Immunohistochemical evidence and ultrastructural compartmentalization of a new antioxidant enzyme in the rat substantia nigra

We previously described in the rat the presence of dehydroascorbate reductase, an enzyme regenerating ascorbic acid, which is constantly lost during oxidative processes occurring at a fast rate within the central nervous system. In the present study, we specifically evaluate the occurrence of this enzyme in the rat substantia nigra by using immunohistochemistry, and by analyzing the neuronal compartmentalization of dehydroascorbate reductase within nigral neurons by immunoblotting and transmission electron microscopy coupled with immunocytochemistry. The enzyme occurs in various portions of the substantia nigra, but it is more abundant in the ventromedial part extending through the ventral tegmental area, and the dorsal portion, involving the pars compacta. Within nigral neurons, the cytosolic enzyme is present in a perinuclear position, close to mitochondria, and in the nuclear membrane; we also found the enzyme in nigral axons close to the myelin sheath. In addition, dehydroascorbate reductase was present in the nucleus of nigral neurons. The nuclear occurrence of the enzyme was confirmed by immunocytochemical labelling and immunoblotting of isolated nuclei. The nuclear enzyme was constantly evident as clusters of immunogold particles on chromatin. This localization suggests new roles for dehydroascorbate reductase (eg. prevention of DNA oxidative damage and regulation of gene transcription).