Intracarotid 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine Administration: Biochemical and Behavioral Observations in a Primate Model of Hemiparkinsonism

Cynomolgus monkeys received intracarotid injections of the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to produce a chronic unilateral model of parkinsonism. Extensive dopamine (DA) depletion was observed in the caudate nucleus and putamen on the side ipsilateral to the injection and this was associated with contralateral tremor, rigidity, and bradykinesia. A dose of 1.25 mg of MPTP caused ipsilateral DA loss of 99.4% in the caudate nucleus, 99.8% in the putamen, and 74.2% in the nucleus accumbens. A dose of 2.5 mg caused ipsilateral DA depletion of 99.3% in the caudate nucleus, 99.5% in putamen, and 90.1% in the nucleus accumbens. The unilateral aspect of the lesion was dose sensitive, with the 2.5-mg dose causing bilateral asymmetric DA depletion. Tissue concentrations of serotonin were not affected by the toxin. These findings confirm that intracarotid injection of MPTP may produce a useful primate model of hemiparkinsonism that can be associated with selective unilateral DA depletion when the appropriate dose of toxin is used.     

In Vivo Electrochemical Studies of Dopamine Overflow and Clearance in the Striatum of Normal and MPTP-Treated Rhesus Monkeys

Abstract: Rapid chronoamperometric recordings, using Nafion-coated carbon-fiber electrodes (30–90 µm o.d.), were used to investigate overflow and uptake of dopamine (DA) in the striatum of normal and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated rhesus monkeys. The monkeys were anesthetized with isoflurane and placed in a stereotaxic apparatus. Magnetic resonance imaging-guided sterile stereotaxic procedures were used for implantations of the electrochemical electrodes coupled with single-barrel micropipettes that were used to apply potassium or DA locally. Potassium evoked a robust overflow of DA-like electrochemical signals into the brain extracellular space in the unlesioned or normal putamen and caudate nucleus of the rhesus monkeys. In contrast, potassium did not produce any detectable changes (> 97% depletion) of DA in the MPTP-lesioned striatum. In addition, the diffusion/clearance of locally applied DA was markedly altered in the lesioned caudate nucleus and putamen compared with unlesioned striatum. Cell counts of the number of residual tyrosine hydroxylase-positive neurons in MPTP-treated monkeys, in conjunction with whole-tissue levels of DA and its metabolites, showed that the MPTP lesions produced extensive damage of the nigrostriatal DA system. These data indicate that residual dopaminergic fibers remaining after MPTP lesions are dysfunctional and have a greatly diminished capacity for high-affinity DA uptake.     

Nigrostriatal reduction of aromatic L-amino acid decarboxylase activity in MPTP-treated squirrel monkeys: in vivo and in vitro investigations

Aromatic L-amino acid decarboxylase (AAAD) activity was examined in vivo with positron emission tomography (PET) using 6-[18F]fluoro-L-DOPA (FDOPA) in squirrel monkeys lesioned with graded doses of the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). In vitro biochemical determinations of AAAD activity in caudate, putamen, substantia nigra, and nucleus accumbens were performed in the same animals to establish a direct comparison of in vivo and in vitro measurements. In vivo and in vitro AAAD activities in caudate/ putamen were substantially reduced in animals treated with the highest dose of MPTP (2.0 mg/kg). The percent change in the striatal FDOPA uptake (K(i)) and decarboxylation rate constant (k3) values resulting from MPTP treatment showed highly significant correlations with in vitro-determined AAAD activities. However, decarboxylase rates within individual animals presented as approximately 10-fold difference between in vivo and in vitro values. Lower in vivo k3 measurements may be attributed to several possibilities, including transport restrictions limiting substrate availability to AAAD within the neuron. In addition, reductions in AAAD activity in the substantia nigra did not parallel reductions in AAAD activity within the striatum, supporting the notion of a nonlinear relationship between nigrostriatal cell degeneration and terminal losses. This work further explores the role of AAAD in Parkinson's disease, a more important factor than previously thought.     

