Dopamine turnover is upregulated in the caudate/putamen of asymptomatic MPTP-treated rhesus monkeys

In Parkinson's disease (PD) and experimental parkinsonism, losses of up to 60% and 80%, respectively, of dopaminergic neurons in substantia nigra, and dopamine (DA) in striatum remain asymptomatic. Several mechanisms have been suggested for this functional compensation, the DA-mediated being the most established one. Since this mechanism was recently challenged by striatal DA analysis in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated monkeys, we present data on several DAergic parameters in three groups of rhesus monkeys: MPTP-treated asymptomatic animals; symptomatic MPTP-treated animals with stable parkinsonism; and untreated sex and age matched controls. We determined ratios of striatal and nigral 3,4-dihydroxyphenyl acetic acid (DOPAC) to DA levels and tyrosine hydroxylase (TH) enzyme activity to DA levels, in addition to the commonly used homovanillic acid (HVA)/DA ratios which, as such, might be less reliable under the conditions of partial denervation. We found that in the asymptomatic MPTP monkeys the DOPAC/DA ratios in putamen and caudate nucleus were shifted with high statistical significance 1.9-5.8-fold, as compared to controls, the shifting of the ratios being in the same range as the 2.6-5.4-fold shifts in the symptomatic animals. Also TH/DA ratios were significantly increased in both, the asymptomatic and the symptomatic MPTP-treated monkeys, with shifts in the putamen and caudate nucleus of 3- and 2.7-7.0-fold, respectively. In the substantia nigra, DOPAC levels and TH activity were strongly decreased after MPTP (-77 to -97%), but the ratios DOPAC/DA and TH/DA were not changed in this brain region. Collectively, our findings support the concept of DAergic compensation of the progressive striatal DA loss in the presymptomatic stages of the parkinsonian disease process.     

Binding and uptake of MPTP by preparations of human and animal brain

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is known to cause parkinsonism in man and animals, producing acute behavioral effects within minutes of administration. This syndrome has been attributed to specific effects on dopaminergic systems. MPTP blocked the binding of haloperidol to membranes from rat and human brain (IC₅₀ = 2.5 μM), but it did not block the binding of flupenthixol to these membranes. These results indicate that MPTP is a ligand for D-2 dopamine receptors but not for D-1 dopamine receptors. Synaptosomes from rat, mouse or guinea-pig corpus striatum or from monkey caudate nucleus exhibited little ability to take up MPTP from the incubation medium. The synaptosomes took up at least 20–50 times more dopamine than MPTP. These results indicate that MPTP could cause acute effects by binding to dopamine receptors and that the specific toxicity MPTP exerts for dopaminergic neuron is not primarily based on the specific uptake of MPTP into these neurons.     

Enhanced susceptibility to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine neurotoxicity in high-fat diet-induced obesity

Currently, obesity is considered a systemic inflammation; however, the effects of obesity on the vulnerability of dopaminergic neurons to oxidative stress are not fully defined. We evaluated the effects of high-fat diet-induced obesity (HF DIO) on neurotoxicity in mice treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Eight weeks after a HF or matched normal diet, a severe decrease in the levels of striatal dopamine and of nigral microtubule-associated protein 2, manganese superoxide dismutase, and tyrosine hydroxylase was observed in obese mice treated with subtoxic doses of MPTP (20 mg/kg) compared with the matched lean group. In addition, the levels of nitrate/nitrite and thiobarbituric acid-malondialdehyde adducts in the substantia nigra of obese mice were reciprocally elevated or suppressed by MPTP. Interestingly, striatal nNOS phosphorylation and dopamine turnover were elevated in obese mice after MPTP treatment, but were not observed in lean mice. The nitrotyrosine immunoreactivity for evaluation of nigral nitrogenous stress in obese mice with MPTP was higher than that in matched lean mice. At higher doses of MPTP (60 mg/kg), the mortality was higher in obese mice than in lean mice. These results suggest that DIO may increase the vulnerability of dopaminergic neurons to MPTP via increased levels of reactive oxygen and nitrogen species, and the role of nNOS phosphorylation in the MPTP toxicities and dopamine homeostasis should be further evaluated.     

