Amitriptyline and desipramine induced ATP depletion and the activation of Bax. (A, B) SH-SY5Y cells were incubated with amitriptyline (Ami, 60 μM) or desipramine (Desi, 50 μM) for 12 and 24 h and then incubated with JC-1 dye [5 μM, (A)] or MitoTracker [100 nM for 12 h, B)] for 30 min. Arrows indicate the cells with JC-1 green monomer fluorescence, suggesting decreased mitochondrial membrane potential (A). Note the decreased fluorescence of MitoTracker in the drug-treated cells in (B). (C) SH-SY5Y cells were incubated with amitriptyline or desipramine and the cellular ATP level was determined using ATPlite assay kit at indicated times. Hydrogen peroxide was used as a positive control for ATP depletion (n = 3, **p < 0.01 and ***p < 0.001 vs. 0 h). (D) To monitor the Bax activation, the cells were treated with amitriptyline or desipramine for 24 h and then processed for immunostaining using anti-active Bax antibodies (arrows). Scale bar = 10 μm.