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Comparison of functional profiles at human recombinant somatostatin sst2 receptor: simultaneous determination of intracellular Ca2+ and luciferase expression in CHO-K1 cells.

Nunn, Caroline and Cervia, Davide and Langenegger, Daniel and Tenaillon, Laurent and Bouhelal, Rochdi and Hoyer, Daniel (2004) Comparison of functional profiles at human recombinant somatostatin sst2 receptor: simultaneous determination of intracellular Ca2+ and luciferase expression in CHO-K1 cells. British Journal of Pharmacology, 142 (1). pp. 150-160. ISSN 0007-1188

Abstract

1. Somatostatin (somatotropin release inhibiting factor; SRIF) acts via five G protein-coupled receptors (sst(1)-sst(5)) that modulate multiple cellular effectors. The aim of this study was to compare two functional effects of the human sst(2) receptor stably expressed in CHO-K1 cells in a single experiment using a duplex assay for intracellular calcium and serum response element (SRE)-driven luciferase expression. 2. Intracellular calcium was measured using a fluorometric imaging plate reader II (FLIPR II). SRIF-14 rapidly and transiently increased intracellular calcium with a pEC(50) of 8.74+/-0.03 (n=52). At 5 h after FLIPR II measurements, luciferase expression was determined. SRIF-14 concentration-dependently increased luciferase expression (pEC(50)=9.06+/-0.03, n=52). 3. Natural and synthetic agonist/antagonist ligands for SRIF receptors were tested in the duplex assay. Correlation of agonist potencies and efficacies between the two responses were significant (r(2)=0.83 and 0.90, pEC(50) and E(max), respectively). 4. Pertussis toxin pretreatment reduced SRIF-14/octreotide-mediated intracellular calcium increases by 45-47% and luciferase expression by 95-98%. 5. Thapsigargin pretreatment abolished the SRIF-14/octreotide-mediated intracellular calcium increase but had no effect on luciferase expression. 6. In conclusion, SRIF stimulates an increase in intracellular calcium and SRE-luciferase expression via human sst(2) receptors in CHO-K1 cells. The increase in luciferase is mediated via G(i)/G(o) while intracellular calcium increase is mediated by both G(i)/G(o) proteins and pertussis toxin-insensitive G proteins, and is mainly via release of calcium from intracellular stores. SRIF ligands display a similar recognition profile suggesting that the ligand/receptor/G protein/effector interaction is similar for the two parameters.

Item Type: Article
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Additional Information: free final full text version available at PubMedCentral; archiving not allowed on institutional repository
Keywords: Somatostatin receptor subtype 2; CHO-K1 cells; FLIPR II; SRE-luciferase; duplex assay; pertussis toxin; thapsigargin
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Date Deposited: 14 Dec 2009 13:54
Last Modified: 31 Jan 2013 01:07
URI: https://oak.novartis.com/id/eprint/769

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