Figure 3.

Microglia limit the progression of CSA microcavities into large lesions in response to morphogenetic constraints

(A) IBA1, Spp1, and Mac2 immunostaining findings indicate that ATM line microcavities (solid arrowheads), visible by Hoechst counterstaining at E14.5 (n = 6, from at least two distinct litters).

(B) Transmitted electron microscopy (EM) also reveals the presence of microcavities (solid arrowheads), lined with amoeboid microglia (green pseudo-color), and containing membrane fragments (open arrowheads) (n = 3, from at least two distinct litters).

(C and D) Coronal hemisections (C) from control and Emx1^cre/+;RhoA^fl/fl mice, the latter displaying a periventricular nodular heterotopia (PVNH) visible from E18.5 onward (dotted lines). Hoechst counterstaining shows the absence of CSA lesions in controls and mutants at E15.5 (open arrowheads), a striking CSA lesion (solid arrowheads) in 50% of the E18.5 mutants and in 100% of the mutants at P8 (n_controls-E15 = 6, n_{heterotopia-E15} = 5, n_controls-E18 = 8, n_{heterotopia-E18} = 10, n_controls-P8 = 10, n_{heterotopia-P8} = 15). Quantification in (D) uses values that represent the scoring of lesion severity, scored from 0 to 2, as detailed in Table S2.

(E) Coronal hemisections of E15.5 brains from Brn4^cre/+; Wnt3a^dta/+ embryos show an ablation of the thalamus (white asterisk) and a global change in brain shape but no marked impact on the CSA (n_controls = 4, n_{thalamusdeleted} = 5).

(F and G) Coronal hemisections (F) of E15. 5 brains from control and Brn4^cre/+; Wnt3a^dta/+ mice exposed to PLX3397 between E12.5 and E15.5. While PLX3397-exposed controls consistently displayed lesions (open arrowheads), PLX3397-treated mutants exhibited smaller lesions or no lesions (solid arrowheads) despite effective local depletion as assessed by IBA1 staining (n_controls = 4, n_{thalamusdeleted} = 5, n_PLX3397 = 8, n_{thalamusdeleted-PLX3397} = 5, from at least two distinct litters). Quantification in (G) uses values that represent the scoring of lesion severity, scored from 0 to 2, as detailed in Table S2.

Graphs show means ± SEM. Mann-Whitney U tests were performed for statistical comparison, ^∗p < 0.05; ^∗∗∗p < 0.001; ns, non significant (p > 0.05). Scale bars: 200 μm (A, left); 100 μm (A and F, high magnifications); 2.5 μm (B); and 500 μm (C, E, and F, low magnification).

Am, amygdala; CSA, cortico-striato-amygdalar boundary; PVNH, periventricular nodular heterotopia; Ncx, neocortex; Str, striatum; Th, thalamus.

See also Figure S3.