Experimental Parkinson's disease in monkeys. Effect of ergot alkaloid derivative on lipid peroxidation in different brain areas

The effects of the Parkinsonism induced by the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) were evaluated in four different monkey brain areas (frontal and occipital cortex, caudate putamen, substantia nigra). The basal and stimulated lipid peroxidation and the reduced glutathione (GSH) concentration were evaluated in three groups of maleMacaca fascicularis monkeys (6 animals/group): (a) controls; (b) MPTP-treated animals; (c) animals treated with MPTP and -dihydroergocryptine (DEK; ergot alkaloid characterized by a dopaminergic agonist action). In MPTP-treated animals the GSH concentration was unchanged or decreased in a non-significant way in the frontal and occipital cortex, and in substantia nigra. The basal thiobabituric acid reactive substance (TBARS) concentrations were significantly higher in the caudate putamen and substantia nigra of MPTP-treated animals. In the MPTP-treated monkeys the DEK administration induced a restoration of basal TBARS values to nearly normal ones. By incubating tissue from different brain areas with FeSO₄ plus ascorbic acid, the stimulation of lipid peroxidation decreased the TBARS production in the substantia nigra of the MPTP-treated animals. These results, taken together, may indicate that an increased lipid peroxidation could possibly play a role in producing the Parkinson-line syndrome by MPTP and that a free radical excess could be responsible for the degeneration of the substantia nigra. The treatment with an ergot alkaloid (i.e., -dihydroergocryptine) partially antagonizes the MPTP-induced increase in basal TBARS concentration in caudate putamen.     

Uptake of 1-Methyl-4-Phenylpyridinium and Dopamine in the Mouse Brain Cell Nuclei

Abstract: The neurotoxin N -methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) causes, via its metabolite 1-methyl-4-phenylpyridinium (MPP⁺), parkinsonism in humans, monkeys, and mice but not in rats. When incubated with mouse brain homogenates, [³H]-MPP⁺ is recovered in relatively large concentrations in the brain cell nucleus. Although isolated cell nuclei from rat and mouse brain contain uptake systems for dopamine (DA), only brain cell nuclei from mice avidly take up [³H]MPP⁺. This nuclear uptake is ATP dependent and can be blocked by ouabain and Nethylmaleimide. It is not, however, affected by neuronal and vesicular blockers such as benztropine. mazindol, and reserpine. Selective uptake of MPP⁺ into brain cell nuclei may provide a new avenue for future investigation into the complex modes of action of the neurotoxin MPTP.     

Physical activity is linked to ceruloplasmin in the striatum of intact but not MPTP-treated primates

Ceruloplasmin is a protective ferroxidase. Although some studies suggest that plasma ceruloplasmin levels are raised by exercise, the impact of exercise on brain ceruloplasmin is unknown. We have examined whether striatal ceruloplasmin is raised with treadmill exercise and/or is correlated with spontaneous physical activity in rhesus monkeys. Parkinson??s disease is characterized by a loss in ceruloplasmin and, similarly, Parkinson??s models lead to a loss in antioxidant defenses. Exercise might protect against Parkinson??s disease and is known to prevent antioxidant loss in experimental models. We have therefore examined whether treadmill exercise prevents ceruloplasmin loss in monkeys treated unilaterally with the dopaminergic neurotoxin MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine). We found that exercise raised ceruloplasmin expression in the caudate and accumbens but not the putamen of intact monkeys. However, putamen ceruloplasmin was correlated with spontaneous activity in a home pen. MPTP alone did not cause unilateral loss of ceruloplasmin but blocked the impact of exercise on ceruloplasmin. Similarly, the correlation between putamen ceruloplasmin and activity was also lost with MPTP. MPTP elicited loss of tyrosine hydroxylase in the treated hemisphere; the remaining tyrosine hydroxylase was correlated with overall daily activity (spontaneous activity plus that induced by the treadmill). Thus, treadmill activity can raise ceruloplasmin but this impact and the link with spontaneous activity are both diminished in Parkinsonian primates. Furthermore, low overall physical activity predicts greater loss of dopaminergic phenotype in MPTP-treated primates. These data have implications for the maintenance of active lifestyles in both healthy and neurodegenerative conditions.     

