Effects of 1-Methyl-4-Phenylpyridinium on Isolated Rat Brain Mitochondria: Evidence for a Primary Involvement of Energy Depletion

Abstract: The effects of 1-methyl-4-phenylpyridinium (MPP⁺) on the oxygen consumption, ATP production, H₂O₂ production, and mitochondrial NADH-CoQ₁ reductase (complex I) activity of isolated rat brain mitochondria were investigated. Using glutamate and malate as substrates, concentrations of 10–100 µ M MPP⁺ had no effect on state 4 (−ADP) respiration but decreased state 3 (+ADP) respiration and ATP production. Incubating mitochondria with ADP for 30 min after loading with varying concentrations of MPP⁺ produced a concentration-dependent decrease in H₂O₂ production. Incubation of mitochondria with ADP for 60 min after loading with 100 µ M MPP⁺ caused no loss of complex I activity after washing of MPP⁺ from the mitochondrial membranes. These data are consistent with MPP⁺ initially binding specifically to complex I and inhibiting both the flow of reducing equivalents and the production of H₂O₂ by the mitochondrial respiratory chain, without irreversibly damaging complex I. However, mitochondria incubated with H₂O₂ in the presence of Cu²⁺ ions showed decreased complex I activity. This study provides additional evidence that cellular damage initiated by MPP⁺ is due primarily to energy depletion caused by specific binding to complex I, any increased damage due to free radical production by mitochondria being a secondary effect.     

Neuroprotective Effect of the Iron Chelator Desferrioxamine Against MPP+ Toxicity on Striatal Dopaminergic Terminals

Abstract: Microdialysis was used to evaluate the effect of desferrioxamine (DES) against 1-methyl-4-phenylpyridinium (MPP⁺) toxicity. The presence of DES (40 fmol-40 nmol/15 min for a total of 90 min) in the Ringer solution, coperfused with MPP⁺ (40 nmol/15 min) on day 1, produced on day 2 a higher extracellular dopamine output after perfusion of MPP⁺ than in control MPP⁺ perfusion experiments, in which no DES was administered on day 1. Both Ringer perfusion alone (control Ringer) and coperfusion of 40 nmol DES with 40 nmol MPP⁺ on day 1 produced on day 2 similar increases in extracellular dopamine output after a second MPP⁺ perfusion. In the control Ringer experiment, note that the MPP⁺ on day 2 is the first MPP⁺ perfusion. Perfusion of 800 fmol FeCl₃/15 min along with 40 nmol MPP⁺ and 400 fmol DES on day 1 completely abolished on day 2 the neuroprotective effect found with 40 nmol MPP⁺ and 400 fmol DES; 800 fmol FeCl₃ did not increase the neurotoxic effect of 40 nmol MPP⁺ perfusion. The ability of DES to protect against MPP⁺ toxicity may indicate a therapeutic strategy in the treatment of diseases when iron is implicated.     

Protection and Potentiation of 1-Methyl-4-Phenylpyridinium-Induced Toxicity by Cytochrome P450 Inhibitors and Inducer May Be Due to the Altered Uptake of the Toxin

Abstract: Earlier studies from our laboratory have demonstrated that 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) toxicity could be modulated by inhibitors and inducer of cytochrome P450 (P450) in an in vitro model consisting of sagittal slices of mouse brain. To understand the molecular mechanisms underlying the role of P450 on MPTP toxicity, it was undertaken to study the effect of the modulators of P450 on the toxicity of the metabolite of MPTP, namely, 1-methyl-4-phenylpyridinium ion (MPP⁺). Incubation of mouse brain slices with various concentrations of MPP⁺ (1–100 µ M ) resulted in dose-dependent inhibition of mitochondrial enzyme NADH-dehydrogenase (NADH-DH) and leakage of the cytosolic enzyme lactate dehydrogenase from the slice into the medium. MPP⁺-induced toxicity was abolished by pretreatment of the slices with inhibitors of monoamine oxidase (MAO; pargyline and deprenyl) or inhibitors of P450 (piperonyl butoxide or SKF-525A) or dopamine uptake blocker (GBR-12909), as measured by the activity of NADH-DH in slices and leakage of lactate dehydrogenase from the slice into the medium. Slices prepared from mice pretreated with phenobarbital (an inducer of P450) potentiated the toxic effects of MPP⁺. Pretreatment of slices with MAO-inhibitor, P450 inhibitors, or dopamine uptake blocker attenuated the uptake of MPP⁺ into the slices. In contrast, MPP⁺ uptake was significantly increased in slices prepared from phenobarbital-pretreated mice. Thus, both MAO and P450 inhibitors abolish the toxicity of MPP⁺ in the sagittal slices of mouse brain by altering the uptake of the toxin into the slices.     