25-Hydroxyvitamin D depletion does not exacerbate MPTP-induced dopamine neuron damage in mice

Recent clinical evidence supports a link between 25-hydroxyvitamin D insufficiency (serum 25-hydroxyvitamin D [25(OH)D] levels <30 ng/mL) and Parkinson's disease. To investigate the effect of 25(OH)D depletion on neuronal susceptibility to toxic insult, we induced a state of 25(OH)D deficiency in mice and then challenged them with the dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). We found there was no significant difference between control and 25(OH)D-deficient animals in striatal dopamine levels or dopamine transporter and tyrosine hydroxylase expression after lesioning with MPTP. Additionally, we found no difference in tyrosine hydroxylase expression in the substantia nigra pars compacta. Our data suggest that reducing 25(OH)D serum levels in mice has no effect on the vulnerability of nigral dopaminergic neurons in vivo in this model system of parkinsonism.     

Evaluation of the capability of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine and pyridine derivatives to evoke parkinsonism

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) produce an irreversible parkinsonian-like syndrome in humans, monkeys and mice C57BL/6. Experimental parkinsonism produced by MPTP on mice C57BL/6 were studied with the aim of working up the method for testing MPTP-like substances. It has been shown that intraperitoneal administration the maximal tolerated doses of MPTP cause significant decrease (by 40-60%) of dopamine content on the mice brain. Number of injections did not influence the results. The similar administration of 4-phenyl-pyridyl and 4,4'-dipyridyl derivates, including known herbicides paraquat and cyperquat, produce neither decrease of dopamine content in the brain, nor the development of parkinsonian-like behavioral syndrome.     

MK-801 Prevents 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine-Induced Parkinsonism in Primates

In cynomologus monkeys, systemic administration of MK-801, a noncompetitive antagonist for the N-methyl-D-aspartate receptor, prevented the development of the parkinsonian syndrome induced by the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). MK-801 also attenuated dopamine depletion in the caudate and putamen and protected dopaminergic neurons in the substantia nigra from the degeneration induced by the neurotoxin. Nevertheless, 7 days after MPTP administration in the caudate and putamen of monkeys also receiving MK-801, the levels of toxic 1-methyl-4-phenylpyridinium were even higher than those measured in monkeys receiving MPTP alone. This indicates that the protective action of MK-801 is not related to MPTP metabolism and strongly suggests that, in primates, the excitatory amino acids could play a crucial role in the mechanism of the selective neuronal death induced by MPTP.     

Neurochemical and Neurogenetic Correlates of Parkinson's Disease

Abstract: We discuss neurochemical and neurogenetic correlates of Parkinson's disease (PD) based on the recent progress in the study of its etiology and pathogenesis. Nigral degeneration with the presence of Lewy bodies in the remaining neurons is the pathologic hallmark of PD, and the resultant loss of striatal dopamine is responsible for most of the clinical manifestations. Although the primary cause is still unknown, mitochondrial respiratory failure and oxidative stress appear to be two major contributors to the nigral cell death. Many endogenous and exogenous compounds with structural similarity to MPTP have been postulated as potential neurotoxins inducing nigral cell death in PD, but there is little evidence of accumulation of such compounds in the nigra. Genetic influence has increasingly been recognized as an important risk factor for PD. In this respect, genetic linkage analysis and molecular cloning of the disease genes in familial parkinsonism are of utmost importance today. Recently, the disease gene for one of the autosomal dominant forms of familial PD was identified, and we cloned the gene for an autosomal recessive type of familial parkinsonism that had been mapped to the long arm of chromosome 6 by our group. Information obtained on familial parkinsonism will contribute to the studies on sporadic PD as well.     

Pramipexole protects against MPTP toxicity in non-human primates

The neurotoxin MPTP induces nigral dopaminergic cell death in primates and produces a partial model of Parkinson's disease (PD). Pramipexole is a D2/D3 dopamine receptor agonist used in the symptomatic treatment of PD, and which also protects neuronal cells against dopaminergic toxins in vitro. We now demonstrate that pramipexole partially prevents MPTP toxicity in vivo in a primate species. Common marmosets were repeatedly treated with pramipexole either before, coincidentally with, or after low-dose MPTP treatment designed to induce a partial lesion of the substantia nigra. Animals pretreated with pramipexole had a significantly greater number of surviving tyrosine hydroxylase (TH) positive neurones in the pars compacta of the substantia nigra. Pramipexole pretreatment also prevented degeneration of striatal dopamine terminals. Treatment with pramipexole concurrently with MPTP or following MPTP did not prevent TH-positive cell loss. Pramipexole pretreatment appears to induce adaptive changes that protect against dopaminergic cell loss in primates.     