Studies on the Neurotoxicity of 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine: Inhibition of NAD-Linked Substrate Oxidation by Its Metabolite, 1-Methyl-4-Phenylpyridinium

Abstract: The effects of the parkinsonism-inducing neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and its 4-electron oxidation product 1-methyl-4-phenylpyridinium (MPP⁺) were studied in isolated mitochondria and in mouse brain striatal slices. ADP-stimulated oxidation of NAD-linked substrates was inhibited in a time-dependent manner by MPP⁺ (0.1–0.5 m M ), but not MPTP, in mitochondria prepared from rat brain, mouse brain, or rat liver. Under identical conditions, succinate oxidation was relatively unaffected. In neostriatal slices prepared from the mouse, a species susceptible to the dopaminergic neurotoxicity of MPTP, incubation with either MPP⁺ or MPTP caused metabolic changes consistent with inhibition of mitochondnial oxidation, i.e., an increase in the formation of lactate and accumulation of the amino acids glutamate and alanine with concomitant decreases in glutamine and aspartate levels. The changes resulting from incubation with MPTP were prevented by the monoamine oxidase inhibitor pargyline, which blocks formation of MPP⁺ from MPTP. The results suggest that compromise of mitochondrial function and its metabolic sequelae within dopaminergic neurons could be an important factor in the neurotoxicity observed after MPTP administration.     

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)     

Potentiation by the Tetraphenylboron Anion of the Effects of 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine and Its Pyridinium Metabolite

The 1-methyl-4-phenylpyridinium species (MPP+) is the four-electron oxidation product of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and is widely assumed to be the actual neurotoxic species responsible for the MPTP-induced destruction of dopaminergic neurons. MPTP is oxidized by the enzyme monoamine oxidase-B to a dihydropyridinium intermediate which is oxidized further to MPP+, an effective inhibitor of the oxidation of the Complex I substrates glutamate/malate in isolated mitochondrial preparations. In the present study, the tetraphenylboron anion (TPB) greatly potentiated the inhibitory effects of MPP+ and other selected pyridinium species on glutamate/malate respiration in isolated mouse liver mitochondria. At 10 microM TPB, the potentiation ranged from approximately 50-fold to greater than 1,000-fold for the several pyridinium species tested. In other experiments, TPB greatly enhanced the accumulation of [3H]MPP+ by isolated mitochondrial preparations. This facilitation by TPB of MPP+ accumulation into mitochondria explains, at least in part, the potentiation by TPB of the above-mentioned inhibition of mitochondrial respiration. Moreover, TPB addition increased the amount of lactate formed during the incubation of mouse neostriatal tissue slices with MPTP and other tetrahydropyridines. The administration of TPB also potentiated the dopaminergic neurotoxicity of MPTP in male Swiss-Webster mice. All of these observations, taken together, are consistent with the premise that the inhibitory effect of MPP+ on mitochondrial respiration within dopaminergic neurons is the ultimate mechanism to explain MPTP-induced neurotoxicity.     

Differential Regulation of Neuronal Nicotinic Receptor Binding Sites Following Chronic Nicotine Administration

Abstract: The deleterious effect of the parkinsonian neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) on dopaminergic neurons of the substantia nigra is well established. In addition, increased glutamatergic drive to basal ganglia output nuclei is considered a likely contributor to the pathogenesis of Parkinson's disease. One possibility for the increased excitatory tone may be related to an impairment in glutamate uptake. As astrocytes possess efficient transport mechanisms for both MPTP and glutamate, we have examined the effect of this agent on d -aspartate uptake into these cells. Treatment of cultures with 50 µ M MPTP for 24 h decreased uptake by 39%. Kinetic analysis revealed that this effect was due to a 35% decrease in V _max with no change in the K _m. Treatment with deprenyl, a monoamine oxidase B inhibitor, produced a complete reversal of MPTP-induced uptake inhibition, but was ineffective following exposure of cells to the MPTP metabolite, 1-methyl-4-phenylpyridinium (MPP⁺). Removal of MPTP from cultures resulted in a complete restoration of glutamate uptake after 24 h. These results show that MPTP reversibly compromises glutamate uptake in cultured astrocytes, which is dependent on the conversion of MPTP to MPP⁺. Such findings suggest that the glutamate transporter in astrocytes plays an important role in MPTP-induced neurotoxicity and possibly in parkinsonism.     