Reactivation of NADH Dehydrogenase (Complex I) Inhibited by 1-Methyl-4-(4'-Alkylphenyl)pyridinium Analogues: A Clue to the Nature of the Inhibition Site

Abstract: Expression of the neurotoxicity of 1-methyl-4-phenyl-1.2,3,6-tetrahydropyridine, following oxidation to l-methyl-4-phenylpyridinium ion (MPP⁺), is believed to involve inhibition of mitochondrial electron transport from NADH dehydrogenase (complex l) to ubquinone. MPP⁺ and its analogues have been shown to Mock electron transport at or near the same site as two powerful inhibitors of mitochondrial respiration, rotenone and piericidin A. All three types of inhibitors combine at two sites on NADH dehydrogenase, a hydrophilic and hydrophobic one, and occupancy of both sites is required for complete inhibition. Tetraphenylboron anion (TPB⁻) in catalytic amounts is known to increase the effectiveness of positively charged MPP⁺ analogues in blodclng mitochondrial respiration. A part of this effect involves facitation of the entry of MPP⁺ oongeners into the hydrophobic site by ion pairing, as has been demonstrated in studies with submitochondrial particles (electron transport particles). This communication documents the fact that TPB⁻, when present in molar excess over the MPP⁺ analogues, reverses the inhibition. This seems to involve again strong ion pairing. removal of the inhibitory analogue from one to the two binding sites, and concentration of the inhibitor in the membrane, so that only the hydrophobic binding site remains occupied, resulting in lowering of the inhibiti to 30–40%.     

Pramipexole Reduces Reactive Oxygen Species Production In Vivo and In Vitro and Inhibits the Mitochondrial Permeability Transition Produced by the Parkinsonian Neurotoxin Methylpyridinium Ion

Abstract: Sporadic Parkinson's disease is associated with a defect in the activity of complex I of the mitochondrial electron transport chain. This electron transport chain defect is transmitted through mitochondrial DNA, and when expressed in host cells leads to increased oxygen free radical production, increased antioxidant enzyme activities, and increased susceptibility to programmed cell death. Pramipexole, a chemically novel dopamine agonist used for the treatment of Parkinson's disease symptoms, possesses antioxidant activity and is neuroprotective toward substantia nigral dopamine neurons in hypoxic-ischemic and methamphetamine models. We found that pramipexole reduced the levels of oxygen radicals produced by methylpyridinium ion (MPP⁺) both when incubated with SH-SY5Y cells and when perfused into rat striatum. Pramipexole also exhibited a concentration-dependent inhibition of opening of the mitochondrial transition pore induced by calcium and phosphate or MPP⁺. These results suggest that pramipexole may be neuroprotective in Parkinson's disease by attenuating intracellular processes such as oxygen radical generation and the mitochondrial transition pore opening, which are associated with programmed cell death.     

Involvement of a Caspase-3-Like Cysteine Protease in 1-Methyl-4-Phenylpyridinium-Mediated Apoptosis of Cultured Cerebellar Granule Neurons

