Comparison of [3H]WIN 35,428 binding, a marker for dopamine transporter, in embryonic mesencephalic neuronal cultures with striatal membranes of adult rats

J Neurochem. 1993 Feb;60(2):469-76. doi: 10.1111/j.1471-4159.1993.tb03174.x.


In contrast to striatal membranes of adult rats, where high- (KD1 = 34 nM) and low- (KD2 = 48,400 nM) affinity binding sites for [3H]WIN 35,428 are present, in primary cultures of ventral mesencephalon neurons (CVMNs) only low-affinity binding sites were found (KD = 336,000 nM). The binding of [3H]WIN 35,428 in CVMNs prepared from rat embryos was reversible, saturable, and located in cytosol. Although dopamine (DA) uptake blockers inhibited [3H]DA uptake at nanomolar concentrations in CVMNs, the displacement of [3H]WIN 35,428 binding in CVMNs by DA uptake inhibitors required 100-8,000 times higher concentrations than were needed to displace [3H]WIN 35,428 binding in striatal membranes. Piperazine derivatives, e.g., GBR-12909, GBR-12935, and rimcazole, inhibited [3H]WIN 35,428 binding in CVMNs more effectively than did cocaine, WIN 35,428, mazindol, nomifensine, or benztropin. A positive correlation (r = 0.779; p < 0.001) was found between drug affinities for the striatal membrane sites labeled by [3H]WIN 35,428 and their abilities to inhibit DA uptake in CVMNs, whereas no correlation existed between the IC50 values of drugs that inhibited [3H]WIN 35,428 binding and [3H]DA uptake in CVMNs. The cytosolic [3H]WIN 35,428 binding sites may be a piperazine acceptor and may not be involved in the regulation of the DA transporter.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Binding, Competitive
  • Carrier Proteins / metabolism*
  • Caudate Nucleus / metabolism*
  • Cell Membrane / metabolism*
  • Cells, Cultured
  • Cocaine / analogs & derivatives*
  • Cocaine / metabolism
  • Corpus Striatum / metabolism*
  • Dopamine / metabolism*
  • Dopamine Plasma Membrane Transport Proteins
  • Embryo, Mammalian
  • Female
  • Kinetics
  • Male
  • Membrane Glycoproteins*
  • Membrane Transport Proteins*
  • Mesencephalon / metabolism*
  • Nerve Tissue Proteins*
  • Neurons / metabolism*
  • Rats
  • Rats, Sprague-Dawley
  • Tritium


  • Carrier Proteins
  • Dopamine Plasma Membrane Transport Proteins
  • Membrane Glycoproteins
  • Membrane Transport Proteins
  • Nerve Tissue Proteins
  • Tritium
  • (1R-(exo,exo))-3-(4-fluorophenyl)-8-methyl-8- azabicyclo(3.2.1)octane-2-carboxylic acid, methyl ester
  • Cocaine
  • Dopamine