Ž . Brain Research 853 2000 156–161 www.elsevier.comrlocaterbres Short communication Immunocytochemical study on the distribution of nitrotyrosine in the brain of the transgenic mice expressing a human CurZn SOD mutation Choong Ik Cha a,e,f, ) , Yoon Hee Chung a , Chung-min Shin a , Dong Hoon Shin d , Yong Sik Kim b , Mark E. Gurney g , Kwang Woo Lee c a Department of Anatomy, Seoul National UniÕersity College of Medicine, Yongon-Dong 28, Chongno-Gu, Seoul 110-799, South Korea b Department of Pharmacology, Seoul National UniÕersity College of Medicine, Yongon-Dong 28, Chongno-Gu, Seoul, South Korea c Department of Neurology, Seoul National UniÕersity College of Medicine, Yongon-Dong 28, Chongno-Gu, Seoul, South Korea d Department of Anatomy, Dankook UniÕersity College of Medicine, Chonan, South Korea e Neuroscience Research Institute, Seoul, South Korea f Biomedical Research Center, KNIH, Seoul, South Korea g CNS Diseases Research Unit Pharmacia Upjohn, Inc., USA Accepted 2 November 1999 Abstract In the previous study, we reported increased NOS expression in the astrocytes in the spinal cord of the transgenic mice that are used as ALS animal model. In the present study, we performed immunocytochemical studies to investigate the changes of nitrotyrosine-immuno- reactivity in the brains of the transgenic mice, and demonstrated in vivo evidence of peroxynitrite-mediated oxidative damage in the pathogenesis of ALS. In the spinal cord of the transgenic mice, immunocytochemistry showed intensely stained nitrotyrosine-IR glial cells with the appearance of astrocytes, but no nitrotyrosine-IR glial cells were observed in the spinal cord of the control mice. In the transgenic mice, nitrotyrosine-IR neurons were observed in the hypoglossal nucleus, lateral reticular nucleus, medullary reticular formation and cerebellar nuclei. Interestingly, nitrotyrosine-IR neurons were observed in the hippocampal formation and septal area of the transgenic mice. In the hippocampus, nitrotyrosine-IR neurons in the CA1 region showed intense staining, and the immunoreactivity was localized mainly in the pyramidal cell layer. Recent studies have shown that antioxidants and selective neuronal NOS inhibitor increase survival in the SOD1 transgenic mouse model of FALS. It is possible that therapy with these agents may slow the neurodegenerative process in human ALS, perhaps through reduction of nitrotyrosine formation. q 2000 Elsevier Science B.V. All rights reserved. Keywords: Amyotrophic lateral sclerosis; Nitrotyrosine; Astrocyte; Transgenic mice; Immunocytochemistry Ž . Amyotrophic lateral sclerosis ALS is a fatal neurode- generative disorder that results in degeneration of motor neurons in the spinal cord, brainstem and cerebral cortex. The disease occurs in both sporadic and familial forms. Approximately 20% of familial cases of ALS are caused by single-site mutations in the CurZn superoxide dismu- Ž . tase SOD1 gene. SOD catalyzes the formation of hydro- gen peroxide through the dismutation of superoxide free radicals and is thus thought to play an important role in mitigating oxidative damage. Currently, there are three major hypotheses that address putative causal factors for the motor neuron degeneration Ž. that occurs in ALS: 1 the autoimmune hypothesis, which ) Corresponding author. Fax: q 82-2-745-9528; e-mail: yoonhee2@snu.ac.kr suggests that antibodies to N- and L-type calcium channels could cause or contribute to the sporadic form of the Ž. disorder; 2 the excitatory amino acid hypothesis, which suggests that excessive synaptic glutamate could contribute Ž. to motor neuron loss; 3 the oxidative stress hypothesis, based on mutation in the CurZn SOD. The pathogenesis of neuronal degeneration in both sporadic and familial ALS associated with mutations in SOD may involve oxida- tive stress. A leading candidate as a mediator of oxidative stress is peroxynitrite, which is formed by the reaction of Ž . superoxide with nitric oxide NO . The production of NO has been implicated in neuronal injury after ischemia, trauma, and numerous neurodegener- w x ative disorders, including ALS 3,5 . Excessive stimulation of glutamate receptors may mediate neuron death by the wx wx production of both NO 9 and superoxide 7 . NO and superoxide combine by a diffusion-limited reaction to form 0006-8993r00r$ - see front matter q 2000 Elsevier Science B.V. All rights reserved. Ž . PII: S0006-8993 99 02302-1