Abstract: Exposure of various neuronal cells or cell lines to high concentrations of 1-methyl-4-phenylpyridinium (MPP⁺), the active metabolite of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), results in cell death. Recently, it has been reported that low concentrations of MPP⁺ induce apoptosis in susceptible neurons. We have further characterized MPP⁺-mediated toxicity of cultured cerebellar granule neurons (CGNs) and found that exposure of CGNs to relatively low concentrations of MPP⁺ results in apoptosis, whereas higher concentrations result in necrosis. Cotreatment of CGNs with MPP⁺ and the tetrapeptide inhibitor of caspase-3-like proteases, acetyl-DEVD-CHO, markedly attenuates apoptotic but not necrotic death of these neurons. The more specific inhibitor of caspase-1-like proteases, acetyl-YVAD-CHO, however, was ineffective against MPP⁺ neurotoxicity. Moreover, cytoplasmic extracts prepared from MPP⁺-treated CGNs contain markedly increased protease activity that cleaves the caspase-3 substrate acetyl-DEVD- p -nitroaniline. Finally, the cytoplasmic concentration of the apoptogenic protein cytochrome c was increased in a time-dependent fashion in MPP⁺-treated CGNs before the onset of apoptosis. Our data confirm that the neurotoxicity of MPP⁺ is due to both necrosis and apoptosis and suggest that the latter is mediated by activation of a caspase-3-like protease.     

Assessment of the direct and indirect effects of MPP+ and dopamine on the human proteasome: implications for Parkinson's disease aetiology

Mitochondrial impairment, glutathione depletion and oxidative stress have been implicated in the pathogenesis of Parkinson's disease (PD), linked recently to proteasomal dysfunction. Our study analysed how these factors influence the various activities of the proteasome in human SH-SY5Y neuroblastoma cells treated with the PD mimetics MPP⁺ (a complex 1 inhibitor) or dopamine. Treatment with these toxins led to dose- and time-dependent reductions in ATP and glutathione and also chymotrypsin-like and post-acidic like activities; trypsin-like activity was unaffected. Antioxidants blocked the effects of dopamine, but not MPP⁺, suggesting that oxidative stress was more important in the dopamine-mediated effects. With MPP⁺, ATP depletion was a prerequisite for loss of proteasomal activity. Thus in a dopaminergic neuron with complex 1 dysfunction both oxidative stress and ATP depletion will contribute independently to loss of proteasomal function. We show for the first time that addition of MPP⁺ or dopamine to purified samples of the human 20S proteasome also reduced proteasomal activities; with dopamine being most damaging. As with toxin-treated cells, chymotrypsin-like activity was most sensitive and trypsin-like activity the least sensitive. The observed differential sensitivity of the various proteasomal activities to PD mimetics is novel and its significance needs further study in human cells.     

Altered Calcium Homeostasis in Cells Transformed by Mitochondria from Individuals with Parkinson's Disease

Abstract: Parkinson's disease may be linked to defects in mitochondrial function. Mitochondrially transformed cells (cybrids) were created from Parkinson's disease patients or disease-free controls. Parkinson's disease cybrids had 26% less complex I activity, but maintained comparable basal calcium and energy levels. Parkinson's disease cybrids recovered from a carbachol-induced increase in cytosolic calcium 53% more slowly than controls even with lanthanum and thapsigargin blockade. Inhibition of complex I with the Parkinson's disease-inducing metabolite 1-methyl-4-phenylpyridinium (MPP⁺) similarly reduced the rate of recovery after carbachol. This MPP⁺-induced reduction in recovery rates was much more pronounced in control cybrids than in Parkinson's disease cybrids. Parkinson's disease cybrids had less carbonyl cyanide m -chlorophenylhydrazone-releasable calcium. Bypassing complex I with succinate partially restored Parkinson's disease cybrid, and MPP⁺ suppressed control cybrid recovery rates. The subtle alteration in calcium homeostasis of Parkinson's disease cybrids may reflect an increased susceptibility to cell death under circumstances not ordinarily toxic.     

