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Research ArticleArticle

Ion Dependence of Carrier-Mediated Release in Dopamine or Norepinephrine Transporter-Transfected Cells Questions the Hypothesis of Facilitated Exchange Diffusion

Christian Pifl and Ernst A. Singer
Molecular Pharmacology November 1999, 56 (5) 1047-1054; DOI: https://doi.org/10.1124/mol.56.5.1047
Christian Pifl
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Ernst A. Singer
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Abstract

The mechanism of release mediated by the human dopamine and norepinephrine transporter (DAT and NET, respectively) was studied by a superfusion technique in human embryonic kidney 293 cells stably transfected with the respective transporter cDNA and loaded with the metabolically inert substrate [3H]1-methyl-4-phenylpyridinium. Release was induced by amphetamine, dopamine, and norepinephrine or by lowering the sodium or chloride concentration in the superfusion buffer (iso-osmotic replacement by lithium and isethionate, respectively). Efflux of [3H]1-methyl-4-phenylpyridinium was analyzed at 30-s time resolution. In both transporters, release induced by the substrates amphetamine, dopamine, and norepinephrine followed the same time course as release induced by the removal of chloride and was faster than that caused by the removal of sodium. In the presence of low sodium (DAT: 10 mM; NET: 5 mM) none of the substrates was able to induce release from either type of cell, but adding back sodium to control conditions promptly restored the releasing action. In the presence of low chloride (DAT: 3 mM; NET: 2 mM), however, amphetamine as well as the catecholamines stimulated release from both types of cell. In contrast with the ion dependence of release observed in superfusion experiments, uptake initial rates of substrates at concentrations used in release experiments were the same or even higher at low sodium than at low chloride. The results indicate a decisive role of extracellular sodium for carrier-mediated release unrelated to the sodium-dependent uptake of the releasing substrate, and suggest a release mechanism different from simple exchange diffusion considering only the amines as substrates.

Footnotes

    • Received February 8, 1999.
    • Accepted August 16, 1999.
  • Send reprint requests to: Dr. Christian Pifl, Institute of Biochemical Pharmacology, Borschkegasse 8a, University of Vienna, A-1090 Vienna, Austria. E-mail: christian.pifl{at}univie.ac.at

  • The American Society for Pharmacology and Experimental Therapeutics
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Molecular Pharmacology: 56 (5)
Molecular Pharmacology
Vol. 56, Issue 5
1 Nov 1999
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Research ArticleArticle

Ion Dependence of Carrier-Mediated Release in Dopamine or Norepinephrine Transporter-Transfected Cells Questions the Hypothesis of Facilitated Exchange Diffusion

Christian Pifl and Ernst A. Singer
Molecular Pharmacology November 1, 1999, 56 (5) 1047-1054; DOI: https://doi.org/10.1124/mol.56.5.1047

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Research ArticleArticle

Ion Dependence of Carrier-Mediated Release in Dopamine or Norepinephrine Transporter-Transfected Cells Questions the Hypothesis of Facilitated Exchange Diffusion

Christian Pifl and Ernst A. Singer
Molecular Pharmacology November 1, 1999, 56 (5) 1047-1054; DOI: https://doi.org/10.1124/mol.56.5.1047
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