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. 2018 May 16;38(20):4791–4810. doi: 10.1523/JNEUROSCI.2423-17.2018

Figure 6.

Figure 6.

Canonical TGF-β/BMP signaling suppresses morphological development in hippocampal neurons through the downregulation of CRMP2. A, Hippocampal neurons were infected with lentiviruses expressing shRNAs against Crmp2. Expression of CRMP2 protein was measured by Western blotting using an anti-CRMP2 antibody. B, Quantification of CRMP2 protein level in A. C, Representative images of GFP (green) and MAP2 (red) staining of 6DIV hippocampal neurons infected with lentiviruses expressing CRMP2, shCRMP2-1, or shCRMP2-2. Scale bar, 50 μm. D, E, Quantification of total dendritic length (D) and branch numbers (E) of 6DIV hippocampal neurons infected with lentiviruses expressing CRMP2, shCRMP2-1, or shCRMP2-2. F, Quantification of dendrite complexity by Sholl analysis of 6DIV hippocampal neurons infected with lentiviruses expressing CRMP2, shCRMP2-1, or shCRMP2-2. G, H, Quantification of total axon length (G) and axon branch numbers (H) of 3DIV hippocampal neurons infected with lentiviruses expressing shCRMP2-1 or shCRMP2-2. I, Representative image of GFP and MAP2 staining of 6DIV hippocampal neurons treated with TGF-β1, and infected with lentiviruses expressing CRMP2. Scale bar, 50 μm. J, K, Quantification of total dendritic length (J) and branch numbers (K) of 6DIV hippocampal neurons treated with TGF-β1, and infected with lentiviruses expressing control or CRMP2. L, Quantification of dendrite complexity by Sholl analysis of 6DIV hippocampal neurons treated with TGF-β1, and infected with lentiviruses expressing control or CRMP2. M, Representative images of GFP and MAP2 staining of 6DIV hippocampal neurons treated with BMP2, and infected with lentiviruses expressing CRMP2. Scale bar, 50 μm. N, O, Quantification of total dendritic length (N) and branch numbers (O) of 6DIV hippocampal neurons treated with BMP2, and infected with lentiviruses expressing CRMP2. P, Quantification of dendrite complexity by Sholl analysis of 6DIV hippocampal neurons treated with BMP2, and infected with lentiviruses expressing CRMP2. Q, Representative images of GFP, tdTomato (red), and MAP2 (cyan) staining of 6DIV hippocampal neurons coinfected with lentiviruses expressing Smad4 and CRMP2. Scale bar, 50 μm. R, S, Quantification of total dendritic length (R) and branch numbers (S) of 6DIV hippocampal neurons coinfected with lentiviruses expressing Smad4 and CRMP2. T, Quantification of dendrite complexity by Sholl analysis of 6DIV hippocampal neurons coinfected with lentiviruses expressing Smad4 and CRMP2. U, Representative images of GFP, tdTomato, and MAP2 staining of 6DIV hippocampal neurons coinfected with lentiviruses expressing Smad1 and CRMP2. Scale bar, 50 μm. V, W, Quantification of total dendritic length (V) and branch numbers (W) of 6DIV hippocampal neurons coinfected with lentiviruses expressing Smad1 and CRMP2. X, Quantification of dendrite complexity by Sholl analysis of 6DIV hippocampal neurons coinfected with lentiviruses expressing Smad1 and CRMP2. Data are presented as the mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001 by one-way ANOVA, Tukey's post-test. N = 3 independent experiments; at least 50 neurons were analyzed in each experiment.