Fig. 5

T. gondii infection decreased VGLUT2 expression in the mouse olfactory bulb. a Brains from mock-infected control mice and mice orally infected with 10 ME49 cysts for 30 and 60 days were collected after perfusion with ice-cold 4% PFA. Brain sections were fixed and stained with anti-VGLUT1 (green), anti-VGLUT2 (red), and mounted with DAPI (blue). Whole brain images were scanned using Leica Biosystems Scanscope and processed with image scope. Representative images were taken from the olfactory bulb at × 20 magnification. n = 4–5 mice per group, the experiment was repeated three times. Scale bar = 50 μm. b, c At least five different fields of the olfactory bulb and pre-frontal cortex were randomly selected from 2 to 4 mice per group. The intensity of VGLUT2 (b) and VGLUT1 (c) signal at different time points post infection was quantified. d Representative images of the olfactory bulb of mock-infected and T. gondii-infected mice stained with anti-GAD67 (green) and VGLUT2 (red). Scale bar = 50 μm, n = 4 mice per group. e The mean fluorescent intensity for GAD67 (green) was measured using Axiovision software from five different fields (n = 4 mice per group). f Representative image from the olfactory bulb of infected mice stained with anti-GAD67 (green) and anti-T. gondii (red). Scale bar = 50 μm. g Western blot analysis of VGLUT1/2 (55 and 65 kDa, respectively) and GAD67 (67 kDa) protein levels using whole brain samples from mock-infected controls and T. gondii-infected mice at days 30 and 60 post infection. GAPDH (37 kDa) was used as loading control. Blot shown is representative of three independent experiments. h–j Densitometric analysis of Western blot data from g. The results are expressed as the ratio of densities of GAD67 or VGLUT1/2-specific bands to GAPDH bands. Data are shown as mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001, **** p < 0.0001