Figure 2. Increased Dendritic Complexity Upon Loss of MeCP2 S421 Phosphorylation.

(A) Dissociated E16+21–22 DIV cortical neurons from MeCP2 S421A knock-in mice or wild-type littermates transfected with GFP to visualize dendritic morphology. Maximal intensity projections of a series of confocal planes for representative 25X images are shown.

(B) Sholl analysis measuring the number of dendritic branch points at 10 μm radial increments from the cell soma reveal an increase in dendritic branching in the MeCP2 S421A cells (p < 0.05, two-way repeated measures ANOVA) due to increased branch points 60–80 μm from the cell soma (p < 0.01, Bonferroni posttests). Wild-type (n=26) and MeCP2 S421A knock-in (n=33) neurons from two independent experiments.

(C) Representative images of single layer V pyramidal neurons expressing GFP under from the Thy1 promoter in 4-week-old forebrains of GFP-M transgenic mice. Tracings are shown of maximal intensity projections of a series of 10X confocal planes taken from coronal cortical sections.

(D) Sholl analysis of dendritic branch points at 10 μm radial increments from the cell soma reveals an increase in dendritic branching of MeCP2 S421A layer V pyramidal neuron apical dendrites 530 (p < 0.05), 540 (p < 0.01), 550 (p < 0.01) and 560 μm (p < 0.05) from the cell soma (two-way repeated measures ANOVA with Bonferroni posttests). Wild-type (n=29) and MeCP2 S421A knock-in (n=16) neurons from three independent littermate pairs.

(E) The maximum number of dendritic intersections ≥ 400 μm from the cell soma observed by Sholl analysis of each GFP-M+ layer V pyramidal cell in MeCP2 WT and S421A knock-in mice. The increase in maximum number of intersections in the MeCP2 S421A knock-in cells compared to WT indicates that the MeCP2 S421 knock-in apical tuft is more complex regardless of the exact distance of the apical tuft from the cell soma. p < 0.01 by Student’s t-test. All data shown in Figure 2 are mean ± SEM.