Supplementary Figure 4: Analysis of hippocampal neurons in Mbe/Mbe mutants

(a-m) Representative images of brains from 8-week old mice stained with sera against calretinin (a-f) and parvalbumin (h-m). Quantification showing the percentage of calretinin positive (g) or parvalbumin positive (n) interneurons in the oriens layer (OL). There was no significant difference when comparing Mbe/Mbe animals and littermate controls (+/+, +/Mbe) (n = 3 animals per genotype; calretinin: one-way ANOVA with a Tukey’s multiple comparisons test; +/+ vs Mbe/Mbe; P = 0.8137; parvalbumin: Kruskal-Wallis test with a Dunn’s multiple comparison test; +/+ vs Mbe/Mbe; P>0.9999). (o) Image of a Golgi stained pyramidal neuron from the hippocampus. Spines located on tertiary apical dendritic branches distanced more than 120μm from the soma (red line) were quantitated. (p) Quantification of spine density in +/+ and Mbe/Mbe animals reveals no significant differences between genotypes (n = 4-6 neurons per animal; n = 3 animals per genotype; two-tailed unpaired t-test; t₄ = 0.3386; P = 0.7519). (q) Schematic representation of CA1 pyramidal neurons that were traced in three dimensions and assessed by Scholl analysis. (r) Quantitation of dendritic intersections from the soma. Ectopic Mbe/Mbe neurons exhibit a significant decrease in complexity, that is most evident 100 μm to 160 μm from the soma (n = 3 cells per animal, n = 3 animals per genotype; two-way ANOVA with a Tukey’s multiple comparisons test; interaction P<0.0001, Supplementary Table 1). The scale bars in (c), and (j) show 500 μm, 100 μm in (f) and (m) and 50 μm in (o). Error bars show mean +/- standard error of the mean.