Neurochemical Research

, Volume 19, Issue 2, pp 201–208 | Cite as

Studies of the biogenic amine transporters. 1. Dopamine reuptake blockers inhibit [3H]mazindol binding to the dopamine transporter by a competitive mechanism: Preliminary evidence for different binding domains

  • Christina M. Dersch
  • Hyacinth C. Akunne
  • John S. Partilla
  • George U. Char
  • Brian R. de Costa
  • Kenner C. Rice
  • F. Ivy Carroll
  • Richard B. Rothman
Original Articles

Abstract

The present study addressed the hypothesis that the DA transporter ligand, [3H]mazindol, labels multiple sites/states associated with the dopamine (DA) transporter in striatal membranes. Incubations with [3H]mazindol proceeded for 18–24 hr at 4δC in 55.2 mM sodium phosphate buffer, pH 7.4, with a protease inhibitor cocktail. In order to obtain data suitable for quantitative curve fitting, it was necessary to repurify the [3H]mazindol by HPLC before a series of experiments. Under these conditions, we observed greater than 80% specific binding. The method of binding surface analysis was used to characterize the interaction of GBR12935, BTCP, mazindol, and CFT with binding site/sites labeled by [3H]mazindol. A one site model fit the data as well as the two site model: Bmax=16911 fmol/mg protein, Kd of [3H]mazindol=75 nM, Ki of GBR12935 =8.1 nM, Ki of CFT=50 nM and Ki of BTCP=44 nM. The inhibitory mechanism (competitive or noncompetitive) of several drugs (GBR12935, CFT, BTCP, cocaine, cis-flupentixol, nomifensine, WIN35,065-2, bupropion, PCP, and benztropine) was determined. All drugs inhibited [3H]mazindol binding by a competitive mechanism. Although the ligand-selectivity of the [3H]mazindol binding site indicates that it is the uptake inhibitor recognition site of the classic DA transporter, the quantitative differences among the ligand-selectivities of different radioligands for the same site suggest that each radioligand labels different overlapping domains of the DA uptake inhibitor recognition site. It is likely that development of domain-selective drugs may further our under-standing of the DA transporter.

Key Words

Cocaine mazindol dopamine transporter BTCP 

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Copyright information

© Plenum Publishing Corporation 1994

Authors and Affiliations

  • Christina M. Dersch
    • 1
  • Hyacinth C. Akunne
    • 1
  • John S. Partilla
    • 1
  • George U. Char
    • 1
  • Brian R. de Costa
    • 2
  • Kenner C. Rice
    • 2
  • F. Ivy Carroll
    • 3
  • Richard B. Rothman
    • 1
  1. 1.Clinical Psychopharmacology Section, NIDA Addiction Research CenterNIHBaltimore
  2. 2.Laboratory of Medicinal Chemistry, NIDDKNIHBethesda
  3. 3.Research Triangle InstituteResearch Triangle Park

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