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 muM for dopamine and a V(max) of 3.2 pmol/2 x 10(5) cells/min. The rank order of potency for various amines was norepinephrine greater-than-or-equal-to 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 K(i) of 0.97 muM. The presence of cocaine lowered the apparent K(m) but did not affect the V(max), indicating competitive inhibition. Tunicamycin inhibited [H-3]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 V(max) but not in the apparent K(m) after tunicamycin treatment, consistent with the notion that tunicamycin treatment results in the loss of a substantial amount of active carrier molecules.