2-Deoxyglucose Enhances 1-Methyl-4-Phenyl-1, 2, 3, 6-Tetrahydropyridine-Induced ATP Loss in the Mouse Brain

Abstract: The effects of 2-deoxyglucose (2-DG), an inhibitor of the uptake and use of glucose, on ATP loss caused by the neurotoxicant 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) were determined in the mouse brain. 2-DG alone had no effect on brain ATP levels, but when administered 30 min before MPTP exposure, 2-DG significantly enhanced MPTP-induced ATP reduction. This was reflected as an increase in ATP loss in the striatum (from 15 to 27%) as well as a significant decrease in ATP in the cerebellar cortex, an area of the brain that was not affected after exposure to MPTP alone. In mice pretreated with 2-DG, striatal ATP levels remained significantly decreased for >8 h after MPTP administration. In contrast, ATP levels in the cerebellar cortex returned to normal values within 4 h from MPTP exposure. Mazindol, a catecholamine uptake blocker, completely protected against MPTP-induced loss of striatal ATP in the absence of 2-DG, but it only partially prevented striatal ATP decrease after administration of both 2-DG and MPTP; mazindol was also ineffective in protecting against ATP loss caused by 2-DG and MPTP in the cerebellar cortex. 2-DG/MPTP-induced ATP loss appeared to be associated with the presence of the 1 -methyl-4-phenylpyridinium (MPP⁺) metabolite because (1) the pattern of ATP recovery in the striatum and cerebellar cortex appeared to reflect the pattern of MPP⁺clearance from these areas of the brain (i.e., significant MPP⁺ levels persisted longer in the striatum than in the cerebellar cortex), and (2) ATP decrease was completely prevented by blocking the conversion of MPTP to MPP⁺with the monoamine oxidase B inhibitor deprenyl. Data indicate that impairment of glucose metabolism dramatically enhances the effects of MPTP/MPP⁺ on cerebral energy supplies, making these effects relatively nonselective for dopaminergic neurons of the nigrostriatal pathway.     

Accumulation of 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine in Cultured Cerebellar Astrocytes

Cultured cerebellar astrocytes rapidly accumulate 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) from the incubation medium, reaching a plateau within 10 min, whereas within that time negligible amounts of 1-methyl-4-phenylpyridinium (MPP+) have entered the astrocytes. MPTP accumulation is essentially independent of temperature and is proportional to extracellular concentration at steady state: The steady-state concentration achieved within these cells is about 50-fold higher at relatively low extracellular concentrations. MPTP appears to accumulate intracellularly within lysosomes, because lysosomotropic agents such as ammonium chloride and chloroquine markedly diminish the accumulation. Moreover, a proton gradient is required, because MPTP accumulation is abolished by the hydrogen ion antiporter monensin. Over an interval of several days, MPTP is converted to MPP+ intracellularly, with a concomitant decrease in medium MPTP and increase in medium MPP+. A constant, small but significant amount of MPP+ is retained intracellularly over a 72-h interval. Increasing the medium MPTP concentrations results in increased conversion of MPTP and enhanced intracellular retention of MPTP and MPP+. Neither MPTP nor MPP+ is neurotoxic to cultured cerebellar astrocytes as determined by cell counts and rate of conversion of MPTP to MPP+.     

