Apoptosis in a neonatal rat model of cerebral hypoxia-ischemia.
OBJECTIVE
The mechanisms of excitotoxic cell death in cerebral ischemia are poorly understood. In addition to necrosis, apoptotic cell death may occur. The purpose of this study was to determine whether an established model of cerebral hypoxia-ischemia in the neonatal rat demonstrates any features of apoptosis.


METHODS
Seven-day-old neonatal rats underwent bilateral, permanent carotid ligation followed by 1 hour of hypoxia, and their brains were examined 1, 3, and 4 days after hypoxia-ischemia. The severity of ischemic damage was assessed in the dentate gyrus and frontotemporal cortex by light microscopy. Immunocytochemistry was performed to detect the cleavage of actin by caspases, a family of enzymes activated in apoptosis. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) reactivity was examined in the cortical infarction bed and dentate gyrus. Neonatal rat brain DNA was run on agarose gel electrophoresis to detect DNA fragmentation. Ethidium bromide-staining and electron microscopy were used to determine whether apoptotic bodies, 1 of the hallmarks of apoptosis, were present.


RESULTS
The frontotemporal cortex displayed evidence of infarction, and in most rats the dentate gyrus showed selective, delayed neuronal death. Immunocytochemistry demonstrated caspase-related cleavage of actin. TUNEL and DNA electrophoresis provided evidence of DNA fragmentation. Ethidium bromide-staining and electron microscopy confirmed the presence of chromatin condensation and apoptotic bodies.


CONCLUSIONS
Features of apoptosis are present in the described model of cerebral hypoxia-ischemia. Apoptosis may represent a mode of ischemic cell death that could be the target of novel treatments that could potentially expand the therapeutic window for stroke.
Pulera
M R
MR
Department of Neurology, Veteran's Affairs Medical Center, Sepulveda, Calif., USA. mpulera@ucla.edu
Adams
L M
LM
Liu
H
H
Santos
D G
DG
Nishimura
R N
RN
Yang
F
F
Cole
G M
GM
Wasterlain
C G
CG
D000818:Animals; D000831:Animals, Newborn / Q000502:physiology*; D017209:Apoptosis / Q000502:physiology*; D001921:Brain / Q000378:metabolism / Q000473:pathology; D002545:Brain Ischemia / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology*; D002544:Cerebral Infarction / Q000473:pathology; D053938:DNA Fragmentation; D004587:Electrophoresis, Agar Gel; D004996:Ethidium; D005260:Female; D000860:Hypoxia / Q000473:pathology / Q000503:physiopathology*; D020287:In Situ Nick-End Labeling; D007202:Indicators and Reagents; D008297:Male; D008854:Microscopy, Electron; D051381:Rats; D017208:Rats, Wistar; D013194:Staining and Labeling
D007202:Indicators and Reagents; D004996:Ethidium