European Journal of Pharmacology, 126 (1986) 211-222 211 Elsevier HIGH-AFFINITY [3HIGBR 12783 BINDING TO A SPECIFIC SITE ASSOCIATED WITH THE NEURONAL DOPAMINE UPTAKE COMPLEX IN THE CENTRAL NERVOUS SYSTEM JEAN-JACQUES BONNET, PHILIPPE PROTAIS, ABDESLAM CHAGRAOUI and JEAN COSTENTIN * Laboratoire de Pharmacodynamie et Physiologie, U.A. C.N.R.S. 1170, U.E.R. de Mbdecine et Pharmacie de Rouen, Avenue de l'Universitb, B.P. 97, 76800 Saint Etienne du Rouvray, France Received 9 January 1986, revised MS received 13 March 1986, accepted 22 April 1986 J.J. BONNET, P. PROTAIS, A. CHAGRAOUI and J. COSTENTIN, High-affinity [~H]GBR 12783 binding to a specific site associated with the neuronal dopamine uptake complex in the central nervous system, European J. Pharmacol. 126 (1986) 211-222. We labelled the neuronal dopamine uptake system by using the potent dopamine uptake inhibitor GBR 12783 in its tritiated form (18.3 Ci/mmol). The binding of [3H]GBR 12783 to rat striatal membranes was saturable and specific with a K d of 1.6 nM and a Brn~xof 10.3 pmol- mg protein -1 as determined by Scatchard analysis. [3H]GBR 12783 binding to rat striatal membranes was inhibited by dopamine uptake inhibitors with IC50 highly correlated with their IC50 for inhibiting [3H]dopamine uptake by a rat striatal synaptosomal preparation. The rank order of potency was the following: GBR 12783 > amfonelic acid > mazindol > pyrovalerone > nomifensine > benztropine > amineptine > methylphenidate > cocaine. Substrates of dopamine uptake competed with [3H]GBR 12783 binding at concentrations higher than those at which they inhibited [3H]dopamine uptake. In rats with a unilateral section of the medial forebrain bundle, the decrease in [3H]GBR 12783 binding to membranes prepared from the ipsilateral striatum was equal to the decrease in [3H]dopamine uptake by a synaptosomal preparation obtained from the same striatum. [3H]GBR 12783 bound in a sodium-dependent manner to membranes prepared from striatum, nucleus accumbens and tuberculum olfactorium. GBR 12783 displayed an approximately 150-fold lower affinity for the cortical norepineph- fine uptake system labelled with [3H]desipramine than for the dopamine transport complex labelled with [3H]GBR 12783. [3H]GBR 12783 appears an attractive tool for the selective characterization of the dopamine uptake system in vitro. In vitro [3H]GBR 12783 binding Dopamine uptake system Catecholamine uptake inhibitors I. Introduction Binding studies have primarily focused on re- ceptors for neurotransmitters. More recently, the labelling of the neuronal uptake systems for several neurotransmitters has awakened vivid interest. Thus, [3H]desipramine and [3H]imipramine have been proposed for the labelling of the norepineph- rine and serotonin uptake systems respectively (Langer et al., 1981; Lee and Snyder, 1981; Re- havi et al., 1982; Sette et al., 1983). Several 3H- ligands for the dopamine (DA) uptake system * To whom all correspondence should be addressed. have also been described: [3H]cocaine (Kennedy and Hanbauer, 1983), [3H]mazindol (Javitch et al., 1984), [3H]nomifensine (Dubocovich and Zahniser, 1985) or [3H]methylphenidate (Janow- sky et al., 1985). All these ligands have a low affinity for the DA transport complex and/or poor selectivity for the DA transport complex compared to the norepinephrine uptake system. Recently, aryl-l,4-dialk(en)yl-piperazine deriva- tives have been described as selective and very potent inhibitors of DA uptake (Van der Zee et al., 1980). We have performed neurochemical studies on GBR 12783, 1-[2-(diphenylmethoxy) ethyl]-4-(3-phenyl-2-propenyl)piperazine (fig. 1), 0014-2999/86/$03.50 © 1986 Elsevier Science Publishers B.V.