at the G-protein level that is induced by the neuropeptide neurotensin after binding to its receptor.

Document type Article de périodique (Journal article) – Article de recherche
Access type Accès libre
Publication date 1997
Journal information "British Journal of Pharmacology" - Vol. 121, no. 8, p. 1817-1823 (1997)
Peer reviewed yes
Publisher JohnWiley & Sons Ltd. (Chichester)
issn 0007-1188
e-issn 1476-5381
Publication status Publié
Affiliations UCL - MD/FSIO - Département de physiologie et pharmacologie
UCL - (SLuc) Service de neurologie
MESH Subject Animals ; CHO Cells ; Cricetinae ; Estrenes - pharmacology ; GTP-Binding Proteins - physiology ; Guanosine 5'-O-(3-Thiotriphosphate) - metabolism ; Guanosine Diphosphate - secretion ; Neurotensin - metabolism ; Pyrrolidinones - pharmacology ; Rats ; Receptors, Neurotensin - agonists - antagonists & inhibitors - metabolism ; Sodium Chloride - pharmacology ; Transfection
Keywords Neurotensin ; SR 48692 ; U-73122 ; [35S]-GTPgammaS ; G-protein
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Bibliographic reference Hermans, Emmanuel ; Geurts, Muriel ; Maloteaux, Jean-Marie. Agonist and antagonist modulation of [35S]-GTP gamma S binding in transfected CHO cells expressing the neurotensin receptor. In: British Journal of Pharmacology, Vol. 121, no. 8, p. 1817-1823 (1997)
Permanent URL http://hdl.handle.net/2078.1/12360