Increased sensitivity to MPTP in human α-synuclein A30P transgenic mice
Introduction
Parkinson's disease (PD) is a progressive neurodegenerative disorder characterized by resting tremor, akinesia, rigidity and postural instability. In PD brains, there is loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc) leading to a dopamine (DA) depletion in the striatum. The etiology of Parkinson's disease remains unknown. Both environmental and genetic factors have been implicated in the pathogenesis of PD. The genetic hypothesis of PD has gained considerable interest during the last decade. Three distinct pathogenic mutations (A53T, A30P and E46K) in the α-synuclein gene, located on chromosome 4, have been identified in several families with autosomal dominant PD and dementia with Lewy bodies (DLB) [18], [26], [42]. The finding of these mutations quickly led to the discovery that α-synuclein accumulates in Lewy bodies (LBs), the pathological hallmark of PD, and Lewy neurites in both familial and sporadic PD and DLB [13], [35], [41], as well as in glial inclusions in multi system atrophy (MSA) [40]. These findings suggest a pathological role for α-synuclein aggregation in these disorders [9]. The mechanisms by which α-synuclein forms neuronal and glial inclusions and becomes associated with neuronal degeneration, or by which α-synuclein mutations result in hereditary forms of PD, remain unknown.
Several transgenic mouse models expressing wild type or mutant human α-synuclein have been reported [17], [22], [24], [38]. All of them point to the fact that high brain expression levels of wild type or mutant α-synuclein in the mouse brain consistently results in aberrant localization of the transgenic protein in the neuronal cytoplasm. The phenotypic consequences of abnormal synuclein cellular localization vary considerably among different transgenic lines and include biochemical evidence of altered α-synuclein processing, pathological changes of striatal denervation, astrocytosis and microgliosis, peripheral neuropathy, and clinical deficits ranging from no deficits to lethal motor dysfunction.
We have recently published that high expression of α-synuclein A30P in the Tg5093 line results in a progressive motor disorder with rigidity, dystonia, gait impairment, and tremor [10]. Histological analysis of this line showed aberrant expression of the protein in cell soma and progressive central nervous system (CNS) gliosis, but no specific deterioration of the nigrostriatal dopaminergic system or overt lesions resembling LBs.
In addition to genetic factors, environmental factors have long been suspected to contribute to the pathogenesis of PD after the report of a parkinsonian syndrome in young adults that were intoxicated by the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a potent complex I inhibitor, which selectively destroys nigrostriatal DA neurons [20]. Several chemical products, including paraquat, diquat and rotenone, used in herbicides and pesticides, are similar structurally to MPTP. However, to date it has not been possible to identify any causative environmental chemical agent in the etiology of PD despite intensive research. Some recent studies have brought renewed interest in the possible role of environmental toxins in PD. A30P α-synuclein expression in PC12 cell lines results in decreased proteasome activity and increased sensitivity to apoptotic cell death after treatment with the proteasome inhibitor lactacystin [37]. The herbicide paraquat enhances α-synuclein fibril formation in mice [21], and chronic administration of rotenone leads to intraneuronal aggregates of α-synuclein in rats [1], [34]. Chronic treatment with MPTP upregulates α-synuclein expression and induces selective redistribution, aggregation and nitration of this protein in the SN in vivo [27], [39]. Moreover, α-synuclein null mice show resistance to MPTP-induced degeneration of nigrostriatal DA neurons when compared to littermates of the same genetic background [5], [30]. Recently, Kuhn et al. observed that α-synuclein was the only gene exclusively up-regulated in MPTP-treated animals displaying cell death [19].
Despite the above observations, whether or not mutant α-synuclein enhances the vulnerability of dopaminergic neurons to environmental toxins in vivo such as MPTP or rotenone remains controversial. While Richfield et al. observed a major effect of α-synuclein on MPTP toxicity [29], two other studies did not detect increased sensitivity to MPTP in transgenic or viral vector-transduced mice overexpressing human A30P and A53T α-synuclein, respectively [6], [28].
We have further studied the possible interaction of genetic and environmental factors in PD by testing whether Tg5093 positive mice are more sensitive than their age-matched littermate controls to MPTP or rotenone, and whether these secondary toxic insults may induce additional phenotypic traits resembling those of human Lewy body disorders in the Tg5093 line.
Section snippets
Transgenic mice
The generation of Tg5093 mice has been previously described [10]. In brief, the cDNA encoding human α-synuclein containing the A30P mutation was expressed in C57B/6jxSJL F3 hybrid mice using a hamster prion protein (PrP) cosmid vector, in which the PrP open reading frame (ORF) was replaced with the human α-synuclein ORF. This vector has been previously shown to drive transgene product expression throughout neurons in the brain, including the substantia nigra (SN) and other vulnerable neuronal
Mortality rates
Mortality rates in Tg+/− MPTP/rotenone intoxicated mice at age 3–4 months were three- to four-fold higher than in Tg−/− MPTP/rotenone intoxicated mice (17%/13% versus 6%/3%, respectively). Similarly, mortality rates in Tg+/− MPTP/rotenone intoxicated mice at 6–8 months of age were two- to three-fold higher than in Tg−/− MPTP/rotenone intoxicated mice (13%/15% versus 5%/6%, respectively). Interestingly, all deaths in MPTP treated animals occurred in males while no gender differences on mortality
Discussion
The main objective of this study was to investigate possible interactions of genetic and environmental factors in PD by testing whether Tg5093 transgenic mice overexpressing human α-synuclein containing the A30P mutation are more sensitive than age-matched littermate controls to neurotoxins like MPTP or rotenone, and whether these secondary toxic insults may result in additional phenotypic traits resembling those of human Lewy body disorders.
The Tg5093 mouse line expressing high levels of
Acknowledgements
We thank Dr. Marta García Granero for the statistical support and Drs. Michael Irizarry and Bradley Hyman for their useful comments and suggestions.
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This project was funded in part by EC grant DIADEM QRLT-2000-026362, Ayuda Serono Fundacion Salud 2000, grant 361/01 Fundacion Echebano and UTE CIMA project.