The parkinsonian neurotoxin rotenone activates calpain and caspase-3 leading to motoneuron degeneration in spinal cord of Lewis rats

Exposure to environmental toxins increases the risk of neurodegenerative diseases including Parkinson's disease (PD). Rotenone is a neurotoxin that has been used to induce experimental Parkinsonism in rats. We used the rotenone model of experimental Parkinsonism to explore a novel aspect of extra-nigral degeneration, the neurodegeneration of spinal cord (SC), in PD. Rotenone administration to male Lewis rats caused significant neuronal cell death in cervical and lumbar SC as compared with control animals. Dying neurons were motoneurons as identified by double immunofluorescent labeling for terminal deoxynucleotidyl transferase, recombinant-mediated dUTP nick-end labeling-positive (TUNEL(+)) cells and choline acetyltransferase (ChAT)-immunoreactivity. Neuronal death was accompanied by abundant astrogliosis and microgliosis as evidenced from glial fibrillary acidic protein (GFAP)-immunoreactivity and OX-42-immunoreactivity, respectively, implicating an inflammatory component during neurodegeneration in SC. However, the integrity of the white matter in SC was not affected by rotenone administration as evidenced from the non co-localization of any TUNEL(+) cells with GFAP-immunoreactivity and myelin basic protein (MBP)-immunoreactivity, the selective markers for astrocytes and oligodendrocytes, respectively. Increased activities of 76 kD active m-calpain and 17/19 kD active caspase-3 further demonstrated involvement of these enzymes in cell death in SC. The finding of ChAT(+) cell death also suggested degeneration of SC motoneurons in rotenone-induced experimental Parkinsonism. Thus, this is the first report of its kind in which the selective vulnerability of a putative parkinsonian target outside of nigrostriatal system has been tested using an environmental toxin to understand the pathophysiology of PD. Moreover, rotenone-induced degeneration of SC motoneuron in this model of experimental Parkinsonism progressed with upregulation of calpain and caspase-3.

[1]  P. Armati,et al.  Capsaicin-induced depolarisation of mitochondria in dorsal root ganglion neurons is enhanced by vanilloid receptors , 2001, Neuroscience.

[2]  Syed F. Ali,et al.  Calpain Upregulation in Spinal Cords of Mice with 1‐Methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP)‐induced Parkinson's Disease , 2000, Annals of the New York Academy of Sciences.

[3]  Syed F. Ali,et al.  Calpain Activation in Apoptosis of Motoneurons in Cell Culture Models of Experimental Parkinsonism , 2006, Annals of the New York Academy of Sciences.

[4]  H. Braak,et al.  α-Synuclein immunopositive Parkinson's disease-related inclusion bodies in lower brain stem nuclei , 2001, Acta Neuropathologica.

[5]  A. Vercesi,et al.  Ca2+‐induced oxidative stress in brain mitochondria treated with the respiratory chain inhibitor rotenone , 2003, FEBS letters.

[6]  Michel Goedert,et al.  Alpha-synuclein and neurodegenerative diseases , 2001, Nature Reviews Neuroscience.

[7]  D. Price,et al.  Parkinson's Disease α-Synuclein Transgenic Mice Develop Neuronal Mitochondrial Degeneration and Cell Death , 2006, The Journal of Neuroscience.

[8]  Hansjürgen Bratzke,et al.  Staging of the intracerebral inclusion body pathology associated with idiopathic Parkinson's disease (preclinical and clinical stages) , 2002, Journal of Neurology.

[9]  P. Mcgeer,et al.  Inflammation and neurodegeneration in Parkinson's disease. , 2004, Parkinsonism & related disorders.

[10]  Todd B. Sherer,et al.  Chronic systemic pesticide exposure reproduces features of Parkinson's disease , 2000, Nature Neuroscience.

[11]  K. Heidenreich,et al.  The pesticide rotenone induces caspase‐3‐mediated apoptosis in ventral mesencephalic dopaminergic neurons , 2003, Journal of neurochemistry.

[12]  A. H. V. Schapira,et al.  MITOCHONDRIAL COMPLEX I DEFICIENCY IN PARKINSON'S DISEASE , 1989, The Lancet.

[13]  E. Wolters,et al.  Enhanced sensitivity of dopaminergic neurons to rotenone-induced toxicity with aging. , 2006, Parkinsonism & related disorders.

[14]  A. Członkowska,et al.  The Inflammatory Reaction Following 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine Intoxication in Mouse , 1999, Experimental Neurology.

[15]  W. Dauer,et al.  Parkinson's Disease Mechanisms and Models , 2003, Neuron.

[16]  S. Khuder,et al.  A meta-analysis of Parkinson's disease and exposure to pesticides. , 2000, Neurotoxicology.

[17]  D. Dickson,et al.  Alpha‐synuclein immunohistochemistry in two cases of co‐occurring idiopathic Parkinson's disease and motor neuron disease , 2005, Movement disorders : official journal of the Movement Disorder Society.

[18]  Todd B. Sherer,et al.  Mechanism of Toxicity in Rotenone Models of Parkinson's Disease , 2003, The Journal of Neuroscience.

