Figure 8.

γC3-mediated dendritic arborization is dependent on Axin1. A, Cortical neurons cultured from P0 pups and nucleofected (∼50% efficiency) at plating with Axin1 shRNA (previously characterized by Chen et al., 2015) or its scrambled control. Neurons were lysed at day in vitro (DIV)8 and lysates analyzed by Western blotting using antibodies against Axin1. As expected, Axin1 levels are drastically reduced in neurons expressing the Axin1 shRNA knock-down construct. B, Quantification of area under the curve for Sholl analysis of control and C3KO cultured cortical neurons transfected with GFP, either Axin1 shRNA or the scrambled control shRNA, ± a full-length γC3 construct and analyzed at DIV8. Bars to the left indicate reduced complexity; bars to the right indicate increased complexity. C, Representative traces of neurons from each transfection condition. C3KO neurons exhibit significantly reduced dendrite complexity in culture as in vivo. Arbor complexity can be rescued by re-expression of γC3, but only if normal Axin1 levels are present. Scale bar: 50 µm. n ≥ 25 neurons per condition. One-way ANOVA; Dunnett's multiple comparisons test. Error bars represent the SEM; *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001; ns = not significant.