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. 2020 Feb 28;23(3):100957. doi: 10.1016/j.isci.2020.100957

Figure 2.

Figure 2

Susd4 KO Mice Exhibit Abnormal Neuronal Morphology and Hypertrophic Microglia

(A) Bielschowsky silver staining of cerebellum. Top panels, representative sagittal view of the whole cerebellum for a WT and a Susd4 KO mouse. Arrows indicate the dense-appearing basket cell layer in the Susd4 KO mouse cerebellum relative to the WT tissue. Scale bar, 200 μm. Lower panels, representative higher magnification view shows Purkinje cell layer and basket cells (arrows). Note “hairy” appearance of basket cells in the Susd4 KO specimen. Scale bar, 20 μm

(B) Representative immunostaining with SMI-31antibody, labeling phosphorylated neurofilament heavy chain (green), and calbindin antibody, labeling Purkinje cells (red), in cerebellum sections from Susd4 KO and WT mice; scale bar, 20 μm.

(C) Representative higher-magnification view showing “hairy” basket cell dendrites (SMI-31 immunostaining in green) surrounding Purkinje cell soma (calbindin immunostaining in red); scale bar, 10 μm.

(D) Golgi-Cox staining of brains. Representative images of dendritic spines for WT and Susd4 KO mice.

(E) Quantification of the number of spines per micrometer dendrite on secondary or tertiary branches from both apical and basal parts of the pyramidal cells in hippocampal CA regions of WT and Susd4 KO Golgi-stained mouse brains. Data represent the mean ± SEM. n = 3 mice per group; ∗∗∗∗: p < 0.0001.

(F) Iba1immunostaining (red) for microglia. Top panels, representative view of hippocampal regions from WT and Susd4 KO mice. Scale bar, 40 μm. Lower panels, representative higher magnification view shows ramified resting microglia (arrows in top panel) in WT in contrast with hypertrophic microglia (arrow heads in top panel) in Susd4 KO. CA, cornu ammonis. Scale bar, 10 μm.

(G) Quantification of Iba1+ cells per square millimeter in hippocampal regions from WT and Susd4 KO mice. Data represent the mean ± SEM. n = 3 mice per group. ∗: p < 0.05.

(H) Quantification of Iba1 fluorescence intensity/microglia from the stained hippocampal sections. Data represent the mean ± SEM. WT values were set to 1.0. n = 3 mice per group. ∗∗∗∗: p < 0.0001. Nuclei were visualized with DAPI staining (blue) for (B), (C), and (F).