Characterization of cocaine-sensitive dopamine uptake in PC12 cells.
Dopamine uptake in rat pheochromocytoma (PC12) cells is a carrier-mediated process which follows Michaelis-Menten kinetics. Uptake was saturable with an apparent Km of 0.71 microM for dopamine and a Vmax of 3.2 pmol/2 x 10(5) cells/min. The rank order of potency for various amines was norepinephrine > or = dopamine > epinephrine. Uptake increased with increasing temperature and showed a sharp break in the Arrhenius plot at 27.5 degrees C. The Q10 was 1.39 above and 2.95 below 27.5 degrees C. Cocaine inhibited uptake in a dose-dependent manner with a Ki of 0.97 microM. The presence of cocaine lowered the apparent Km but did not affect the Vmax, indicating competitive inhibition. Tunicamycin inhibited [3H]dopamine accumulation in a dose- and time-dependent fashion suggesting the dopamine uptake site in PC12 cells is an asparagine-linked glycoprotein. Kinetic analysis showed a decrease in Vmax but not in the apparent Km after tunicamycin treatment, consistent with the notion that tunicamycin treatment results in the loss of a substantial amount of active carrier molecules.
Zhu
J
J
Department of Pharmacology, University of Nebraska Medical Center, Omaha 68198-6260.
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D000818:Animals; D001667:Binding, Competitive; D003042:Cocaine / Q000494:pharmacology*; D004298:Dopamine / Q000378:metabolism*; D004837:Epinephrine / Q000378:metabolism; D006358:Hot Temperature; D007700:Kinetics; D016716:PC12 Cells / Q000378:metabolism*; D051381:Rats; D013816:Thermodynamics; D014415:Tunicamycin / Q000494:pharmacology
D014415:Tunicamycin; D003042:Cocaine; D004298:Dopamine; D004837:Epinephrine