[19]  N. Banik,et al.  Calpain and its involvement in the pathophysiology of CNS injuries and diseases: therapeutic potential of calpain inhibitors for prevention of neurodegeneration. , 2003, Current drug targets. CNS and neurological disorders.

[20]  A. Probst,et al.  α‐Synuclein pathology of the spinal and peripheral autonomic nervous system in neurologically unimpaired elderly subjects , 2006, Neuropathology and applied neurobiology.

[21]  S. Imam,et al.  Calpain Upregulation and Neuron Death in Spinal Cord of MPTP‐Induced Parkinsonism in Mice , 2002, Annals of the New York Academy of Sciences.

[22]  E. Hirsch,et al.  Increased m-calpain expression in the mesencephalon of patients with parkinson's disease but not in other neurodegenerative disorders involving the mesencephalon: a role in nerve cell death? , 1996, Neuroscience.

[23]  N. Yanagisawa.  Natural history of Parkinson's disease: From dopamine to multiple system involvement , 2006 .

[24]  E. Hirsch,et al.  The Role of Glial Reaction and Inflammation in Parkinson's Disease , 2003, Annals of the New York Academy of Sciences.

[25]  H. Shibasaki,et al.  Effects of mitochondrial dysfunction on glutamate receptor-mediated neurotoxicity in cultured rat spinal motor neurons , 2004, Brain Research.

[26]  Vladimir N. Uversky,et al.  Neurotoxicant-induced animal models of Parkinson’s disease: understanding the role of rotenone, maneb and paraquat in neurodegeneration , 2004, Cell and Tissue Research.

[27]  M. Beal.  Mitochondria, Oxidative Damage, and Inflammation in Parkinson's Disease , 2003, Annals of the New York Academy of Sciences.

[28]  Hua-qing Liu,et al.  A delivery strategy for rotenone microspheres in an animal model of Parkinson's disease. , 2006, Biomaterials.

[29]  Wenhua Liu,et al.  Selective Vulnerability of Dopaminergic Neurons to Microtubule Depolymerization* , 2005, Journal of Biological Chemistry.

[30]  E. Hogan,et al.  Combined TUNEL and double immunofluorescent labeling for detection of apoptotic mononuclear phagocytes in autoimmune demyelinating disease. , 2000, Brain research. Brain research protocols.

[31]  J. Paul Robinson,et al.  Mitochondrial Complex I Inhibitor Rotenone Induces Apoptosis through Enhancing Mitochondrial Reactive Oxygen Species Production* , 2003, The Journal of Biological Chemistry.

[32]  Hitoshi Takahashi,et al.  The intermediolateral nucleus and Clarke’s column in Parkinson’s disease , 1997, Acta Neuropathologica.

[33]  E. Yoshikawa,et al.  Microglial activation and dopamine terminal loss in early Parkinson's disease , 2005, Annals of neurology.

[34]  Kiowa S. Bower,et al.  Synergistic effects of pesticides and metals on the fibrillation of alpha-synuclein: implications for Parkinson's disease. , 2002, Neurotoxicology.

[35]  B. Liu,et al.  Distinct Role for Microglia in Rotenone-Induced Degeneration of Dopaminergic Neurons , 2002, The Journal of Neuroscience.

[36]  David S. Park,et al.  Inhibition of Calpains Prevents Neuronal and Behavioral Deficits in an MPTP Mouse Model of Parkinson's Disease , 2003, The Journal of Neuroscience.

[37]  M. Duchen,et al.  Mitochondria, Ca2+ and neurodegenerative disease. , 2002, European journal of pharmacology.

[38]  D. Higgins,et al.  In Vivo Labeling of Mitochondrial Complex I (NADH:UbiquinoneOxidoreductase) in Rat Brain Using [3H]Dihydrorotenone , 2000, Journal of neurochemistry.

[39]  M. Oda,et al.  Diminution of dopaminergic neurons in the substantia nigra of sporadic amyotrophic lateral sclerosis , 1993, Neuropathology and applied neurobiology.

[40]  W. Schmidt,et al.  A rat model of Parkinsonism shows depletion of dopamine in the retina , 2007, Neurochemistry International.

[41]  N. Banik,et al.  The localization of mcalpain in myelin: immunocytochemical evidence in different areas of rat brain and nerves , 1995, Brain Research.

[42]  P. Carvey,et al.  Progressive Dopamine Neuron Loss in Parkinson's Disease: The Multiple Hit Hypothesis , 2006, Cell transplantation.

[43]  O. Isacson,et al.  Neuroinflammation of the nigrostriatal pathway during progressive 6‐OHDA dopamine degeneration in rats monitored by immunohistochemistry and PET imaging , 2002, The European journal of neuroscience.

[44]  C. Olanow,et al.  Inhibition of proteasome activity sensitizes dopamine neurons to protein alterations and oxidative stress , 2004, Journal of Neural Transmission.

[45]  Y. Hiraku,et al.  Mechanism for generation of hydrogen peroxide and change of mitochondrial membrane potential during rotenone-induced apoptosis. , 2003, Life sciences.

[46]  M. Delong,et al.  Pathophysiology of parkinsonian motor abnormalities. , 1993, Advances in neurology.

