DELAYED NEURONAL DEATH IN THE CA1 PYRAMIDAL CELL LAYER OF THE GERBIL HIPPOCAMPUS FOLLOWING TRANSIENT ISCHEMIA IS APOPTOSIS

The CA1 pyramidal neurons in the hippocampus are selectively vulnerabl e to transient ischemic damage. in experimental animals, the CA1 pyram idal neurons undergo cell death several days after brief forebrain isc hemia. It remains, however, unknown whether this delayed neuronal deat h is necrosis or apoptosis. To investigate the degenerating processes of the CA1 pyramidal neurons in gerbil hippocampus after brief ischemi a, lysosomal and nuclear alterations in the cells were examined using immunocytochemistry, in situ nick-end labeling, and Southern blotting. By light and electron microscopy, immunoreactivity for cathepsins B, H, and L, representative lysosomal cysteine proteinases, increased in the CA1 pyramidal neurons 3 d after ischemic insult, which showed cell shrinkage. By morphometric analysis, the volume density of cathepsin B-positive lysosomes markedly increased 3 d after ischemic insult, whi le that of autophagic vacuole-like structures also increased at this s tage, suggesting that cathepsin B-immunopositive lysosomes increasing in the neurons after ischemic insult are mostly autolysosomes. Nuclei of the CA1 neurons were nick-end labeled by biotinylated dUTP mediated by terminal deoxy-transferase 3 and 4 d after ischemic insult, but no t in the prior stages. Simultaneously, dense chromatin masses appeared in nuclei of the neurons. By Southern blotting, laddering of DNA occu rred only in CA1 hippocampal tissues obtained 4 d after ischemic insul t. Confocal laser scanning microscopy demonstrated that the fragmented DNA in the CA1 pyramidal layer was phagocytosed by microglial cells. The results suggest that delayed death of the CA1 pyramidal neurons af ter brief ischemia is not necrotic but apoptotic.