Identification of CCK-A receptors on chief cells with use of a novel, highly selective ligand

Abstract

Functional studies suggest that guinea pig chief cells have both cholecystokinin-A (CCK-A) and CCK-B receptors (CCK-A-R and CCK-B-R, respectively). However, all efforts to directly characterize the specific CCK-A-R using binding have been unsuccessful. Recent studies describe specific CCK-A-R agonists such as A-71378 ([desamino-Nle28,31-(N-methyl)Asp32]CCK heptapeptide]. In the present study, [D-Tyr-Gly]A-71378 was synthesized, which has > 300-fold selectivity for CCK-A-R and can be iodinated. [D-Tyr-Gly]A-71378 was equipotent to A-71378 in stimulating pepsinogen release from purified guinea pig chief cells. Binding of 125I-labeled [D-Tyr-Gly]A-71378 was saturable and specific. Potencies for inhibiting binding were as follows: [D-Tyr-Gly]A-71378 = A-71378 = 4x CCK octapeptide (CCK-8) > 1,000x des(SO4)-CCK-8, gastrin. In contrast, for 125I-gastrin binding they were CCK-8 > gastrin-17-I > des(SO4)-CCK-8 >> A-71378 or [D-Tyr-Gly]A-71378. Binding of [D-Tyr-Gly]A-71378 was best fitted by a two-site model. In contrast, 125I-gastrin binding was fitted with a single-site model. For inhibiting binding of 125I-[D-Tyr-Gly]A-71378, the CCK antagonists had relative affinities of L-364,718 >> L-365,260, and the reverse was true with 125I-gastrin. Correlation of binding with changes in biological activity suggested low-affinity CCK-A-R were mediating these changes. These results demonstrate directly for the first time that guinea pig chief cells possess CCK-A-R and CCK-B-R. The pharmacology of these CCK-A-R resembles those on other tissues. This novel, highly selective CCK-A ligand should be useful because it will identify CCK-A-R when they make up as little as 0.2% of the total CCK receptor number.