Figure 2. OGD in lG partly preserves Δψ_m (i.e. delays depolarization) caused by subsequent NaN₃ treatment.

(A) Sodium azide (NaN₃) causes concentration-dependent decline of Δψ_m in both experimental conditions. However, OGD in lG medium significantly delayed the decrease in JC-1 fluorescence ratio caused by treatment with 5 mM NaN₃ during simulated reperfusion. (B) Expectedly, both NaN₃ and H₂O₂ show concentration-dependent effect by lowering the JC-1 red/green fluorescence ratio. Astrocytes were stained with JC-1 either before or during the treatment with NaN₃ (marked as pre-dye loading and dye loading under ETC inhibition, respectively). NaN₃ has not affected the cell membrane organization allowing at the same time JC-1 to enter the cytoplasm and mitochondria within. (C) Representative fluorescent micrographs of astrocytes labeled with JC-1 Original micrographs were converted to tritanope color palette (ImageJ 1.48a). Depolarization is visible in the normoxic group (b, c) (seen as concentration-dependent decrease in magenta and increase in blue color), but it is more pronounced than in OGD group after treatment with NaN₃ (e, f). Some mitochondria remained partly depolarized. Data are expressed as a percentage normalized to the red/green fluorescence ratio values of untreated control (the first bar from left). Significant differences are indicated by **p<0.01 with respect to untreated control, ##p<0.01 between treatment and its respective control, ΔΔp<0.01 between different inhibitor concentrations.