Release of dopamine in the ventral tegmental area of rat

In this preliminary report we showed that 3,4-dihydroxyphenylacetic acid (DOPAC), the major metabolite of dopamine (DA), is present in the ventral tegmental area. This finding indicates that in the ventral tegmental area, which contains the cell bodies of dopaminergic neurons of the mesocortical and mesolimbic DA systems, DA may be released by a mechanism similar to that operating in the nerve endings. However, haloperidol, which increases DOPAC levels in the substantia nigra, failed to do so in the ventral tegmental area. The results support the contention that DA neurons in the ventral tegmental area have distinctive features from nigral DA neurones.     

Cerebral Metabolic Effects of Monoamine Oxidase Inhibition in Normal and 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine Acutely Treated Monkeys

The neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induces dopaminergic cell death in the substantia nigra pars compacta (SNpc) and clinical parkinsonism in humans and experimental animals. Pretreatment with monoamine oxidase inhibitors prevents this cell death and associated parkinsonism by blocking the oxidation of MPTP to a toxic intermediate. The 2-deoxyglucose method was used to study the acute effects of MPTP in the monkey brain and the effects of monoamine oxidase inhibition on local cerebral glucose utilization in both normal and MPTP-treated monkeys. MPTP administration alone caused a major increase in glucose utilization in the SNpc and smaller increases in some subnuclei within the ventral tegmental area in which eventual dopaminergic cell loss also occurs. Pretreatment with pargyline abolished these metabolic increases, a finding suggesting both that the oxidized product of MPTP generates the metabolic increases and that the increased glucose consumption may contribute to cell toxicity. On the other hand, in most cortical, thalamic, striatal, brainstem, and cerebellar areas MPTP alone caused reductions in glucose utilization, and pargyline failed to prevent these effects. Pargyline alone depressed metabolism in the locus coeruleus and a few other monoaminergic structures.     

Celastrol protects against MPTP- and 3-nitropropionic acid-induced neurotoxicity

Oxidative stress and inflammation are implicated in neurodegenerative diseases including Parkinson's disease (PD) and Huntington's disease (HD). Celastrol is a potent anti-inflammatory and antioxidant compound extracted from a perennial creeping plant belonging to the Celastraceae family. Celastrol is known to prevent the production of proinflammatory cytokines, inducible nitric oxide synthase and lipid peroxidation. Mice were treated with celastrol before and after injections of MPTP, a dopaminergic neurotoxin, which produces a model of PD. A 48% loss of dopaminergic neurons induced by MPTP in the substantia nigra pars compacta was significantly attenuated by celastrol treatment. Moreover, celastrol treatment significantly reduced the depletion in dopamine concentration induced by MPTP. Similarly, celastrol significantly decreased the striatal lesion volume induced by 3-nitropropionic acid, a neurotoxin used to model HD in rats. Celastrol induced heat shock protein 70 within dopaminergic neurons and decreased tumor necrosis factor-alpha and nuclear factor kappa B immunostainings as well as astrogliosis. Celastrol is therefore a promising neuroprotective agent for the treatment of PD and HD.     

A single intramuscular injection of rAAV‐mediated mutant erythropoietin protects against MPTP‐induced parkinsonism

