Comparison of functional profiles at human recombinant somatostatin sst sub(2) receptor: simultaneous determination of intracellular Ca super(2+) and luciferase expression in CHO-K1 cells

• Published in: British journal of pharmacology Vol. 142; no. 1; pp. 150 - 160
• Main Authors: Nunn, C, Cervia, D, Langenegger, D, Tenaillon, L, Bouhelal, R, Hoyer, D
• Format: Journal Article
• Language: English
• Published: 01.05.2004
• ISSN: 0007-1188
• DOI: 10.1038/sj.bjp.0705735

Somatostatin (somatotropin release inhibiting factor; SRIF) acts via five G protein-coupled receptors (sst sub(1)-sst sub(5)) that modulate multiple cellular effectors. The aim of this study was to compare two functional effects of the human sst sub(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. Intracellular calcium was measured using a fluorometric imaging plate reader II (FLIPR II). SRIF-14 rapidly and transiently increased intracellular calcium with a pEC sub(50) of 8.74 plus or minus 0.03 (n = 52). At 5 h after FLIPR II measurements, luciferase expression was determined. SRIF-14 concentration-dependently increased luciferase expression (pEC sub(50) = 9.06 plus or minus 0.03, n = 52). 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 super(2) = 0.83 and 0.90, pEC sub(50) and E sub(max), respectively). Pertussis toxin pretreatment reduced SRIF-14/octreotide-mediated intracellular calcium increases by 45-47% and luciferase expression by 95-98%. Thapsigargin pretreatment abolished the SRIF-14/octreotide-mediated intracellular calcium increase but had no effect on luciferase expression. In conclusion, SRIF stimulates an increase in intracellular calcium and SRE-luciferase expression via human sst sub(2) receptors in CHO-K1 cells. The increase in luciferase is mediated via G sub(i)/G sub(o) while intracellular calcium increase is mediated by both G sub(i)/G sub(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.