Figure 4
Functional characterization of dopamine (DA) response in JumpIn-DAT cells. (a) Representative xCELLigence growth curves after cell seeding ± 1 µg/ml dox, inhibitor pretreatment and dopamine addition. (b) Effect of dox on JumpIn-DAT cell response (TRACT assay) upon stimulation with 10 µM dopamine (set at 100%) (■) and effect of dox on cell surface expression of DAT detected by HA-tag ELISA (teal ♦, expressed as fold over mock). Data are shown as mean ± SD (HA-tag ELISA) or SEM (TRACT assay) of two or three separate experiments performed in quintuplicate or duplicate, respectively. (c) Representative vehicle-corrected xCELLigence traces of JumpIn-DAT cells in the absence of dox (–dox) and (d) JumpIn-DAT cells in the presence of 1 µg/ml dox (+dox) after stimulation with increasing concentrations of dopamine. Data is normalized prior to agonist addition at time = 0 min. (e) Concentration-effect curves of dopamine on JumpIn-DAT cells ± dox are shown as the net AUC of the first 30 min after stimulation normalized to the cell response of 316 µM dopamine. (f) Cell response of 31.6 µM dopamine (red bar, set at 100%) on dox-treated JumpIn-DAT cells pretreated for 1 h with 1 µM of one of following GPCR antagonists: SCH23390 (dopamine D1-like), raclopride (dopamine D2-like), doxazosin (alpha-1 adrenergic), yohimbine (alpha-2 adrenergic), propranolol (beta adrenergic). Data are shown as mean ± SEM of three to nine individual experiments each performed in duplicate. Comparison of multiple mean values to vehicle control was done using a one-way ANOVA with Dunnett’s post-hoc test. ***p < 0.001.