Differential Interaction of 1-Methyl-4-Phenylpyridinium Ion with the Putatively Vesicular Binding Site of [3H]Tyramine in Dopaminergic and Nondopaminergic Brain Regions

Abstract: The neurotoxin 1-methyl-4-phenylpyridinium ion (MPP⁺) in the brain striatum has recently been shown to bind at a putatively vesicular site labeled by [³H]tyramine ([³H]TY). Whereas in the rat and mouse striatum MPP⁺ antagonized TY binding competitively, in the cerebellum there was a mixed-type antagonism, which suggests the simultaneous occupancy of two different sites. K _i values from displacement curves revealed a fourfold difference in the affinity of MPP⁺ for TY sites in the two brain regions. The degeneration of central noradrenergic terminals induced by an intraperitoneal injection of the toxin N -(2-chloroethyl)- N -ethyl-2-bromobenzylamine in rats decreased by 80% the maximal number of cerebellar TY binding sites, while not affecting striatal binding. Furthermore, guanethidine, a marker for noradrenaline (NA) vesicles, potently inhibited TY binding in NA-innervated regions, such as the cerebellum and the parietal cortex, and poorly in the striatum. It is concluded (a) that both MPP⁺ and TY may also label NA vesicles and (b) that the vesicular carriers for dopamine and NA have different characteristics, which may underlie a regional specificity in the rate of endovesicular sequestration of MPP⁺, with either neurodegenerative or neuroprotective consequences, depending on the brain area involved.     

Involvement of multiple protein kinases in cPLA2 phosphorylation, arachidonic acid release, and cell death in in vivo and in vitro models of 1-methyl-4-phenylpyridinium-induced parkinsonism – the possible key role of PKG

The study was aimed at investigating in vivo and in vitro the involvement of the cGMP/cGMP-dependent protein kinase (PKG) signaling pathway in MPP⁺-induced cytosolic phospholipase A₂ (cPLA₂) activation of dopaminergic neurons. MPP⁺ activated neuronal nitric oxide synthase (NOS)/soluble guanylyl cyclase/cGMP pathway in mouse midbrain and striatum, and in pheochromocytoma cell line 12 cells, and caused an upward shift in [Ca²⁺]_i level in the latter. The activation was accompanied by increases in total and phosphorylated cPLA₂, and increased arachidonic acid release. Effects of selective inhibitors [2-oxo-1,1,1-trifluoro-6,9-12,15-heneicosatetraene (AACOCF₃), (E)-6-(bromomethylene)tetrahydro-3-(1-naphthalenyl)2h-pyran-2-one (BEL)] indicated the main impact of cPLA₂ on arachidonic acid release in pheochromocytoma cell line 12 cells. Treatment of the cells with the protein kinase inhibitors GF102610x, UO126, and KT5823, and with the nitric oxide synthase (NOS) inhibitor NNLA revealed the involvement of protein kinase C (PKC) and extracellular signal-regulated kinases 1 and 2 (ERK 1/2), with the possible key role of PKG, in cPLA₂ phosphorylation at Ser505. Inhibitors of cPLA₂ and PKG increased viability and reduced MPP⁺-induced apoptosis of the cells. Our results indicate that the neuronal NOS/cGMP/PKG pathway stimulates cPLA₂ phosphorylation at Ser505 by activating PKC and ERK1/2, and suggest that up-regulation of this pathway in experimental models of Parkinson's disease may mediate dopaminergic neuron degeneration and death through activation of cPLA₂.     

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.     

Effects of NH+4-N and NO+3-N on growth and metabolism of Sphagnum magellanicum

Photosynthetic CO₂-fixation, chlorophyll content, growth rate and nitrate reductase activity were used to examine the influence of NH⁺₄-N and NO⁻₃-N on Sphagnum magellanicum cultivated under defined conditions in phytotrons. NO⁻₃-concentrations up to 322 μ M were found to be favourable. Increased NH⁺₄ concentrations, however, resulted in growth inhibition and decreased chlorophyll content at concentrations ≧ 255 μ M ; e.g. 600 μ M NH⁺₄ caused a 20% reduction of nitrate reductase activity and net photosynthesis. For raised bog Sphagna an improved standard nutrient solution is proposed with the following ion concentrations (μ M ): 55 Na⁺; 17 K⁺; 95 NH⁺₄; 22 Ca²⁺; 22 Mg²⁺; 2 Fe³⁺; 20 Cl⁻; 100 NO⁻₃; 57 SO^2-₄; 7.4 H₂PO⁻₄; trace elements: A-Z solution (Hoagland) 50 μl 1000 ml⁻¹; pH 5.8.     

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.     

