Age-related changes in ultrastructural features of cathepsin B- and D-containing neurons in rat cerebral cortex

The present study examines age-related changes in the subcellular localizat ion of cathepsin B (cath B) and cathepsin D (cath D), as well as morphologi cal features of the cathepsin-immunoreactive (ir) neurons in rat cerebral c ortex. Sprague-Dawley rats were studied at 3 and 26 months. By immunoelectr on microscopy cath B- or cath D-immunoreactivities were found in many, but not all, pyramidal neurons. In young rat cerebral cortical neurons, cath B was observed not only in lysosomal systems such as multivesicular bodies, d ense bodies, and lipofuscin granules, but also in extralysosomal sites. By contrast, cath D was confined mainly to lysosomal systems in young rats. In aged rats, cath B showed a similar pattern in its subcellular localization compared to the young control, but some of the dense bodies containing cat h B was closely apposed to the outer nuclear envelope. These cells exhibite d a relatively normal appearance. Regardless of subcellular localization, a pproximately 10% of cath B-ir neurons displayed ultrastructural disturbance s presumed to indicate an early stage of degeneration. The nucleus was inde nted, nuclear boundary was indistinct, nuclear pore structures appeared sep arately with high frequency, and the endoplasmic reticulum appeared to be a ffected. In addition to its presence in lysosomal structures, cath D-immuno reactivity in aged cerebral cortex was noted prominently in the cytosol as diffuse granules. About 37% of cath D-ir cells showed this age-related chan ge. Among the neurons with the diffusely scattered form of cath D, approxim ately 70% of cells exhibited the degenerating features. These cells were ch aracterized by large amounts of diffuse cath D, reduced cellular size, loss of the nuclear boundary, scattered nuclear pore structures, an often fragm entation of the nucleus, disturbances of endoplasmic reticular system, and in advanced stages, condensed nucleus and poor preservation of almost cytop lasmic organelles. Though some of these features were also found in cath B- ir neurons, findings of overt degeneration, such as fragmented and condense d nuclei and impaired almost cytoplasmic organelles, were generally not obs erved in cath B-ir neurons. In addition, lipofuscin aggregates containing c ath D were observed frequently in the extracellular space close to sites of ruptured plasma membrane, whereas in the sections stained with anti-cath B antibodies, large-sized lipofuscin aggregates showed only very weak or no cath B-immunoreactivity at all. Taken together, the present results suggest that cath D and cath B may be regulated differently and play their specifi c roles in the aging of the brain, especially, the change in location of ca th D from the lysosomal system to the cytosol in the aged brain may play an important role in age-related cell death. (C) 1999 Published by Elsevier S cience B.V. All rights reserved.