title: Binding of the [125I]3 beta-(iodophenyl)tropan-2 beta-carboxylic acid isopropyl ester to the dopamine transporter at a physiologically relevant temperature: mutually exclusive binding and different ionic requirements for various uptake blockers and substrates.
abstract: The present study describes the binding of cocaine analog [125I]3 beta-(iodophenyl)tropan-2 beta-carboxylic acid isopropyl ester ([125I]RTI-121), a highly selective ligand for the dopamine transporter (DAT), to rat striatal synaptosomal membranes at 37 degrees C. Saturation analysis of [125I]RTI-121 binding revealed a single binding site with similar affinity for RTI-121 at both 50 and 134 mM NaCl. However, the density of binding sites was reduced at 134 mM NaCl. Various uptake blockers and substrates of the DAT monophasically inhibited the specific binding of [125I]RTI-121. Increasing the NaCl concentration from 50 mM to 134 mM enhanced the affinity of the substrate dopamine and amphetamine for the DAT, without affecting that of the uptake blockers. At 134 mM NaCl, the copresence of GBR12935, BTCP, cocaine, amphetamine, or dopamine decreased the affinity of RTI-121 to the extent predicted by a model in which the binding of all compounds is mutually exclusive. This, along with a different NaCl sensitivity for blockers and substrates, suggests that the two categories of compounds recognize nonidentical but overlapping binding domains on the DAT. In contrast, the mutually exclusive binding with similar NaCl sensitivity for RTI-121 and the other uptake blockers tested here suggests the involvement of common binding domains in the recognition of these blockers.
authors.affiliation: Department of Pharmacology, Nanjing Medical University School of Basic Medical Sciences, People's Republic of China.
authors.forename: N H
authors.initials: NH
authors.lastname: Chen
authors.forename: Y L
authors.initials: YL
authors.lastname: Wang
authors.forename: M E
authors.initials: ME
authors.lastname: Reith
mesh_terms: D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D002352:Carrier Proteins / Q000187:drug effects*; D003042:Cocaine / Q000031:analogs & derivatives* / Q000494:pharmacology; D050483:Dopamine Plasma Membrane Transport Proteins; D004305:Dose-Response Relationship, Drug; D008297:Male; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; D051381:Rats; D017207:Rats, Sprague-Dawley; D013574:Synaptosomes / Q000187:drug effects*
chemical_list: D002352:Carrier Proteins; D050483:Dopamine Plasma Membrane Transport Proteins; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; C493216:Slc6a3 protein, rat; C079213:RTI 121; D003042:Cocaine