Fig. 4

T. gondii infection disrupted expression of the NMDA receptor in the mouse brain. a Brains from mock-infected control mice and mice orally infected with 10 ME49 cysts for 15, 30, and 60 days were collected after perfusion with ice-cold 4% PFA. Brain sections were fixed and stained with neurotracer (green), anti-NMDAR antibody (red), and counterstained with DAPI (blue). Representative images were taken from hippocampal and cortical regions at × 20 magnification n = 3–4 mice/group. The experiment was repeated twice. The inset image from day 60 shows co-localization of NMDAR and neurotracer. b, c Intensity of NMDAR (Texas Red) from hippocampus (b) and cortex (c) at different times post infection were analyzed. Five different fields were randomly selected from two different animals per group. d, e Number of neurotracer positive cells (AF488) from hippocampus and cortex, respectively. n = 2–4 per group. f Western blot analysis of NMDAR (105 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. g Densitometric analysis of Western blot data from f using ImageJ. The results are expressed as the ratio of densities of NMDAR-specific bands to GAPDH bands. h Brain sections from infected mice were stained with antibodies to T. gondii (red) and NMDAR (green). Brain sections were counterstained with DAPI (blue). i Mouse cortical neuronal cells (E15 cortical neurons) were infected with 1 MOI of T. gondii (mCherry pru strain, red) for 24–72 h. Infected neurons were fixed, permeabilized, and stained with anti-MAP-2 (white) and anti-NMDAR (green) antibodies. Cells were counterstained with DAPI (blue). Representative images of the axons of infected neurons 24–72 h post infection are shown. Images are representative of two independent experiments. Data are shown as mean ± SEM. *p < 0.05