Effects of l-Methyl-4-Phenyl-l,2,3,6-Tetrahydropyridine in the Dog: Effect of Pargyline Pretreatment

Adult beagle dogs of either sex were injected with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-HCl (2.5 mg/kg, i.v.) alone or after pretreatment with pargyline (5.0 mg/kg, s.c., twice), with pargyline alone, or were uninjected. Groups were killed 2 h, 3 weeks, or 3 months after injection, and several brain areas were assayed for biogenic amines and their synthetic and degradative enzymes. MPTP caused a massive and permanent loss of striatal dopamine, tyrosine hydroxylase, and 3,4-dihydroxyphenylalanine decarboxylase activities and the loss of cells within the substantia nigra pars compacta. Dopamine and norepinephrine also were depleted to various degrees in cortex, olfactory bulb, and hypothalamus; however, dopamine beta-hydroxylase activity in cortex was normal. There was no cell loss in the ventral tegmental area or locus ceruleus. The activities of monoamine oxidase (MAO)-A and MAO-B in cortex and caudate were not affected by MPTP. Despite a permanent loss of the nigrostriatal system, the dogs exhibited only a transient hypokinesia lasting 1-2 weeks. Pargyline pretreatment prevented the loss of striatal dopamine and cells from the substantia nigra, but did not prevent a prolonged but reversible decrease in the concentration of dopamine metabolites. It is argued that this apparent inhibition of MAO is due not to suicide inactivation of the enzyme by MPTP, but to reversible inhibition by accumulation of the pyridinium metabolite, 1-methyl-4-phenylpyridinium, selectivity in aminergic terminals.     

单侧帕金森病猴模型的行为、神经递质、多巴胺受体、病理及局部脑血流研究

经右侧颈总动脉注射甲基-苯基-四氢吡啶(MPTP)使猴产生左侧肢体动作减少、行动迟缓、震颤及向右侧缓慢旋转。应用美多巴和阿朴吗啡显著地改善单侧帕金森病(PD)症状,同时引起向左侧快速旋转,并呈明显的剂量依赖性;应用苯丙胺引起向右侧快速旋转。连续应用美多巴诱致单侧PD猴产生舞蹈手足徐动症。高效液相色谱测定显示右侧壳核、尾状核和黑质多巴胺(DA)含量显著降低。光镜发现右侧黑质神经元变性。SPECT活体显像发现病损侧纹状体D_2DA受体活性在病损初期无改变,病损严重时超敏,以及病损侧脑血流灌注减低。实验表明MPTP可建成理想的能形象地模拟人类PD的单侧PD猴模型。SPECT是活体研究PD病理生理的有效检测手段。     

Quinolinic acid but not MK-801 protects the dopaminergic system from 1-methyl-4- phenyl-1,2,3,6-tetrahydropyridine-induced toxicity in goldfish retina

The toxicity of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, intravitreally injected in goldfish eye, involves interplexiform retinal neurons and depletes tyrosine hydroxylase immunoreactivity and dopamine levels. This induced neurotoxicity was prevented by the concomitant administration in nontoxic doses (10 μg) of quinolinic acid, an endogenous structural analogue of N-methyl -aspartate with excitotoxic properties. Quinolinic acid is ineffective on the retinal degeneration induced by 1-methyl-4- phenylpyridinium ion. This fact suggests that quinolinic acid inhibits the MAO-B oxidation of 1-methyl-4- phenyl-1,2,3,6-tetrahydropyridine. MK-801, a noncompetitive antagonist of glutamate NMDA-receptors, exerts partial protective effects on MPTP-induced delayed toxicity in mammals. In the goldfish eye, MK-801, injected in low concentration, and in conjunction with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine or 1-methyl-4-phenylpyridinium ion, did not prevent retinal neurodegeneration. Ten μg of MK-801 alone did not affect retinal neurons, while a higher concentration (20 μg) causes the chromatolysis of some photoreceptor nuclei.     

Chronic D1 and D2 dopaminomimetic treatment of MPTP-denervated monkeys: effects on basal ganglia GABA(A)/benzodiazepine receptor complex and GABA content.