Erythropoietin (Epo) is neuroprotective in a number of preparations, but can lead to unacceptably high and even lethal hematocrit levels. Recent reports show that modified Epo variants confer neuroprotection in models of glaucoma and retinal degeneration without raising hematocrit. In this study, neuroprotective effects of two Epo variants (EpoR76E and EpoS71E) were assessed in a model of Parkinson's disease. The constructs were packaged in recombinant adeno‐associated viral (rAAV) vectors and injected intramuscularly. After 3 weeks, mice received five daily injections of 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP) and were killed 5 weeks later. The MPTP‐lesioned mice pretreated with rAAV.eGFP (negative control) exhibited a 7‐ to 9‐Hz tremor and slower latencies to move on a grid test (akinesia). Both of these symptomatic features were absent in mice pretreated with either modified Epo construct. The rAAV.eGFP‐treated mice lesioned with MPTP exhibited a 41% reduction in tyrosine hydroxylase (TH)‐positive neurons in the substantia nigra. The rAAV.EpoS71E construct did not protect nigral neurons, but neuronal loss in mice pretreated with rAAV.EpoR76E was only half that of rAAV.eGFP controls. Although dopamine levels were normal in all groups, 3,4‐dihydroxyphenylacetic acid (DOPAC) was significantly reduced only in MPTP‐lesioned mice pretreated with rAAV.eGFP, indicating reduced dopamine turnover. Analysis of TH‐positive fibers in the striatum showed normalized density in MPTP‐lesioned mice pretreated with rAAV.EpoS71E, suggesting that enhanced sprouting induced by EpoS71E may have been responsible for normal behavior and dopaminergic tone in these mice. These results show that systemically administered rAAV‐generated non‐erythropoietic Epo may protect against MPTP‐induced parkinsonism by a combination of neuroprotection and enhanced axonal sprouting.     

The TrkB-Positive Dopaminergic Neurons are Less Sensitive to MPTP Insult in the Substantia Nigra of Adult C57/BL Mice

Tyrosine kinase receptors TrkB and TrkC mediate neuroprotective effects of the brain-derived neurotrophic factor (BDNF) and neurotrophins in the dopaminergic nigro-striatal system, but it is obscure about their responses or expression changes in the injured substantia nigra under Parkinson’s disease. In present study, immunofluorescence, Fluoro-Jade staining and laser scanning confocal microscopy were applied to investigate distribution and changes of TrkB and TrkC in the dopamine neurons of the substantia nigra by comparison of control and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model. It revealed that TrkB and TrkC-immunoreactivities were substantially localized in cytoplasm and cell membrane of the substantia nigra neurons of control adults. While neurons double-labeled with tyrosine hydroxylase (TH)/TrkB, or TH/TrkC were distributed in a large numbers in the substantia nigra of controls, they apparently went down at 36.2–65.7% of normal level, respectively following MPTP insult. In MPTP model, cell apoptosis or degeneration of nigral neurons were confirmed by caspase-3 and Fluoro-Jade staining. More interestingly, TH/TrkB-positive neurons survived more in cell numbers in comparison with that of TH/TrkC-positive ones in the MPTP model. This study has indicated that TrkB-containing dopamine neurons are less sensitive in the substantia nigra of MPTP mouse model, suggesting that specific organization of Trks may be involved in neuronal vulnerability to MPTP insult, and BDNF-TrkB signaling may play more important role in protecting dopamine neurons and exhibit therapeutic potential for Parkinson’s disease.     

Chemical neurotransmission in the parkinsonian brain

In Parkinson's disease the progressive loss of nigrostriatal dopamine neurons leads to striatal dopamine deficiency and correlates with the severity of parkinsonian disability. The findings concerning dopamine receptors both in vitro and in vivo are not consistent, possibly reflecting differences in patient populations, but the presynaptic defect in dopaminergic neurotransmission is greater than that seen in postsynaptic receptor binding studies. The cholinergic neurons in the extrapyramidal nuclei are relatively well preserved, but subcortico-cortical and -hippocampal cholinergic neurons degenerate in relation to the degree of dementia. The decreased GABA receptor binding in the parkinsonian substantia nigra possibly reflects the loss of nigral dopamine neurons, since nigral GABA receptors are located on these neurons. Of the various neuropeptides, the concentration of met- and leu-enkephalin seems to be reduced in the striatum. In the substantia nigra the concentration of substance P decreases, together with the met-enkephalin and cholecystokinin levels. The concentration of somatostatin decreases in the frontal cortex and hippocampus of demented patients. With the exception of the association between cortical somatostatin deficiency and intellectual deterioration, the role of the neuropeptides in the pathophysiology and clinical features of Parkinson's disease are not yet fully understood.     

