Reduced expression and activity of cytochrome c oxidase before the onset of neurodegeneration. (A) Representative ×63 confocal images of tyrosine hydroxylase (TH) and mtCOI immunostaining in the substantia nigra (SN) of 5-month-old control, K320E/Wt^DaN and K320E/K320E^DaN mice. Scale bar = 10 µm. (B) Normalized mean fluorescent intensity (FI) of mtCOI signal in SN and ventral tegmental area (VTA) dopaminergic neurons (69–96 SN neurons, 88–125 VTA neurons, from 3–5 mice per group). P-values were calculated using one-way ANOVA with Bonferroni’s correction for multiple comparison. Data are presented as superplots including mean ± standard error of the mean (SEM). (C) Representative images of a dual staining for cytochrome c oxidase (COX) and succinate dehydrogenase (SDH) enzyme activity revealing blue, COX-deficient cells in the SN and VTA (red arrowheads) of 5-month-old K320E/Wt^DaN and K320E/K320E^DaN mice. Scale bar = 500 μm and 20 μm (insets).