Two region-dependent pathways of eosinophilic neuronal death after transient cerebral ischemia

Various types of eosinophilic neurons (ENs) are found in the post-ischemic brain. We examined the temporal profile of ENs in the core and peripheral regions of the ischemic cortex, and analyzed the relationship to the expression of various cell death-related factors. Unilateral forebrain ischemia was induced in Mongolian gerbils by transient common carotid artery occlusions, and the brains from 3 h to 2 weeks post-ischemia were prepared for morphometric and immunohistochemical analysis of ENs. ENs with minimally abnormal nuclei and swollen cell bodies appeared at 3 h in the ischemic core and at 12 h in the periphery. In both locations multiple cell death-related factors including calcium, micro-calpain, cathepsin D, 78 kDa glucose-regulated protein (GRP78) and ubiquitin were activated. In the ischemic core, pyknosis and irregularly atrophic cytoplasm peaked at 12 h, which was associated with significant increases in staining for calcium and micro-calpain. ENs with pyknosis and scant cytoplasm peaked at 4 days and were positive for TUNEL and calcium staining. In the ischemic periphery, ENs had slightly atrophic cytoplasm and sequentially developed pyknosis, karyorrhexis and karyolysis over 1 week. These cells were positive for TUNEL and calcium staining. All types of EN were negative for caspase 3. There may be two region-dependent pathways of EN changes in the post-ischemic brain: pyknosis with cytoplasmic shrinkage in the core, and nuclear disintegration with slightly atrophic cytoplasm in the periphery. This difference coordinates different activation patterns of cell death-related factors in ENs.