The effect of various chronic dopaminergic treatments in 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) monkeys on the brain gamma-aminobutyric acid type A (GABA(A)) /benzodiazepine receptor complex and GABA content was investigated in order to assess the GABAergic involvement in dopaminomimetic-induced dyskinesia. Three MPTP monkeys received for one month pulsatile administrations of the D1 dopamine (DA) receptor agonist SKF 82958 whereas three others received the same dose of SKF 82958 by continuous infusion. A long acting D2 DA receptor agonist, cabergoline, was given to another three animals. Untreated MPTP as well as naive control animals were also included. Pulsatile SKF 82958 relieved parkinsonian symptoms but was also associated with dyskinesia in two of the three animals whereas animals treated continuously with SKF 82958 remained as untreated MPTP monkeys. Chronic cabergoline administration improved motor response with no persistent dyskinesia. MPTP treatment induced a decrease of 3H-flunitrazepam binding in the medial anterior part of caudate-putamen and an increase in the internal segment of globus pallidus (GPi) which was in general unchanged by pulsatile or continuous SKF 82958 administration. Throughout the striatum, binding of 3H-flunitrazepam remained reduced in MPTP monkeys treated with cabergoline but was not significantly lower than untreated MPTP monkeys. Moreover, cabergoline treatment reversed the MPTP-induced increase in 3H-flunitrazepam binding in the GPi. GABA concentrations remained unchanged in the striatum, external segment of globus pallidus and GPi following MPTP denervation. Pulsatile but not continuous SKF 82958 administration decreased putamen GABA content whereas cabergoline treatment decreased caudate GABA. No alteration in GABA levels were observed in the GPe and GPi following the experimental treatments. These results suggest that: (1) D2-like receptor stimulation with cabergoline modulates GABA(A) receptor density in striatal subregions anatomically related to associative cortical afferent and (2) the absence of dyskinesia in dopaminomimetic-treated monkeys might be associated with the reversal of the MPTP-induced upregulation of the GABA(A)/benzodiazepine receptor complex in the Gpi.     

Analysis of γ-Aminobutyric AcidA Receptor Subunits in the Mouse Cochlea by Means of the Polymerase Chain Reaction

Abstract: Unlike 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), which produces consistent decreases in levels of striatal dopamine (DA) with considerably smaller and more variable effects on mouse brain levels of serotonin (5-HT) and norepinephrine (NE), a novel amine-substituted MPTP analogue, 1-methyl-4-(2'-aminophenyl)-1,2,3,6-tetrahydropyridine (2'-NH₂-MPTP), administered in a standard mouse dosing paradigm for MPTP (20 mg/kg X 4) did not affect striatal DA but led to marked reductions (60–70%) in levels of 5-HT, 5-hydroxyindoleacetic acid (5-HIAA), and NE measured in frontal cortex and hippocampus 1 week after treatment. Another 2'-substituted MPTP analogue, 1-methyl-4-(2'-methylphenyl)-1,2,3,6-tetrahydropyridine, affected cortical and hippocampal 5-HT, 5-HIAA, and NE only minimally, while markedly reducing the DA content in striatum (90%), thus indicating that the substituent (-NH₂ versus -CH₃) at the 2'position is important for the differential effects of these MPTP analogues. In a replication study with a 3-week end point, hippocampal and cortical 5-HT, 5-HIAA, and NE levels remained depressed with no indication of recovery. These results suggest that 2'-NH₂-MPTP may be a novel, regionally selective neurotoxin for serotonergic and norad-renergic nerve terminals.     

Effects of 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine and 1-Methyl-4-Phenylpyridinium Ion on Activities of the Enzymes in the Electron Transport System in Mouse Brain

The effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and 1-methyl-4-phenylpyridinium ion (MPP+) on activities of enzyme complexes in the electron transport system were studied using isolated mitochondrial preparations from C57BL/6J mouse brains. Both MPTP and MPP+ dose-dependently inhibited activity of NADH-ubiquinone oxidoreductase (EC 1.6.5.3). The inhibition was reversible. Preincubation of freeze-thawed mitochondria with MPTP or MPP+ had no effect on the inhibition; however, when nonfrozen mitochondria were used, NADH-ubiquinone oxidoreductase activity was reduced to 46% of that in the nonincubated sample after a 5-min preincubation with MPTP and to 77% of that in the nonincubated sample after a 5-min preincubation with MPP+. Kinetic analyses revealed that inhibition of MPTP was noncompetitive and that of MPP+ uncompetitive with respect to NADH. On the other hand, inhibition of MPTP was uncompetitive and that of MPP+ noncompetitive with respect to ubiquinone. Succinate-ubiquinone oxidoreductase (complex II), dihydroubiquinone-cytochrome c oxidoreductase (complex III), and ferrocytochrome c-oxygen oxidoreductase (EC 1.9.3.1) activities were either slightly inhibited or not inhibited by MPTP or MPP+. The significance of these findings is discussed in relation to the mechanism of MPTP-induced neuronal degeneration.     

