Figure 3

Down-regulation of perineuronal nets (PNNs) after 8 weeks of 4-methylumbelliferone (4-MU) feeding in uninjured animals, with or without daily treadmill training, and the re-appearance of perineuronal nets (PNNs) after 2 months of wash-out period. (A) Representative fluorescent images showing Wisteria floribunda agglutinin (WFA) and aggrecan (ACAN) positive PNNs around cells in the ventral horns and their colocalization (WFA + /ACAN +) in thoracic spinal cord (Th8) in uninjured animals in all 5 groups after 8 weeks of feeding and after 2 months of wash-out period. Green arrows indicated the WFA positive PNNs enwrapped cells; red arrows indicated ACAN positive PNNs enwrapped cells and gray arrows indicated cells where WFA/ACAN positive signal colocalizes. Scale bar 100 μm. (B–D) Quantitative analysis of WFA positive or ACAN positive PNNs. Data showed mean ± SEM (n = 3 animals per groups, 3 sections per animal). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, two-way ANOVA, Dunnett's multiple comparison. (E) Representative confocal images showing WFA positive and ACAN positive PNNs surrounding ChAT positive motoneurons in the thoracic rat spinal cord (Th8) in placebo, 4-MU treated and wash-out group in ventral horn. Scale bar 30 μm.