Abstracts

Seizure-induced neuronal death is morphologically necrotic and caspase-independent, despite internucleosomal DNA cleavage (DNA laddering). We investigated whether seizure-induced neuronal necrosis, like cerebral ischemia, involves translocation of lysosomal cathepsin B and DNAse II to the cytoplasm and nucleus, where the former could hydrolyze cytoplasmic proteins and the latter could cause DNA laddering. Adult male Wistar rats were subjected to 3-h lithium-pilocarpine-induced status epilepticus (LPCSE), after which seizures were stopped with diazepam and phenobarbital. Twenty-four h later rats were killed by in situ transcardiac perfusion-fixation of their brains, with subsequent removal and processing for H [amp] E staining, TUNEL and cathepsin B and DNase II immunoreactivity. Twenty-four brain regions were assessed for the degree of neuronal death, using H [amp] E and TUNEL, and for translocation of cathepsin B and DNase II by brightfield immunohistochemical and immunofluorecent light microscopy and by immunogold electron microscopy (IEM) in the piriform cortex. The extent of neuronal necrosis and TUNEL positivity was similar to what we have described previously. In the neurons of control rats (n=3) cathepsin B and DNase II immunoreactivity by light microscopy appeared in a punctate distribution in the cytoplasm, and both were found in the cytoplasm but not the nuclei of neurons in the piriform cortex by IEM. Twenty-four h after SE, there was a striking translocation of DNase II and cathepsin B immunoreactivity to the pyknotic nuclei of necrotic neurons, as well as a diffuse increased immunoreactivity of both in the cytoplasm (n=3). These results indicate that following SE, lysosomal acid hydrolases such as cathepsin B can gain access to and damage nuclear as well as cytoplasmic proteins, and that lysosomal DNase II also translocates to nuclei, where it may be at least in part responsible for seizure-induced DNA laddering. (Supported by The Department of Veterans Affairs.)