Glial Cell Line-Derived Neurotrophic Factor Exerts Neurotrophic Effects on Dopaminergic Neurons In Vitro and Promotes Their Survival and Regrowth After Damage by 1-Methyl-4-Phenylpyridinium

Abstract: The effect of glial cell line-derived neurotrophic factor (GDNF) on the growth of mesencephalic dopaminergic neurons and on their survival following exposure to the neurotoxin 1-methyl-4-phenylpyridinium (MPP⁺) was examined in vitro. In cultures developing under normal conditions, GDNF at 1 ng/ml optimally improved the survival and stimulated the growth of dopaminergic neurons without affecting glial growth. In cultures treated with MPP⁺, GDNF could not prevent toxicity to dopaminergic neurons. The uptake of [³H]dopamine and the number of tyrosine hydroxylase-positive neurons were similarly reduced by MPP⁺ in the presence or absence of GDNF. However, after removal of MPP⁺, GDNF protected dopaminergic neurons from the continuous cell death and stimulated the regrowth of dopaminergic fibers damaged by MPP⁺. We conclude that GDNF supports the growth of normally developing dopaminergic neurons and stimulates their survival and recovery after damage. These findings suggest that GDNF could be useful in the development of therapeutic approaches to Parkinson's disease, which is characterized by dopaminergic cell loss.     

Novel imine antioxidants at low nanomolar concentrations protect dopaminergic cells from oxidative neurotoxicity

Strong evidence indicates that oxidative stress may be causally involved in the pathogenesis of Parkinson's disease. We have employed human dopaminergic neuroblastoma cells and rat primary mesencephalic neurons to assess the protective potential of three novel bisarylimine antioxidants on dopaminergic cell death induced by complex I inhibition or glutathione depletion. We have found that exceptionally low concentrations (EC₅₀ values ∼20 nM) of these compounds (iminostilbene, phenothiazine, and phenoxazine) exhibited strong protective effects against the toxicities of MPP⁺, rotenone, and l -buthionine sulfoximine. Investigating intracellular glutathione levels, it was found that MPP⁺, l -buthionine sulfoximine, and rotenone disrupted different aspects of the native glutathione equilibrium, while the aromatic imines did not further influence glutathione levels or redox state on any baseline. However, the imines independently reduced protein oxidation and total oxidant flux, saved the mitochondrial membrane potential, and provided full cytoprotection under conditions of complete glutathione depletion. The unusually potent antioxidant effects of the bisarylimines could be reproduced in isolated mitochondria, which were instantly protected from lipid peroxidation and pathological swelling. Aromatic imines may be interesting lead structures for a potential antioxidant therapy of Parkinson's disease and other disorders accompanied by glutathione dysregulation.     

Involvement of matrix NADP turnover in the oxidation of NAD-linked substrates by pea leaf mitochondria

The involvement of the internal rotenone-insensitive NADPH dehydrogenase on the inner surface of the inner mitochondrial membrane [ND_in(NADPH)] in the oxidation of strictly NAD⁺-linked substrates by pea ( Pisum sativum L.) leaf mitochondria was measured. As estimated by the inhibition caused by 5 μ M diphenyleneiodonium (DPI) in the presence of rotenone to inhibit complex I, the activity of ND_in(NADPH) during glycine oxidation (measured both as O₂ uptake and as CO₂ release) was 40–50 nmol mg⁻¹ protein min⁻¹. No significant activity of ND_in(NADPH) could be detected during the oxidation of 2-oxoglutarate, another strictly NAD⁺-linked substrate; this was possibly due to its relatively low oxidation rate. Control experiments showed that, even at 125 μ M , DPI had no effect on the activity of glycine decarboxylase complex (GDC) and lipoamide dehydrogenase. The relative activity of complex I, ND_in(NADPH), and ND_in(NADH) during glycine oxidation, estimated using rotenone and DPI, differed depending on the pyridine nucleotide supply in the mitochondrial matrix. This was shown by loading the mitochondria with NAD⁺ and NADP⁺, both of which were taken up by the organelle. We conclude that the involvement of NADP turnover during glycine oxidation is not due to the direct production of NADPH by GDC but is an indirect result of this process. It probably occurs via the interconversion of NADH to NADPH by the two non-energy-linked transhydrogenase activities recently identified in plant mitochondria.     

