Evidence for a displaceable non-specific [3H]neurotensin binding site in rat brain

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Abstract
Levocabastine is a potent antihistamine drug, structurally unrelated to neurotensin. In rat and mouse brain but not in other animal species, it inhibited 60% of the [3H]neurotensin binding displaced by unlabelled neurotensin or neurotensin (8–13). The levocabastine-sensitive site or “site 1” displayed high affinity properties for levocabastine (IC50=25 nM) and was highly stereospecific (IC50-value higher than 10 μM for one of the isomers). Binding to the “site 1” in rat brain corresponded to the [3H]neurotensin binding displaceable by 1 μM levocabastine, whereas binding to the “site 2” corresponded to the binding displaced by 1 μM neurotensin when the “site 1” was occluded by 1 μM levocabastine. Both “site 1” and “site 2” appeared to be saturable. Scatchard plots obtained in rat bulbus olfactorius allowed to calculate a K D-values of 7.1 nM and a B max-values of 37.2 fmol/mg original tissue for “site 1”, while “site 2” displayed a K D-value of 0.7 nM and a B max-value of 16.3 fmol/mg original tissue. The regional distributions of both sites showed marked differences. The “site 1” was homogeneously distributed throughout all rat brain areas, whereas the amount of “site 2” binding was markedly different in separate brain areas: bulbus olfactorius and substantia nigra had the highest amounts (8.9 and 7.8 fmol/mg tissue) while cerebellum had the lowest (0.4 fmol/mg tissue). In spite of its high affinity and stereospecificity, “site 1” has to be considered as an acceptor or recognition site for [3H]neurotensin because of its species-link, low saturability and homogeneous distribution in all rat brain areas. On the other hand, “site 2” had the characteristics of a physiological receptor: high affinity, saturability in the low nanomolar range and marked regional distribution in rat brain. “Site 2” corresponds therefore most probably to the physiological neurotensin receptor. The foregoing experiments provide evidence for the presence of a drug displaceable, non-specific (=unrelated to a physiological receptor) neurotensin binding site in rat brain; levocabastine should be an important tool to occlude this site in order to reveal, by means of in vitro binding assays, the specific neurotensin binding site in rat brain.