Figure 4. Regional expression of VGLUT3, VGLUT3A224V/+, VGLUT3A224V/A224V, and VGLUT3A224V/− in the CNS of mice at different ages. A–J, Detection and quantification of VGLUT3, VGLUT3A224V/+, VGLUT3A224V/A224V, and VGLUT3A224V/− mRNA and protein (Prot) expression by in situ hybridization or immunoautoradiography on coronal mouse brain sections taken at P10 (A, B) and at 3 months (C–F), 6 months (G, H), or 12 months (I, J) in the striatum (Str), hippocampus (Hi), dorsal raphe (DR), and median raphe (MR). WT, VGLUT3A224V/+, and VGLUT3A224V/A224V mice expressed a similar level of transcripts (B, D, H, J, left; Mann–Whitney U test for B, J; Kruskal–Wallis test for D, H; p > 0.05, n = 8). B, D, H, right, F, Loss of the protein in all areas from P10 until 1 year in VGLUT3A224V/A224V mice (n = 8 for each genotype, Mann–Whitney U test or Kruskal–Wallis test, *p < 0.05, **p < 0.01, ***p < 0.001). In VGLUT3A224V/+ mice, there is a 34% decrease in VGLUT3 in all areas at 3 and 6 months (D, H). E, F, In this experiment, the protein expression of VGLUT3 was compared in WT, VGLUT3A224V/A224V, and VGLUT3A224V/− mice (n = 8) in the striatum, hippocampus, and dorsal raphe. VGLUT3 expression decreased by 70% in VGLUT3A224V/A224V mice and further decreased by 84% in VGLUT3A224V/− mice in the striatum (Kruskal–Wallis test; in the striatum, WT vs VGLUT3A224V/+, p = 0.0043; WT vs VGLUT3A224V/A224V, p = 0.0043; WT vs VGLUT3A224V/−, p = 0.0012; in the hippocampus, WT vs VGLUT3A224V/+, p = 0.01; WT vs VGLUT3A224V/A224V, p = 0.0043; WT vs VGLUT3A224V/−, p = 0.0012; in raphe nuclei, WT vs VGLUT3A224V/+, p = 0.0571; WT vs VGLUT3A224V/A224V, p = 0.0159; WT vs VGLUT3A224V/−, p = 0.0286). K, L, Western blot detection (K) and quantification (L) of VGLUT3 in the cortex, striatum, and hippocampus of VGLUT3A224V/+ and VGLUT3A224V/A224V mice at 3 months (n = 5; Kruskal–Wallis test, **p < 0.01).