Differences in interactions with the dopamine transporter as revealed by diminishment of Na(+) gradient and membrane potential: dopamine versus other substrates.
In heterologous cells expressing the dopamine transporter (DAT), simultaneous elevation of intracellular Na(+) and depolarization of the membrane with gramicidin reduced the potency of various DAT substrates, including dopamine, d-amphetamine, beta-phenethylamine, p-tyramine, and MPP(+), in inhibiting binding of the cocaine analog [(3)H]CFT, with the greatest reduction observed for d-amphetamine. In rat striatal synaptosomes, gramicidin exerted similar effects; in addition, the potency of d-amphetamine was reduced by the Na(+)-channel activator veratridine. The latter effect was counteracted by the Na(+)-channel blocker tetrodotoxin. In broken membranes, where, as the situation with gramicidin, both sides of the non-polarized membrane were exposed to 130 mM Na(+), gramicidin was ineffective. Dopamine had a potency for membrane preparations that was not significantly different from that for control cells or synaptosomes, while other substrates had potencies for membrane preparations that were reduced to a level similar to those observed in gramicidin-treated cells or synaptosomes. These results suggest that diminishing Na(+) gradient and membrane potential may convert DAT to a conformational state that dopamine could easily bind to when gaining free access to its intracellular portion. In contrast, non-dopamine substrates may not be able to readily interact with this state from either side of the membrane.
Zhen
Juan
J
Department of Biological Sciences, Illinois State University, Normal, IL, USA.
Chen
Nianhang
N
Reith
Maarten E A
ME
D000704:Analysis of Variance; D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D002118:Calcium / Q000494:pharmacology; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D003042:Cocaine / Q000031:analogs & derivatives / Q000493:pharmacokinetics; D003342:Corpus Striatum / Q000166:cytology; D004298:Dopamine / Q000378:metabolism*; D050483:Dopamine Plasma Membrane Transport Proteins / Q000378:metabolism*; D018765:Dopamine Uptake Inhibitors / Q000493:pharmacokinetics / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D006096:Gramicidin / Q000494:pharmacology; D006801:Humans; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology*; D011485:Protein Binding / Q000187:drug effects / Q000502:physiology; D011869:Radioligand Assay / Q000379:methods; D051381:Rats; D017207:Rats, Sprague-Dawley; D012964:Sodium / Q000378:metabolism*; D013574:Synaptosomes / Q000187:drug effects / Q000378:metabolism; D014316:Tritium / Q000493:pharmacokinetics
D050483:Dopamine Plasma Membrane Transport Proteins; D018765:Dopamine Uptake Inhibitors; D014316:Tritium; D006096:Gramicidin; C021279:(1R-(exo,exo))-3-(4-fluorophenyl)-8-methyl-8- azabicyclo(3.2.1)octane-2-carboxylic acid, methyl ester; D012964:Sodium; D003042:Cocaine; D002118:Calcium; D004298:Dopamine