MPTP-induced model of Parkinson's disease in cytochrome P450 2E1knockout mice
Introduction
The discovery that a contaminant found in an illicit substance of abuse, MPTP, was able to induce a severe syndrome of parkinsonism in humans gave a considerable impulse to research on the pathophysiology of Parkinson's disease (PD) (Davis et al., 1979, Langston et al., 1983). Elucidation of the biochemical steps leading to the MPTP-induced selective degeneration of the nigrostriatal dopamine (DA) pathway has provided new clues to DA neuron vulnerability (Kopin, 1992). Furthermore, marked species differences in MPTP toxicity have been described, and this differential sensitivity to the neurotoxin has provided further information about the genetic factors determining cell susceptibility to xenobiotic insults (Corsini et al., 2002). MPTP is metabolized to its toxic compound, MPP+, via an intermediate 1-methyl-4-phenyl-2,3-dihydropyridinium (MPDP+), and this oxidation is catalyzed by monoamine oxidase type B (Chiba et al., 1984, Markey et al., 1984). MPP+ accumulates in DA neurons through the high-affinity DA transporter (DAT) (Javitch and Snyder, 1984, Javitch et al., 1985). Inside the neuron, MPP+ is taken up by vesicular monoamine transporter 2 (VMAT2) and stored in synaptic vesicles (Kopin, 1992, Staal and Sonsalla, 2000). This storage is responsible for DA displacement and toxin inactivation (Del Zompo et al., 1991, Del Zompo et al., 1992, Stern-Bach et al., 1992). Alternatively, MPP+ is taken up by mitochondria, where it accumulates, directly inhibiting complex I (Nicklas et al., 1985, Ramsay and Singer, 1986, Sonsalla and Nicklas, 1992). Accordingly, MPP+ toxicity has been directly associated with failure of energy supplies in some in vitro models (Di Monte et al., 1986, Denton and Howard, 1987). Recently, other pieces of the puzzle have been pieced together, such as the role of nitric oxide and glial activation (Mohanakumar et al., 2002, Teismann et al., 2003). More recently, MPTP toxicity in mice has been linked to the presence within the brain of an enzyme of the P450 system, CYP2E1 (Vaglini et al., 2004).
By using CYP2E1 inhibitors, the authors were able to markedly enhance the typical MPTP lesion of the substantia nigra (SN) in mice and they suggested that this enzyme may have a detoxicant role related to the efflux transporter of the toxin.
CYP2E1 has been widely identified in the rat, as well as the human, brain (Hansson et al., 1990, Brzezinski et al., 1999, Joshi and Tyndale, 2006, Upadhya et al., 2000), and in neurons of the SN it has been associated with tyrosine hydroxylase (Riedl et al., 1996, Watts et al., 1998). It was shown that CYP2E1 in rat basal ganglia is induced by ethanol or nicotine administration (Anandatheerthavarada et al., 1993a, Anandatheerthavarada et al., 1993b, Sohda et al., 1993). Cyp2e1 mRNA expression was demonstrated in different brain areas including the SN (Farin and Omiecinski, 1993). A functional role of CYP2E1 in DA metabolism in the SN has been proposed recently (Nissbrandt et al., 2001, Niazi Shahabi et al., 2003, Niazi Shahabi et al., 2008).
In order to further understand the role of CYP2E1 in the MPTP mechanism of nigral lesions, in this study we investigated the response of Cyp2e1(−/−) mice to the toxic insult by MPTP in a typical paradigm of an acute challenge and we compared the results obtained in these animals with those observed in their wild-type counterparts. Here we report that Cyp2e1(−/−) mice not only fail to be hypersensitive to MPTP, but are even less sensitive to the typical nigral damage produced by the toxin.
Section snippets
Materials
MPTP hydrochloride, MPP+, phosphate buffer, paraformaldehyde, sucrose, heptane sulfonic acid, acetonitrile, triethylamine, and MPP+ were obtained from Sigma Aldrich (St. Louis, MO, USA) Anti-rabbit TH antibody was obtained from Chemicon ICN (Temecula, CA, USA). Anti-rabbit biotinylate IgG and avidin–biotin complex were from Vector Laboratories (Burlingame, CA, USA).
Knockout mice
Male Cyp2e1 knockout mice (129/SV-Cyp2e1tm1Gonz) (Cyp2e1(−/−) stock number: 002910) and their wild-type counterparts (129S1/SvImJ) (
Changes in striatal levels of DA and its metabolites
The effect of the acute administration of MPTP on the level of striatal DA and DA metabolites in the wild-type and Cyp2e1(−/−) mice is shown in Fig. 1. Seven days after treatment with MPTP, striatal DA content in wild-type mice was 42% of that in untreated controls (treated wild-type, 67.1 ± 2.5 ng/mg protein; controls, 116.1 ± 8.3 ng/mg protein). In knockout mice, administration of the single dose of MPTP was associated with a DA content 30% lower than that in untreated controls (treated knockout,
Discussion
CYP2E1 is an enzyme of the P450 family whose occurrence in high concentration is typical in the liver. Its presence in extrahepatic organs such as kidney and lung has been reported (Thomas et al., 1987, de Waziers et al., 1990). The enzyme has many endogenous and exogenous substrates, some of which are toxic and, generally, small non-polar molecules (Lieber, 1997). The many endogenous molecules include ketones (e.g. acetone) and fatty acids (e.g. arachidonic acid). The exogenous compounds
Acknowledgements
The authors would like to thank Ms Natalia Jaksic for her assistance in the preparation of the manuscript and Mrs Ornella Baroni for her valuable help in taking care of the animals.
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