[47]  B. Boeve,et al.  α-Synuclein pathology in the spinal cords of neurologically asymptomatic aged individuals , 2006, Neurology.

[48]  M. L. Schmidt,et al.  α-Synuclein in Lewy bodies , 1997, Nature.

[49]  J. H. Lucas,et al.  Inosine and guanosine preserve neuronal and glial cell viability in mouse spinal cord cultures during chemical hypoxia , 1999, Brain Research.

[50]  W. Schmidt,et al.  Rotenone destroys dopaminergic neurons and induces parkinsonian symptoms in rats , 2002, Behavioural Brain Research.

[51]  E. Hogan,et al.  Purification of a calcium-activated neutral proteinase from bovine brain , 1983, Neurochemical Research.

[52]  Todd B. Sherer,et al.  An In Vitro Model of Parkinson's Disease: Linking Mitochondrial Impairment to Altered α-Synuclein Metabolism and Oxidative Damage , 2002, The Journal of Neuroscience.

[53]  M. Duchen,et al.  Interplay between mitochondria and cellular calcium signalling , 2004, Molecular and Cellular Biochemistry.

[54]  Wei Duan,et al.  Early induction of calpains in rotenone-mediated neuronal apoptosis , 2006, Neuroscience Letters.

[55]  G. Meredith,et al.  Behavioral models of Parkinson's disease in rodents: A new look at an old problem , 2006, Movement disorders : official journal of the Movement Disorder Society.

[56]  Y. Kagawa,et al.  Deficiencies in complex I subunits of the respiratory chain in Parkinson's disease. , 1989, Biochemical and biophysical research communications.

[57]  M. Lupi,et al.  Low levels of ALS-linked Cu/Zn superoxide dismutase increase the production of reactive oxygen species and cause mitochondrial damage and death in motor neuron-like cells , 2005, Journal of the Neurological Sciences.

[58]  E. Sribnick,et al.  Immunofluorescent labeling of increased calpain expression and neuronal death in the spinal cord of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated mice , 2004, Brain Research.

[59]  Diana Brahams,et al.  Medicine and the Law , 1983, The Lancet.

[60]  I. Miyazaki,et al.  Rotenone induces aggregation of γ-tubulin protein and subsequent disorganization of the centrosome: Relevance to formation of inclusion bodies and neurodegeneration , 2005, Neuroscience.

[61]  H. Braak,et al.  Staging of brain pathology related to sporadic Parkinson’s disease , 2003, Neurobiology of Aging.

[62]  D. Allen,et al.  Caspase and calpain function in cell death: bridging the gap between apoptosis and necrosis , 2005, Annals of clinical biochemistry.

[63]  M. Lupi,et al.  Neurodegeneration induced by complex I inhibition in a cellular model of familial amyotrophic lateral sclerosis , 2006, Brain Research Bulletin.

[64]  O. Andreassen,et al.  Transgenic ALS Mice Show Increased Vulnerability to the Mitochondrial Toxins MPTP and 3-Nitropropionic Acid , 2001, Experimental Neurology.

[65]  Y. Kalkonde,et al.  Chemokines in the MPTP model of Parkinson’s disease: Absence of CCL2 and its receptor CCR2 does not protect against striatal neurodegeneration , 2007, Brain Research.

[66]  James Lowe,et al.  Parkinsonism in motor neuron disease: case report and literature review , 2004, Acta Neuropathologica.

[67]  T. Peng,et al.  Mitochondrial dysfunction in Parkinson's disease. , 1999, Biochemical Society symposium.

[68]  M. Gurney,et al.  Midbrain dopaminergic neuronal degeneration in a transgenic mouse model of familial amyotrophic lateral sclerosis , 1997, Annals of neurology.

[69]  B. Crain,et al.  Degeneration of the substantia nigra in familial amyotrophic lateral sclerosis. , 1991, Clinical neuropathology.

[70]  M. Esposti.  Inhibitors of NADH-ubiquinone reductase: an overview. , 1998 .

[71]  M. Weinberg,et al.  Activation of multiple molecular mechanisms for apoptosis in human malignant glioblastoma T98G and U87MG cells treated with sulforaphane , 2006, Neuroscience.

[72]  C. C. Johnson,et al.  The risk of Parkinson's disease with exposure to pesticides, farming, well water, and rural living , 1998, Neurology.

[73]  G. Núñez,et al.  Caspases: the proteases of the apoptotic pathway , 1998, Oncogene.

[74]  E. Sribnick,et al.  Estrogen prevents glutamate-induced apoptosis in C6 glioma cells by a receptor-mediated mechanism , 2006, Neuroscience.

[75]  G. Fiskum,et al.  Mitochondrial Mechanisms of Neural Cell Death and Neuroprotective Interventions in Parkinson's Disease , 2003, Annals of the New York Academy of Sciences.

[76]  F. Gonzalez-Lima,et al.  Neurodegeneration Produced by Rotenone in the Mouse Retina: A Potential Model to Investigate Environmental Pesticide Contributions to Neurodegenerative Diseases , 2006, Journal of toxicology and environmental health. Part A.