The efficacy of the mitochondrial blockers, cyanide and rotenone, was confirmed using a Seahorse Cell Mito Stress Test in an Agilent Seahorse XFp setup. Oxygen consumption rate (OCR) in astrocytes was measured upon addition of indicated inhibitors and mitochondrial respiration was blocked using the same agents (2.5 mM NaCN or 5 µM rotenone) as in ATP and VRAC activity assays. As a positive control we used combination of antimycin A and rotenone from the Cell Mito Stress kit. Data are the mean values ±SEM of six independent wells in three independent cell preparations. (top inset) Graphical representation of mitochondrial processes targeted by the Cell Mito Stress Test components. Abbreviations: Ant. A, antimycin; FCCP, mitochondrial uncoupling agent; NaCN, sodium cyanide; Oligo., oligomycin; Rot, rotenone.