FIGURE 3.
Effect of MPP+ on DAcyt homeostasis. A, oxidation profile of MPP+ shows that the presence of the toxin did not interfere with cyclic voltammetry measurements of DA. B, MPP+ (10 μm, 1 h) induced a large increase in DAcyt, which was prevented by the DAT inhibitor nomifensine (Nmf; 5 μm, 15-min pretreatment). *, p < 0.01 compared with all other groups by one-way ANOVA. Ctrl, control. C, time course of changes in DAcyt following cell treatment with 10 or 50 μm MPP+. The curves are statistically different by two-way ANOVA (p < 0.01). D, DAcyt in neurons exposed to 10 μm MPP+ for 1 h in the presence and absence of the VMAT2 inhibitor reserpine (Res; 2 μm, 60-min pretreatment) or the MAO inhibitor pargyline (PGL; 10 μm, 15-min pretreatment). In B–D, cells were pretreated with 100 μm l-DOPA for 1 h before the recordings to elevate DAcyt to levels detectable by IPE. Incubations with all drugs were done at 37 °C, whereas recordings were performed at room temperature. E, an in vitro assay of MAO-A activity performed at room temperature using a chemiluminescent substrate shows that MPP+ (IC50 = 1 μm) is a more potent enzyme inhibitor than pargyline (IC50 = 5.5 μm) or DA (IC50 = 150 μm). F, the Lineweaver-Burk plot demonstrates that MPP+ is a competitive inhibitor of MAO.