The aim of this study is to investigate of inducible nitric oxide synthase (iNOS) and nitrotyrosine (NT) expression in the cerebellum of dogs naturally infected with canine distemper virus (CDV), a disease characterized by demyelination in the white matter of the cerebellum. Cerebellum samples collected from naturally infected twenty-three which were 14 acute and 9 chronically infected. with CDV and six non-infected dogs. The presence of CDV infection was confirmed using histopathology and immunohistochemical localization of CDV antigen in glial cells of the white matter of the cerebellum. The CDV infected cerebella were also categorized Luxol Fast Blue staining according to the severity of demyelination as mild (n=7), moderate (n=9), and severe (n=7). The streptavidin-biotin-peroxidase technique was used to detect immunolocalisation of the CDV antigen, iNOS, NT and glial fibrillary acidic protein (GFAP). CDV antigens, immunopositive astrocytes, oligodendrocytes and granular neurons were seen in the white matter of the infected dogs. In the control dogs, iNOS and NT immunopositivity were not seen in the white matter of the cerebellum. In CDV infected dogs, iNOS immunopositivity was observed especially in astrocytes in white matter of cerebellum. The number of iNOS immunopositive astrocytes were significantly higher in chronic cases compared to acute cases of CDV infected dogs (p < 0.05). iNOS immunopositive gemistocytic astrocytes were also immunopositive for NT. iNOS and NT immunopositivity was not detected outside the region of the demyelinated areas. The iNOS immunopositive total cell number was found to be significantly higher in severely affected demyelination areas (p < 0.05). These data indicated that there was a relationship between degrees of the CDV associated with demyelination and the level of iNOS immunopositivity in the glial cells.