Histological preparation

Two, four, seven, and fourteen days after shamCCI or mCCI, rats were deeply anesthetized with isoflurane followed by transcardial infusion with phosphate-buffered saline (PBS) and then paraformaldehyde (4% w/v). The brain was removed, incubated overnight in paraformaldehyde, and cryoprotected with 15% (w/v) then 30% (w/v) sucrose in PBS. Coronal sections were collected at −4.9 mm from bregma and sagittal sections were collected between 1.0 and 1.9 mm from midline.

For electron microscopy, rats were deeply anesthetized with isoflurane 4 or 14 days after shamCCI or mCCI followed by transcardial infusion with PBS and then gluteraldehyde (4% (w/v)). A 1.0 × 0.5 × 0.5-µm tissue block was removed from the trunk of the corpus callosum 0–0.1 mm from midline. The block was incubated in gluteraldehyde overnight at 4℃. Blocks were moved to PBS, stained with 0.1% OsO₄ (w/v) and 1-µm sagittal sections prepared.