CHARACTERIZATION AND DISTRIBUTION OF BINDING-SITES FOR A NEW NEUROTENSIN RECEPTOR ANTAGONIST LIGAND, [H-3] SR-48692, IN THE GUINEA-PIG BRAIN

SR 48692, a selective nonpeptide antagonist of neurotensin (NT) receptors was developed recently. In the present work we studied the binding properties of the corresponding radioligand, [H-3]SR 48692, in the adult guinea pig brain. The characterization of [H-3]SR 48692 binding was carried out on brain membrane preparations and the distribution of [H-3]SR 48692 binding sites was determined by receptor au toradiography and compared to that of [I-125]NT binding sites. In brain homogenates, [3H]SR 48692 bound to a single population of sites with a K-d of 2.19 nM and a maximal binding capacity of 1.15 pmol/mg of protein. This maximal binding capacity value was 20 times higher than that observed for [I-125]NT. NT agonists were able to interact competitively with the entire population of binding sites labeled by [H-3]SR 48692, but their affinities were much lower than those observed for [I-125]NT. By contrast, NT antagonists exhibited similar abilities to inhibit the binding of both radioligands. The addition of unlabeled NT in saturation assays revealed a competitive inhibition of [H-3]SR 48692 binding, suggesting that agonist and antagonist ligand bind to overlapping domains of the NT receptor, The autoradiographic distribution of the low-affinity NT binding sites detected by [H-3]SR 48692 (96% of the receptors) was very similar to the distribution of high-affinity receptors labeled with [I-125]NT (4% of the receptors). In addition, the binding of [H-3]SR 48692 was insensitive to guanyl nucleotides. Taken together, these findings suggest that the binding sites detected by [H-3]SR 48692 in the guinea pig brain mainly represent the uncoupled form of the NT receptor.