Distribution of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine in experimental animals studied by postiron emission tomography and whole body autoradiography

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is a selective potent neurotoxin which has induced a syndrome similar to parkinsonism both in man and in monkeys. At autopsy degeneration of pigmented nerve cells in the pars compacta of the substantia nigra has been confirmed. The regional distribution of intravenously administered 1-(11C-methyl)-4-phenyl-1,2,3,6-tetrahydropyridine (11C-MPTP) in the brain of Rhesus monkeys was studied by positron emission tomography and the whole body distribution in mice was documented by autoradiography and by impulse counting of selected tissues. A very rapid and high uptake of 11C-MPTP derived radioactivity was seen in areas corresponding to striatum and midbrain, including the substantia nigra area. No elimination from these regions was seen during the study period of 2 h. The uptake was in the order of 7-8 times the homogenous distribution of the radioactivity in the monkey. The uptake was generally high also in other regions of the brain, but there some elimination could be distinguished. Pretreatment of the monkey with spiperone, a selective dopamine receptor antagonist, did not alter uptake nor the kinetics of the 11C-MPTP derived radioactivity. Thus 11C-MPTP does not have a high affinity for postsynaptic dopamine receptors. A remarkably high uptake of 11C-MPTP derived radioactivity was seen in the eye of the monkey. The selective uptake of radioactivity in the eye was also confirmed in pigmented but not in albino mice. The melanin affinity of MPTP may cause high intracellular concentrations of the compound or its metabolites in the melanin containing nerve cells in substantia nigra, which may explain the serious vulnerability of these neurons to MPTP.     

Deficiency of inducible nitric oxide synthase protects against MPTP toxicity in vivo

MPTP produces clinical, biochemical, and neuropathologic changes reminiscent of those that occur in idiopathic Parkinson's disease (PD). In the present study we show that MPTP treatment led to activation of microglia in the substantia nigra pars compacta (SNpc), which was associated and colocalized with an increase in inducible nitric oxide synthase (iNOS) expression. In iNOS-deficient mice the increase of iNOS expression but not the activation of microglia was blocked. Dopaminergic SNpc neurons of iNOS-deficient mice were almost completely protected from MPTP toxicity in a chronic paradigm of MPTP toxicity. Because the MPTP-induced decrease in striatal concentrations of dopamine and its metabolites did not differ between iNOS-deficient mice and their wild-type littermates, this protection was not associated with a preservation of nigrostriatal terminals. Our results suggest that iNOS-derived nitric oxide produced in microglia plays an important role in the death of dopaminergic neurons but that other mechanisms contribute to the loss of dopaminergic terminals in MPTP neurotoxicity. We conclude that inhibition of iNOS may be a promising target for the treatment of PD.     

1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine Metabolism and l-Methyl-4-Phenylpyridinium Uptake in Dissociated Cell Cultures from the Embryonic Mesencephalon

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a contaminant found in a synthetic illicit drug, can elicit in humans and monkeys a severe extrapyramidal syndrome similar to Parkinson's disease. It also induces alterations of the dopamine (DA) pathways in rodents. MPTP neurotoxicity requires its enzymatic transformation into 1-methyl-4-phenylpyridinium (MPP+) by monoamine oxidase followed by its concentration into target cells, the DA neurons. Here, we show that mesencephalic glial cells from the mouse embryo can take up MPTP in vitro, transform it into MPP+, and release it into the culture medium. MPTP is not taken up by neurons from either the mesencephalon or the striatum in vitro (8 days in serum-free conditions). However, mesencephalic neurons in culture revealed a high-affinity uptake mechanism for the metabolite MPP+, similar to that for DA. The affinity (Km) for DA uptake is fivefold higher than that for MPP+ (0.2 and 1.1 microM, respectively), whereas the number of uptake sites for MPP+ is double (Vmax = 25 and 55 pmol/mg of protein/min for DA and MPP+, respectively). Mazindol, a DA uptake inhibitor, blocks the uptake of DA and MPP+ equally well under these conditions. Moreover, by competition experiments, the two molecules appear to use the same carrier(s) to enter DA neurons. Small concentrations of MPP+ are also taken up by striatal neurons in vitro. The amount taken up represented less than 10% of the MPP+ uptake in mesencephalic neurons. Depolarization induced by veratridine released comparable proportions of labeled DA and MPP+ from mesencephalic cultures.(ABSTRACT TRUNCATED AT 250 WORDS)     

