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Notes: Summary Dopaminergic binding sites were studied in slices from rat striatum incubated in a physiological medium and using the two highly selective ligands 3H-apomorphine and 3H-domperidone. The clearly biphasic or stretched inhibition of the specific binding of these two ligands by domperidone or apomorphine, respectively allowed to define three distinct classes of binding site. It was demonstrated, by comparing the binding of the 3H-ligand added at the beginning of slice incubation or just before homogenisation of tissue and filtration, that the “specific” bindings only occurred during the incubation of slices. The inhibition constants (K i values) of dopaminergic agents for the three classes of binding site as also the dissociation constants (K d values) of 3H-ligands and the maximal capacity (B max) of the three classes of binding site were closely similar to those of binding sites previously demonstrated on rat striatal membranes, namely D-2, D-3 and D-4 sites (Sokoloff et al. 1980a, b). Their identification on a preparation in which the cellular organisation is largely preserved rules out the possibility that these sites represent an artifact due to membrane preparation. Unexpectedly the addition of guanyl nucleotides like GTP or GppNHp to the slice preparation decreased the binding of 3H-apomorphine to the high affinity sites (particularly to the D-2 sites) while D-4 site binding was correspondingly increased. The guanylnucleotide effect apparently took place before cell disruption and occurred at concentrations similar to those required in striatal membrane preparations. These observations, together with those indicating the presence of high affinity binding sites for dopaminergic agonists in intact striatal cells, suggest that a putative nucleotide regulatory unit of dopamine receptors, is not fully occupied by intracellular GTP but could be interacted with from the external face of the cell membrane.

Notes: Summary In order to document the hypothesis that anti-psychotics may interact with more than one class of cerebral dopamine receptors, the relative potencies of a series of compounds were compared in three behavioral tests and in binding studies with two radioligands. Apomorphine (0.6 mg/kg) simultaneously clicited in rat two kinds of facial stereotypies (sniffing and licking) and a stereotyped climbing behavior, allowing to compare in the same animals the relative potencies of various antipsychotics against the three behaviors. Only some substituted benzamides (Sulpiride, LUR 2366 and DAN 2163) antagonised at significantly lower dosages climbing than sniffing (or licking). The possibility that this discriminant potency might be related to a distinct affinity for two classes of dopamine receptors was investigated by binding studies on striatal membranes with 3H-apomorphine and 3H-domperidone. From lesion and subcellular fractionation studies, two classes of binding sites both labeled with 3H-domperidone but distinguished by apomorphine i.e. D-2 sites with nM affinity and D-4 sites with μM affinity for the dopamine agonist (according to the nomenclature of Sokoloff et al. 1980b) appear to be differently localised in striatum. Thus D-2 sites, whose number decreases after kainate lesion, are not significantly modified following cortical lesions and preferentially sediment with heavy primary subcellular fractions. In contrast D-4 sites, less affected by kainate lesions, are significantly decreased following cortical lesions (−30%) and preferentially sediment with the light subcellular fractions. In addition the apparently heterogenous recognition of total 3H-domperidone binding sites (i.e. the sum of D-2 and D-4 sites) by dopamine and apomorphine persists in the presence of guanosine-5′-triphosphate (pseudo-Hill coefficient of 0.60 instead of 0.55). This suggests that D-2 and D-4 sites cannot be considered as two discrete states of the same receptor strictly convertible one into the other by guanylnucleotides. Whereas most dopamine antagonists inhibited D-2 and D-4 site binding with similar affinities, the three benzamide derivatives with the largest selectivity in behavioral tests displayed 2–3-fold higher affinity for D-4 than for D-2 sites and the ratios of ID50 values of the whole series of antagonists against sniffing (or licking) and climbing behaviors were correlated (P〈0.01) with the ratios of K i values regarding D-2 and D-4 site binding. Also, sulpiride and LUR 2366 unlike haloperidol and metoclopramide, inhibited the total specific 3H-domperidone binding in a biphasic manner. However, the distinction by sulpiride and LUR 2366 of low and high affinity sites did not superimpose that of D-2 and D-4 sites, as distinguished by agonists. In this test the relative proportion of low affinity sites was 2-fold higher than that of D-2 sites and K i values for high affinity sites were lower than that for D-4 sites. Also the heterogeneity of 3H-domperidone sites regarding affinity of LUR 2366 persisted in the presence of low concentrations of apomorphine. Hence low affinity sites for discriminant benzamide derivatives may exist in two forms, distinguished by agonists and possibly interconvertible by GTP. Thus the hypothesis that two classes of central dopamine receptor can be distinguished by some substituted benzamides, but perhaps display no great difference in affinity of agonists in their physiological state, fits partic-ularly well with behavioral data.