Functional mutations in mouse norepinephrine transporter reduce sensitivity to cocaine inhibition.
The transporters of dopamine, norepinephrine and serotonin are molecular targets of cocaine, amphetamine, and therapeutic antidepressants. The residues involved in binding these drugs are unknown. We have performed several rounds of random and site-directed mutagenesis in the mouse norepinephrine transporter and screened for mutants with altered sensitivity to cocaine inhibition of substrate uptake. We have identified a triple mutation that retains close to wild-type transport function but displays a 37-fold decrease in cocaine sensitivity and 24-fold decrease in desipramine sensitivity. In contrast, the mutant's sensitivities to amphetamine, methamphetamine, and methylphenidate are only slightly changed. Our data reveal critical residues contributing to the potent uptake inhibitions by these important drugs. Furthermore, this drug-resistant triple mutant can be used to generate a unique knock-in mouse line to study the role of norepinephrine transporter in the addictive effects of cocaine and the therapeutic effects of desipramine.
Wei
Hua
H
Department of Pharmacology, The Ohio State University, 333 West 10th Avenue, 5072 Graves Hall, Columbus, OH 43210, United States.
Hill
Erik R
ER
Gu
Howard H
HH
D000818:Animals; D019921:Biotinylation; D003042:Cocaine / Q000494:pharmacology*; D004298:Dopamine / Q000378:metabolism; D018765:Dopamine Uptake Inhibitors / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D016297:Mutagenesis, Site-Directed / Q000379:methods; D009154:Mutation; D009638:Norepinephrine / Q000378:metabolism; D050484:Norepinephrine Plasma Membrane Transport Proteins / Q000737:chemistry / Q000235:genetics*; D011485:Protein Binding / Q000235:genetics; D014162:Transfection / Q000379:methods; D014316:Tritium / Q000378:metabolism
D018765:Dopamine Uptake Inhibitors; D050484:Norepinephrine Plasma Membrane Transport Proteins; D014316:Tritium; D003042:Cocaine; D004298:Dopamine; D009638:Norepinephrine