Fig. 3

Maternal separation (MS) induces microglial alterations in prefrontal cortex and exacerbates the amyloid pathology in 5xFAD females. (A, top) Quantification of the percentage of Iba1 + area relative to the total PFC area and (A, bottom) CD68 + area relative to the total Iba1 + area in 2–3 sections/animal; n = 4–7 animals/group. B Representative microphotographs of microglia in the prefrontal cortex (PFC) of each experimental group at 4 months old using Iba1 and CD68 antibodies. Green: CD68; red: Iba1; blue: DAPI; Scale bar: 20 μm. C Representative microphotographs of microglia and Ab plaques in PFC from 5xFAD female and male mice at 4 months old. Microglia were stained using Iba1 (white) and Gal-3 (green) antibodies; Ab plaques were stained using 6E10 antibody (red) and DAPI (blue) for nuclei; scale bar: 20 μm. D Quantification of the percentage of 6E10 + area and (E) Gal-3 + area in 2–3 sections/animal and relative to the total area of PFC (for 6E10, n = 6–8 animals/group; for Gal-3: n = 3–5 animals/group). F (left) Ab plaque density in PFC of 5xFAD mice was represented (2–3 sections/animal, n = 5–8 animals/group). F (right) Representative microphotographs of Ab plaques using Congo Red Staining in the PFC of 5xFAD mice. G Representative blots of the brain fraction S3 corresponded to protofibrils from PFC of 5xFAD mice. H Relative intensity of APP in PFC sequential protein fraction isolation from 4-month-old mice normalized to Actin (soluble fraction: n = 3–4 animals/group; protofibrils: n = 4–6 animals/group; dense plaques: n = 3–4 animals/group; technical duplicates). Data are shown as mean ± SD. *P < 0.05