Resistance to MPTP-neurotoxicity in α-synuclein knockout mice is complemented by human α-synuclein and associated with increased β-synuclein and Akt activation

Genetic and biochemical abnormalities of α-synuclein are associated with the pathogenesis of Parkinson's disease. In the present study we investigated the in vivo interaction of mouse and human α-synuclein with the potent parkinsonian neurotoxin, MPTP. We find that while lack of mouse α-synuclein in mice is associated with reduced vulnerability to MPTP, increased levels of human α-synuclein expression is not associated with obvious changes in the vulnerability of dopaminergic neurons to MPTP. However, expressing human α-synuclein variants (human wild type or A53T) in the α-synuclein null mice completely restores the vulnerability of nigral dopaminergic neurons to MPTP. These results indicate that human α-synuclein can functionally replace mouse α-synuclein in regard to vulnerability of dopaminergic neurons to MPTP-toxicity. Significantly, α-synuclein null mice and wild type mice were equally sensitive to neurodegeneration induced by 2'NH(2)-MPTP, a MPTP analog that is selective for serotoninergic and noradrenergic neurons. These results suggest that effects of α-synuclein on MPTP like compounds are selective for nigral dopaminergic neurons. Immunoblot analysis of β-synuclein and Akt levels in the mice reveals selective increases in β-synuclein and phosphorylated Akt levels in ventral midbrain, but not in other brain regions, of α-synuclein null mice, implicating the α-synuclein-level dependent regulation of β-synuclein expression in modulation of MPTP-toxicity by α-synuclein. Together these findings provide new mechanistic insights on the role α-synuclein in modulating neurodegenerative phenotypes by regulation of Akt-mediated cell survival signaling in vivo.     

Histofluorescence studies of monoamine neurons in the rat brain after cobaltous acetate intoxication

The aim was to study changes in brain monoamine neurons in an experimental animal model with an extrapyramidal motor syndrome of the parkinsonian type. The neurological signs were observed in rats after acute cobaltous acetate intoxication under mild ischemic conditions. Histofluorescence studies showed a decrease in catecholamine fluorescence (which signifies a decrease in the amine content) in the hypothalamus and mesencephalic reticular formation, but not in the substantia nigra or basal ganglia. Serotonin fluorescence was increased in nerve cell bodies of the dorsal and median raphe nuclei and in nerve terminals in some thalamic and preoptic regions. Histological staining of sections adjacent to the fluorescent ones showed no neuronal loss and some pathology of myelin. The disturbing effect of cobaltous ions on the neuronal transmission, and/or the imbalance between dopamine and serotonin in the extrapyramidal motor syndrome observed in poisoned rats have been discussed.     

Overexpression of Parkinson's disease-associated alpha-synucleinA53T by recombinant adeno-associated virus in mice does not increase the vulnerability of dopaminergic neurons to MPTP

Mutations in the alpha-synuclein gene are linked to a rare dominant form of familial Parkinson's disease, and alpha-synuclein is aggregated in Lewy bodies of both sporadic and dominant Parkinson's disease. It has been proposed that mutated alpha-synuclein causes dopaminergic neuron loss by enhancing the vulnerability of these neurons to a variety of insults, including oxidative stress, apoptotic stimuli, and selective dopaminergic neurotoxins, such as 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). To test this hypothesis in vivo, we overexpressed human alpha-synuclein(A53T) in the substantia nigra of normal and MPTP-treated mice by rAAV-mediated gene transfer. Determination of dopaminergic neuron survival, striatal tyrosine hydroxylase fiber density, and striatal content of dopamine and its metabolites in rAAV-injected and uninjected hemispheres demonstrated that alpha-synuclein(A53T) does not increase the susceptibility of dopaminergic neurons to MPTP. Our findings argue against a direct detrimental role for (mutant) alpha-synuclein in oxidative stress and/or apoptotic pathways triggered by MPTP, but do not rule out the possibility that alpha-synuclein aggregation in neurons exposed to oxidative stress for long periods of time may be neurotoxic.