Induction by low Na+ or Cl- of cocaine sensitive carrier-mediated efflux of amines from cells transfected with the cloned human catecholamine transporters.
1. COS-7 cells transfected with the cDNA of the human dopamine transporter (DAT cells) or the human noradrenaline transporter (NAT cells) were loaded with [3H]-dopamine or [3H]-noradrenaline and superfused with buffers of different ionic composition. 2. In DAT cells lowering the Na+ concentration to 0, 5 or 10 mM caused an increase in 3H-efflux. Cocaine (10 microM) or mazindol (0.3 microM) blocked the efflux at low Na+, but not at 0 Na+. Lowering the Cl- concentration to 0, 5 or 10 mM resulted in an increased efflux, which was blocked by cocaine or mazindol. Desipramine (0.1 microM) was without effect in all the conditions tested. 3. In NAT cells, lowering the Na+ concentration to 0, 5 or 10 mM caused an increase in 3H-efflux, which was blocked by cocaine or mazindol. Desipramine produced a partial block, its action being stronger at 5 or 10 mM Na+ than at 0 mM Na+. Efflux induced by 0, 5 or 10 mM Cl- was completely blocked by all three uptake inhibitors. 4. In cross-loading experiments, 5 mM Na(+)- or 0 Cl(-)-induced efflux was much lower from [3H]-noradrenaline-loaded DAT, than NAT cells and was sensitive to mazindol, but not to desipramine. Efflux from [3H]-dopamine-loaded NAT cells elicited by 5 mM Na+ or 0 Cl- was blocked by mazindol, as well as by desipramine. 5. Thus cloned catecholamine transporters display carrier-mediated efflux of amines if challenged by lowering the extracellular Na+ or Cl-, whilst retaining their pharmacological profile. The transporters differ with regard to the ion dependence of the blockade of reverse transport by uptake inhibitors.
Pifl
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C
Institute of Biochemical Pharmacology, University of Vienna, Austria.
Agneter
E
E
Drobny
H
H
Reither
H
H
Singer
E A
EA
D000588:Amines / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport / Q000502:physiology; D002352:Carrier Proteins / Q000378:metabolism*; D002395:Catecholamines / Q000378:metabolism; D002478:Cells, Cultured; D002712:Chlorides / Q000494:pharmacology*; D003001:Cloning, Molecular; D003042:Cocaine / Q000494:pharmacology*; D050483:Dopamine Plasma Membrane Transport Proteins; D006801:Humans; D007668:Kidney / Q000378:metabolism*; D008251:Macaca; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; D012964:Sodium / Q000494:pharmacology*; D014162:Transfection
D000588:Amines; D002352:Carrier Proteins; D002395:Catecholamines; D002712:Chlorides; D050483:Dopamine Plasma Membrane Transport Proteins; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; C493214:SLC6A3 protein, human; D012964:Sodium; D003042:Cocaine