Fig. 5.

Inhibitory and releasing effect of aminorex. Uptake of substrate (SERT: 5-HT; NET: MPP+; DAT: dopamine) at increasing aminorex concentrations is shown in (A). IC₅₀ values for aminorex (SERT: 18.39 ± 1.12 μM; NET: 0.33 ± 1.07 μM; DAT: 0.855 ± 1.20 μM) are suggesting a comparable inhibitory effect to cocaine for NET and DAT. For SERT the inhibitory effect was more than 10-fold higher compared to cocaine (all experiments were performed three times). Substrate efflux after treatment with aminorex was measured for SERT (B), NET (C) and DAT (D). In brief, HEK 293 cells stably expressing human SERT, NET or DAT were preloaded with 0,4 μM [3H]5-HT or 0,1 μM [3H]MPP+ for 20 min at 37 °C in a final volume of 0.1 mL/well, transferred to small superfusion chambers (0.2 ml) and superfused with KRH buffer (25 °C, 0.7 ml × min⁻¹). After a washout period of 40 min to reach stable baseline for efflux, 10 μM monensin or buffer (control) was added after 4 min followed by 3 or 10 μM aminorex (SERT and NET) and 50 μM aminorex (DAT) respectively for 10 min. In addition we used 3 μM D-amphetamine as an positive control. Collection of fractions was carried out in 2 min interval. At the end of the experiment, cells were lysed in 1% SDS. While levamisole lead to an efflux of substrate via SERT and NET, which was enhanced by addition of monensin, no releasing effect of aminorex was observed at DAT (p = 0.9981). A slight increase could be observed in NET (p = 0.001) and aminorex could trigger a strong release in SERT (p < 0.0001). Data are shown as mean ± SEM of three independent experiments. p-values were calculated using a Two-way ANOVA (Bonferroni post-test).