Fibroblast growth factor 9 prevents MPP+-induced death of dopaminergic neurons and is involved in melatonin neuroprotection in vivo and in vitro

Oxidative stress and down-regulated trophic factors are involved in the pathogenesis of nigrostriatal dopamine(DA)rgic neurodegeneration in Parkinson's disease. Fibroblast growth factor 9 (FGF9) is a survival factor for various cell types; however, the effect of FGF9 on DA neurons has not been studied. The antioxidant melatonin protects DA neurons against neurotoxicity. We used MPP⁺ to induce neuron death in vivo and in vitro and investigated the involvement of FGF9 in MPP⁺ intoxication and melatonin protection. We found that MPP⁺ in a dose- and time-dependent manner inhibited FGF9 mRNA and protein expression, and caused death in primary cortical neurons. Treating neurons in the substantia nigra and mesencephalic cell cultures with FGF9 protein inhibited the MPP⁺-induced cell death of DA neurons. Melatonin co-treatment attenuated MPP⁺-induced FGF9 down-regulation and DA neuronal apoptosis in vivo and in vitro . Co-treating DA neurons with melatonin and FGF9-neutralizing antibody prevented the protective effect of melatonin. In the absence of MPP⁺, the treatment of FGF9-neutralizing antibody-induced DA neuronal apoptosis whereas FGF9 protein reduced it indicating that endogenous FGF9 is a survival factor for DA neurons. We conclude that MPP⁺ down-regulates FGF9 expression to cause DA neuron death and that the prevention of FGF9 down-regulation is involved in melatonin-provided neuroprotection.     

The N-Methyl-D-Aspartate Antagonist MK-801 Fails to Protect Dopaminergic Neurons from 1-Methyl-4- Phenylpyridinium Toxicity In Vitro

Abstract: Recent reports suggest that NMDA receptor antagonists when administered in vivo can protect dopaminergic neurons from the toxic actions of MPP⁺. In the present study the possible neuroprotective effects against MPP⁺ toxicity of the noncompetitive NMDA receptor antagonist MK-801 was studied in primary cultures of fetal rat mesencephalic dopamine neurons. MK-801 failed to protect dopaminergic neurons from MPP⁺ toxicity at concentrations that completely block NMDA-induced toxicity of these same neurons. In contrast to work carried out in cerebellar granule cells, MPP⁺ toxicity of mesencephalic dopamine neurons was unaffected by preexposure to subtoxic concentrations of either NMDA or cycloheximide. Our findings suggest that the toxic effects of MPP⁺ on dopaminergic neurons are not mediated through a direct interaction with the NMDA subtype of glutamate receptor.     

Neurotoxic Effect of Intranigral Injection of 1-Methyl-4-Phenylpyridinium on GABA-Containing Neurons and Its Relation to Circling Behavior

Abstract: The ionic species 1-methyl-4-phenylpyridinium (MPP⁺) seems to be the metabolite responsible for the damage to dopaminergic neurons occurring after administration of the parkinsonian drug 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine. In the present study we show that the unilateral stereotaxic microinjection of MPP⁺ into the substantia nigra pars reticulata in rats produces immediately intense and long-lasting (up to 96 h) contralateral turning behavior in a dose-dependent manner. This behavioral effect was correlated with a dose- and time-dependent decrease (up to 90%) of glutamate decarboxylase activity and with a notable loss of neurons in the injected nigra reticulata. GABA levels in the injected nigra were also decreased, whereas the dopamine concentration in the ipsilateral striatum was not affected at 24 h, when maximal behavioral effects were observed. The circling behavior was prevented by the dopamine carrier blocker nomifensine only during the first 2 h, whereas the dopamine receptor antagonist haloperidol was ineffective. The results indicate that MPP⁺ is toxic for inhibitory GABAergic neurons in the nigra pars reticulata and, furthermore, suggest that disruption of the function of these GABAergic neurons may be involved in the abnormal motor behavior produced by the injection of MPP⁺ in the substantia nigra.