Figure 3. Progressive degeneration of DA neurons in the midbrain of G59S Tg and G71R Tg mice. (A) Immunohistochemical images show TH staining in the coronal midbrain sections of 24-month-old nTg, WT Tg, G59S Tg, and G71R Tg mice. Scale bar: (left) 500 μm (low-magnification images) and (right) 50 μm (high-magnification images). Note the dystrophic TH-positive DA neurites in the SNr of G59S Tg and G71R Tg mice. (B) Bar graphs depict the unbiased stereological estimation of the number of TH-positive DA neurons in the SNc of 24-month-old nTg, tTA, tetO-WT, WT Tg, tetO-G59S, G59S Tg, tetO-G71R, and G71R Tg mice (n = 4 per genotype). Data are presented as mean ± SEM. One-way ANOVA with Dunnett’s multiple comparisons test was used for statistical analysis (the mean of each genotype was compared with the mean of nTg). ****P < 0.0001. ns: Not significant. (C) Bar graphs depict the unbiased stereological estimation of the number of TH-positive DA neurons in the SNc of 1-, 6-, and 12-month-old nTg, WT Tg, G59S Tg, and G71R Tg mice (n = 4 per genotype per time point). Data are presented as mean ± SEM. At each time point, one-way ANOVA with Dunnett’s multiple comparisons test was used for statistical analysis (the mean of each genotype was compared with the mean of nTg). **P = 0.0099, ***P = 0.0001. (D) Bar graphs depict the unbiased stereological estimation of the number of TH-positive DA neurons in the VTA of 24-month-old nTg, WT Tg, G59S Tg, and G71R Tg mice (n = 4 per genotype). Data are presented as mean ± SEM. One-way ANOVA with Dunnett’s multiple comparisons test was used for statistical analysis (the mean of each genotype was compared with the mean of nTg). ****P < 0.0001. (E) Bar graphs quantify the area percentage of dystrophic TH-positive DA neurites in the SNr of 2- and 24-month-old nTg, WT Tg, G59S Tg, and G71R Tg mice (at each time point, n = 4 animals per genotype and 5 sections per animal). Dystrophic DA neurites were defined as neuritic varicosity ≥ 25 μm2. Data are presented as mean ± SEM. At each time point, one-way ANOVA with Dunnett’s multiple comparisons test was used for statistical analysis (the mean of each genotype was compared with the mean of nTg). *P = 0.0259, ****P < 0.0001. TH: Tyrosine hydroxylase.