Substrate release by p-chloroamphetamine and PAL-1045. (A) Release of [³H]MPP⁺ from preloaded HEK293 cells expressing SERT. [³H]MPP⁺ release is plotted as percent [³H]MPP⁺ over time. The experimental conditions are indicated in the black bars: 3 µM p-chloroamphetamine (open squares), 3 µM p-chloroamphetamine + 10 µM monensin (open circles), 3 µM PAL-1045 (open diamonds), and 3 µM PAL-1045 + 10 µM monensin (open triangles). 100 µM ouabain (open bar) was present starting at t = −40 min and throughout the experiment. For statistical analysis, the total area under the curve after basal release (i.e., fractions t = 0 to t = 4 min) was calculated for each individual trace. **, P < 0.01; n.s., not significant. One-way ANOVA followed by Bonferroni’s multiple comparison test. (B) Calculated [³H]MPP⁺ release rate for one cell in femtomole/minute (=V_max). (A and B) Data are means ± SD, n = 8–9 independent observations per condition.