APOPTOTIC DNA FRAGMENTATION IN THE RAT CEREBRAL-CORTEX INDUCED BY PERMANENT MIDDLE CEREBRAL-ARTERY OCCLUSION

Recent investigations have demonstrated internucleosomal DNA fragmenta tion in ischemic neuronal tissue. This type of fragmentation is charac teristic of programmed cell death or apoptosis and suggests that neuro nal death in stroke may be more complex than simple necrotic death. Th e present experiments provide a detailed examination of the regional l ocalization and time course for apoptotic DNA fragmentation in the cer ebral cortex following focal cerebral ischemia. Spontaneously hyperten sive rats were subjected to permanent right middle cerebral artery occ lusion and the cerebral cortices were examined for evidence of DNA fra gmentation using electrophoretic, flow cytometric, and histological ap proaches. An electrophoretic examination of cortical DNA at 24 h after the occlusion indicated that the majority of nucleosomal ladders were in the transition zone or penumbra and the core of the infarction, wi th no fragmentation apparent in the contralateral normal cortex. A flo w cytometric analysis of DNA fragmentation in intact cells revealed a similar pattern, with increased fragmentation observed in ischemic cor tex vs, the contralateral cortex. Saggital sections taken 1.5 mm later al to midline were collected from animals at 1, 4, and 24 h after the infarction and DNA fragmentation was examined histologically by termin al deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling (TUNEL) staining. Quantitative analysis of these sections indicated t hat DNA fragmentation can be observed in the anterior and central area of the infarctions as soon as 1 h after the occlusion and that the ex tent and magnitude of the fragmentation increases at 4 and 24 h. Thus, nucleosomal DNA fragmentation accompanies neuronal death following fo cal cerebral ischemia and supports the hypothesis that programmed cell death may contribute to the death of neurons in stroke.