Whole head were harvested after intracardiac perfusion with 2% formaldehyde in PBS and then fixed with 2% formaldehyde in PBS for 24–72 hours. We next embedded the brains in paraffin, and 5-μm-thick paraffin sections were cut using microtome and mounted on glass slides. For IHC (immunohistochemistry), tissues were deparaffinized, and antigen retrieval was performed in a pH 9 solution (DAKO) at 96°C. To prevent nonspecific staining, sections were incubated with 5% normal donkey serum (Jackson ImmunoResearch) in PBS before incubation with the respective primary and secondary antibodies. The primary antibodies used are listed in Table S3 . Apoptag (ApopTag Peroxidase In Situ Apoptosis Detection Kit, #S7100, Millipore) was used as a marker for apoptosis.
The number of neurons in the cortex was assessed by quantifying the number of NeuN+ nuclei in mosaics of the coronal sections of the region of the cortex exposed to the compression apparatus, i.e. the anterior cingulate area, the primary and secondary motor area and the first part of primary somatosensory area. The structures were identified using the mouse brain atlas map.
The number of neurons in the cortex was assessed by quantifying the number of NeuN+ nuclei in mosaics of the coronal sections of the region of the cortex exposed to the compression apparatus, i.e. the anterior cingulate area, the primary and secondary motor area and the first part of primary somatosensory area. The structures were identified using the mouse brain atlas map.