Mitochondrial membrane depolarization and mitochondrial or cardiomyocytes respiration. Respiration was measured using complex I substrates glutamate/malate/pyruvate in the absence (Control) or presence of 0.5 mM isoflurane (Iso). Respiration state 3 was initiated with addition of ADP (A, 1 mM for cardiomyocytes or B, 0.2 mM for mitochondria] and expressed as nmol nmol O₂ min⁻¹ mg protein⁻¹. Each value was expressed as the mean ± SD, one-way ANOVA, Bonferroni post hoc test, ***P < 0.0001 versus Control wild type (WT), n = 5 separate experiments. Insets (A, B) represent the value of state 2 (respiration without ADP) and respiratory control index (RCI) in all groups. Mitochondrial membrane potential (ΔΨm) (C) was assessed by fluorescence of tetramethylrhodamine methyl ester perchlorate (TMRM) without Iso (Control) or with 0.5 mM Iso (Iso). Each value was expressed as the mean ± SD, two-tailed Student t test, *P = 0.043: Iso WT versus Control WT, **P = 0.017: Iso CypD-KO versus Control cyclophilin D knockout (CypD-KO).