Fig. 6

Mitochondrial transport and function are impaired at glucolipotoxicity-stressed RGCs in a GSK3β-dependent manner. (a) Mitochondrial axonal trafficking was determined by infecting RGCs with an adenovirus vector carrying a foreign gene for mitochondrial complex IV (Ad-GFP-Mito). Representative fluorescence images for GFP-labeled mitochondria within axons (upper panels) and kymograph images of axonal mitochondrial movement (middle and bottom panels) are shown. Traces of moving mitochondria are indicated with white arrow. The average transport speed of movable mitochondria was calculated and expressed as mito velocity (μm/min). (b) Activity of mitochondrial complex I and complex IV was measured by spectrophotometry and expressed as nmol/min/mg protein. (c) Representative images of double immunofluoresence for Tau 5 (red) and complex I (green). Scale bar, 40 μm. (d) Western blotting for complex I and complex IV from whole cell lysates (Total) or synaptosome fractions (Syn). Intensities were quantified and normalized against the level of GAPDH and expressed as percentage of protein abundance under stimulation relative to control. Data are means ± SEM of three (a, d) or four (b) independent experiments. ^*P < 0.05 and ^**P < 0.01 vs control; ^#P < 0.05 and ^##P < 0.01 vs HG + PA. NS, no significant difference