Evidence for apoptosis after intracerebral hemorrhage in rat striatum

The overall hypothesis that cell death after intracerebral hemorrhage is me diated in part by apoptolic mechanisms was tested. Intracerebral hemorrhage was induced in rats using stereotactic infusions of 0.5 U of collagenase ( 1-mu L volume) into the striatum. After 24 hours, large numbers of TUNEL-po sitive stained cells with morphologies suggestive of apoptosis were present in the center and periphery of the hemorrhage. Double staining with Nissl and immunocytochemical labeling with antibodies against neuronal nuclei and glial fibrillary acidic protein suggested that these TUNEL-positive cells were mostly neurons and astrocytes. Electrophoresis of hemorrhagic brain ex tracts showed evidence of DNA laddering into similar to 200-bp fragments. W estern blots showed cleavage of the cytosolic caspase substrate gelsolin. T he density of TUNEL-positive cells at 24 and 48 hours after hemorrhage was significantly reduced by treatment with the broad-spectrum caspase inhibito r zVADfmk. It was unlikely that apoptotic changes were due to neurotoxicity of injected collagenase because TUNEL-positive cells and DNA laddering wer e also obtained in an alternative model of hemorrhage where autologous bloo d. was infused into the striatum. Furthermore, equivalent doses of collagen ase did not induce cell death in primary neuronal cultures. These results p rovide initial evidence that apoptotic mechanisms may mediate some of the i njury in brain after intracerebral hemorrhage.