Selective Destruction of Cultured Dopaminergic Neurons from Fetal Rat Mesencephalon by 1-Methyl-4-Phenylpyridinium: Cytochemical and Morphological Evidence

Dopaminergic neurons in cultures of dissociated cells from fetal rat mesencephalon were exposed to the principal metabolite of the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), 1-methyl-4-phenyl-pyridinium ion (MPP+), and several of its structural analogues. At concentrations between 0.01 and 0.1 microM, MPP+ inhibited catecholamine accumulation as visualized by cytofluorescence. Between 0.1 and 10.0 microM, MPP+ resulted in disappearance of tyrosine hydroxylase immunoreactivity without affecting other cells in the cultures. At concentrations higher than 10 microM, MPP+ was toxic to all cells present in the cultures. The effect of low concentrations of MPP+ on catecholamine cytofluorescence of the dopaminergic neurons was partially reversible. The intermediate concentrations produced irreversible structural changes of tyrosine hydroxylase-positive cells, resulting in complete disappearance of these neurons. The morphological changes were specific to the dopaminergic neurons and were not evident in other cells viewed with phase contrast microscopy. Of the structural analogues tested, the 1-ethyl analogue of MPP+ was effective in selectively destroying dopaminergic neurons in our culture system. The antioxidants L-acetyl-carnitine, beta-carotene, and alpha-tocopherol failed to protect against MPP+ neurotoxicity when co-incubated with the toxin.     

Prolonged Alterations in Canine Striatal Dopamine Metabolism Following Subtoxic Doses of l-Methyl-4-Phenyl- 1,2,3,6-Tetrahydropyridine (MPTP) and 4''-Amino-MPTP Are Linked to the Persistence of Pyridinium Metabolites

Single toxic doses of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP).HCl (2.5 mg/kg i.v.) and 4'-amino-MPTP.2HCl (22.5 mg/kg) induce loss of striatal dopamine (DA) and tyrosine hydroxylase (TH) activity and of nigral DA neurons in the dog. To examine the subacute neurochemical changes induced by low doses of MPTP and 4'-amino-MPTP, dose-response studies of these compounds were carried out in the dog, using 6- and 3-week survival times for these two compounds, respectively. Low single doses of MPTP (1.0, 0.5, and 0.1 mg/kg i.v.) and 4'-amino-MPTP (15, 7.5, and 3.75 mg/kg i.v.) did not cause depletion of canine striatal DA or TH or a loss of nigral neurons. However, levels of the DA metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) were decreased in a dose-related fashion, with significant loss of DOPAC being evident 6 weeks after the lowest administered dose of MPTP and 3 weeks after 4'-amino-MPTP. This selective loss of DA metabolites following nontoxic doses of MPTP and 4'-amino-MPTP led to a shift in the ratio of DA to DOPAC or HVA, which was characteristic for each compound. The measurement of striatal 1-methyl-4-phenylpyridinium (MPP+) and 4'-amino-MPP+ levels revealed that high concentrations (up to 150 microM) persist in the striatum for weeks following administration of a single nontoxic dose of MPTP or 4'-amino-MPTP. A causal relationship between the striatal concentration of MPP+ or 4'-amino-MPP+ and the change in DA metabolism as reflected in the DA/DOPAC ratio is suggested by a significant correlation between these measures. It is suggested that presynaptic sequestration and retention of MPP+ and 4'-amino-MPP+ by striatal DA terminals result in the inhibition of the monoamine oxidase contained within these terminals.