January 1999, Volume 126, Issue 1, Pages 79 - 86

Original Article

Proliferative effects of cholecystokinin in GH3 pituitary cells mediated by CCK2 receptors and potentiated by insulin Alison J. Smith1 & Ruth M. McKernan Merck, Sharp and Dohme Research Laboratories, Neuroscience Research Centre, Terlings Park, Eastwick Road, Harlow, Essex, CM20 2QR     1Author for correspondence

Keywords

CCK;   glycine-extended gastrin;   proliferation;   CCK2 receptor;   insulin;   mitogenic pathways;   L-365,260

Abstract

1   Proliferative effects of CCK peptides have been examined in rat anterior pituitary GH3 cells, which express CCK2 receptors. 2   CCK-8s, gastrin(1-17) and its glycine-extended precursor G(1-17)-Gly, previously reported to cause proliferation via putative novel sites on AR4-2J and Swiss 3T3 cells, elicited significant dose dependent increases of similar magnitude in [3H]thymidine incorporation over 3 days in serum-free medium of 39±10% (P<0.01, n=20), 37±8% (P<0.01, n=27) and 41±6% (P<0.01, n=36) respectively. 3   CCK-8s and gastrin potentially stimulated mitogenesis (EC50 values 0.12 nM and 3.0 nM respectively), whilst G-Gly displayed similar efficacy but markedly lower potency. L-365,260 consistently blocked each peptide. The CCK2 receptor affinity of G-Gly in GH3 cells was 1.09 µM (1.01;1.17, n=6) and 5.53 µM (3.71;5.99, n=4) in guinea-pig cortex. 4   1 µM G-Gly weakly stimulated Ca2+ increase, eliciting a 104±21% increase over basal Ca2+ levels, and was blocked by 1 µM L-365,260 whilst CCK-8s (100 nM) produced a much larger Ca2+ response (331±14%). 5   Insulin dose dependently enhanced proliferative effects of CCK-8s with a maximal leftwards shift of the CCK-8s curve at 100 ng ml-1 (17 nM) (EC50 decreased 500 fold, from 0.1 nM to 0.2 pM; P<0.0001). 10 µg ml-1 insulin was supramaximal reducing the EC50 to 5 pM (P=0.027) whilst 1 ng ml-1 insulin was ineffective. Insulin weakly displaced [125I]BHCCK binding to GH3 CCK2 receptors (IC50 3.6 µM). 6   Results are consistent with mediation of G-Gly effects via CCK2 receptors in GH3 cells and reinforce the role of CCK2 receptors in control of cell growth. Effects of insulin in enhancing CCK proliferative potency may suggest that CCK2 and insulin receptors converge on common intracellular targets and indicates that mitogenic stimuli are influenced by the combination of extracellular factors present.

Received 6 July 1998; Revised 2 October 1998; Accepted 6 October 1998

^© Macmillan Publishers Ltd 1999