Rotenone induces oxidative stress and dopaminergic neuron damage in organotypic substantia nigra cultures.
Rotenone, a pesticide and complex I inhibitor, causes nigrostriatal degeneration similar to Parkinson disease pathology in a chronic, systemic, in vivo rodent model [M. Alam, W.J. Schmidt, Rotenone destroys dopaminergic neurons and induces parkinsonian symptoms in rats, Behav. Brain Res. 136 (2002) 317-324; R. Betarbet, T.B. Sherer, G. MacKenzie, M. Garcia-Osuna, A.V. Panov, J.T. Greenamyre, Chronic systemic pesticide exposure reproduces features of Parkinson's disease, Nat. Neurosci. 3 (2000) 1301-1306; S.M. Fleming, C. Zhu, P.O. Fernagut, A. Mehta, C.D. DiCarlo, R.L. Seaman, M.F. Chesselet, Behavioral and immunohistochemical effects of chronic intravenous and subcutaneous infusions of varying doses of rotenone, Exp. Neurol. 187 (2004) 418-429; T.B. Sherer, J.H. Kim, R. Betarbet, J.T. Greenamyre, Subcutaneous rotenone exposure causes highly selective dopaminergic degeneration and alpha-synuclein aggregation, Exp. Neurol. 179 (2003) 9-16.]. To better investigate the role of mitochondria and complex I inhibition in chronic, progressive neurodegenerative disease, we developed methods for long-term culture of rodent postnatal midbrain organotypic slices. Chronic complex I inhibition over weeks by low dose (10-50 nM) rotenone in this system lead to dose- and time-dependent destruction of substantia nigra pars compacta neuron processes, morphologic changes, some neuronal loss, and decreased tyrosine hydroxylase (TH) protein levels. Chronic complex I inhibition also caused oxidative damage to proteins, measured by protein carbonyl levels. This oxidative damage was blocked by the antioxidant alpha-tocopherol (vitamin E). At the same time, alpha-tocopherol also blocked rotenone-induced reductions in TH protein and TH immunohistochemical changes. Thus, oxidative damage is a primary mechanism of mitochondrial toxicity in intact dopaminergic neurons. The organotypic culture system allows close study of this and other interacting mechanisms over a prolonged time period in mature dopaminergic neurons with intact processes, surrounding glia, and synaptic connections.
Testa
Claudia M
CM
Center for Neurodegenerative Disease, Emory University, Atlanta, GA 30322, USA. ctesta@emory.edu
Sherer
Todd B
TB
Greenamyre
J Timothy
JT
D000818:Animals; D000831:Animals, Newborn; D000975:Antioxidants / Q000494:pharmacology; D015153:Blotting, Western / Q000379:methods; D002452:Cell Count / Q000379:methods; D004298:Dopamine / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D005786:Gene Expression Regulation / Q000187:drug effects; D007150:Immunohistochemistry / Q000379:methods; D009410:Nerve Degeneration / Q000139:chemically induced* / Q000517:prevention & control; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D009924:Organ Culture Techniques; D018384:Oxidative Stress / Q000187:drug effects* / Q000502:physiology; D051381:Rats; D012402:Rotenone / Q000009:adverse effects / Q000494:pharmacology*; D013378:Substantia Nigra / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D013997:Time Factors; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism; D024502:alpha-Tocopherol / Q000494:pharmacology
D000975:Antioxidants; D012402:Rotenone; D014446:Tyrosine 3-Monooxygenase; D024502:alpha-Tocopherol; D004298:Dopamine