[Study on the radioligand binding assay of cholecystokinin receptor in rat cerebral cortex]

A radioligand binding assay system for determining the characterization of CCK receptor is presented. Using Bolton-Hunter reagent, we prepared a biologically active, specific 125I-BH-CCK8. The iodination mixture was then transferred to a column of Sephadex G-25 and examined by silica thin layer chromatography. Its specific activity and radiochemical purity were 3.4 TBq/mmol and 96% respectively. Binding of 125I-BH-CCK8 to the membrane of rat cerebral cortex was rapid, reversible, time-temperature dependent, saturable and specific. The labeled CCK was shown to have biological activity as measured by the CCK receptor radioassay. Under our laboratorial conditions, the CCK binding required an hour to reach equilibrium at 4 degrees C. We chose polyethylene glycol 6,000 and gamma-globulin protein for the separation of B and F. Scatchard plot of CCK binding was linear with a Kd value of 1.098 nmol/L and Bmax of 197.5 fmol/mg protein. The results of this study support the stand-point that CCK may function as a regulatory peptide in brain and hence may be of use for clarifying the CCK receptor's function in central nervous system.