CHARACTERIZATION OF TYPE-A AND TYPE-B CCK RECEPTOR-BINDING SITES IN RAT VAGUS NERVE

We employed quantitative receptor autoradiography to analyze pharmacological properties of I-125-Bolton Hunter cholecystokinin (CCK-8)-labeled binding sites in sections of rat cervical vagus nerve that had been ligated 24 h prior to extraction. Binding densities were detected in segments of nerve proximal and distal to the ligature. Analysis was confined to proximal segments. Saturation and competitive binding studies were carried out using sulphated CCK-8 and two selective CCK receptor antagonists: MK-329, to define. type-A (CCK(A)) binding sites; and, L-365,260, to define type-B (CCK(B)) binding sites. Sulphated CCK-8 was the most potent inhibitor of vagal I-125-CCK binding (IC50 = 2 nM). Nonlinear curve fitting analysis of the CCK binding data favored the presence of a single class of vagal CCK receptors (KD(i) = 1 nM). However, both MK-329 (IC50 = 18 nM) and L-365,260 (IC50 = 45 nM) competed for vagal I-125-CCK binding indicating the presence of CCK(A) and CCK(B) binding sites. Co-analysis of the antagonist binding data suggested that CCK(A) and CCK(B) receptors were transported in equal concentrations within the vagus. MK-329 bound with high affinity to CCK(A) sites (K(i) = 3 nM) and low-affinity to CCK(B) sites (K(i) = 462 nM) while L-365,260 bound with high affinity to CCK(B) sites (K(i) = 10 nM) and low-affinity to CCK(A) sites (K(i) = 775 nM). These same ligands were used to characterize the specificity of I-125-CCK binding in the medial and lateral divisions of the nucleus of the solitary tract (NTS), two regions innervated by primary vagal afferents carrying CCK receptors. Sulphated CCK-8 was the most potent inhibitor of I-125-CCK binding in both regions of the NTS (IC50 = 0.6 nM, medial; IC50 = 0.4 nM, lateral). In the medial NTS, MK-329 was a potent inhibitor (IC50 = 8.7 nM) while L-365,260 was a weak inhibitor (IC50 = 341 nM) of I-125-CCK binding. In contrast, in the lateral NTS, L-365,260 was a potent inhibitor (IC50 = 21.8 nM) while MK-329 was a weak inhibitor (IC50 > 1,000 nM) of I-125-CCK binding. These results are consistent with the existence of two populations of vagal afferent fibers projecting to different regions of the NTS. One, containing CCK(A) receptors, projects principally to the medial NTS, while the other containing CCK(B) receptors, projects principally to the lateral NTS.