Melatonin fails to protect against long-term MPTP-Induced dopamine depletion in mouse striatum

Several laboratories recently have reported that melatonin may possess neuroprotective properties. The present paper presents the results of our studies on the long-termin vivo effects of melatonin in a well-defined neurotoxicity model using 1-methyl-4-phenyl-l,2,3,6-tetrahydropyridine (MPTP) in the C57BL/6 mouse. MPTP is bioactivated by brain monoamine oxidase B (MAO-B) to its neurotoxic pyridinium metabolite l-methyl-4-phenylpyridi-nium (MPP⁺) which destroys dopaminergic nerve terminals leading to the depletion of neostriatal dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC). Our initial study compared striatal DA and DOPAC levels in MPTP-only-treated animals and animals treated with melatonin 30 min prior to and 3 times hourly post-MPTP. DA/DOPAC levels measured 7 days after MPTP were similar in both groups. A second study was designed to address the possibility that melatonin cleared from the brain prior to MPP⁺. Animals, that had been administered the same regimen of melatonin as in the first study plus a fourth post-MPTP melatonin dose, were maintained on melatonin in drinking water until 5 days post-MPTP. Striatal DA/DOPAC levels of these melatonin-plus-MPTP treated animals also were the same as the MPTP-only-treated animals.In vitro studies confirmed that melatonin is not an inhibitor of MAO-B. These data demonstrate that melatonin does not have any significant protective effects against the long-term striatal DA and DOPAC depletion induced by MPTP in the C57BL/6 mouse.     

Attenuation of 1-methyl-4-phenylpyridinium (MPP+) neurotoxicity by deprenyl in organotypic canine substantia nigra cultures

Summary Systemic administration of MPTP to experimental animals induces neurodegeneration of dopaminergic neurons in the central nervous system. MPTP crosses the blood-brain barrier where it is taken up by astrocytes and converted to MPP⁺ by monamine oxidase-B (MAO-B). Subsequently, MPP⁺ is selectively taken up by dopaminergic neurons upon which it exerts intracellular neurotoxic effects. Systemic administration of the selective MAO-B inhibitor deprenyl prevents the conversion of MPTP to MPP⁺ and by this mechanism is able to protect against MPTP neurotoxicity. Deprenyl has also been reported to exert neuroprotective effects that are independent of its MAO-B inhibitory properties, but since MPP⁺ itself does not cross the blood-brain barrier it is difficult to directly study the MAO-B independentin vivo effects of MPP⁺ itself. One approach is to use organotypic tissue cultures of the canine substantia nigra (CSN) which permit administration of precise concentrations of pharmacological agents directly to mature, well-developed and metabolically active dopaminergic neurons. These neurons as well as other components of the cultures exhibit morphological and biochemical characteristics identical to theirin vivo counterparts. This study was undertaken to evaluate the neuroprotective effects of deprenyl in MPP⁺-treated cultures by measuring changes in the levels of HVA as an indicator of dopamine release and metabolism by dopaminergic neurons and to correlate this indication of dopaminergic function with morphological evidence of survival or loss of dopaminergic neurons in mature CSN cultures. Mature CSN cultures, at 44 days in vitro (DIV), were exposed to either MPP⁺ alone, deprenyl alone or simultaneously to both deprenyl and MPP⁺ or to MPP⁺ following 4 day pretreatment with deprenyl. Exposure to MPP⁺ alone caused significant reduction in HVA levels, evidence of widespread injury and ultimate disappearance of large neurons in the cultures. These effects were attenuated by simultaneous exposure to MPP⁺ and deprenyl and the destructive effects of MPP⁺ appeared to be prevented by pretreatment with deprenyl. Thus the neuroprotective effects of deprenyl on MPP⁺-induced reduction of HVA levels in living cultures appears similar to the effects of deprenyl on dopamine levels and tyrosine hydroxylase activity reported by others in cultures previously exposed to deprenyl and MPP⁺. These studies also confirm that the neuroprotective effects of deprenyl against MPP⁺ in dopaminergic neurons are, at least in part, independent of deprenyl's inhibition of MAO-B.     

Acute effects of 1-methyl-4-phenylpyridinium ion (MPP+) on dopamine and serotonin metabolism in rat striatum as assayed in vivo by a micro-dialysis technique

Summary The acute effect of 1-methyl-4-phenylpyridinium ion (MPP⁺), a neurotoxin derived from 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP), was examined by the in vivo micro-dialysis technique. A dialysis cannula was implanted into rat striatum, and the changes in the concentrations of dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), and 5-hydroxyindoleacetic acid (5-HIAA) in the perfusate every 20 min after administration of MPP⁺ were determined by high-performance liquid chromatography with electrochemical detection (HPLC-ED). After MPP⁺ administration the levels of DOPAC, HVA and 5-HIAA were markedly decreased. On the contrary the level of DA was markedly increased and reached a maximum 40 min after beginning of the MPP⁺ administration. By postmortem analysis of the striatal tissue MPP⁺ was proved to cause the inhibition of monoamine oxidase (MAO), especially MAO-B. These results suggest that the acute biochemical changes induced by MPP⁺ in vivo were MAO inhibition and release of DA.     

(+)MK-801 prevents the DDC-induced enhancement of MPTP toxicity in mice

In order to reach deeper insight into the mechanism of diethyldithiocarbamate (DDC)-induced enhancement of MPTP toxicity in mice, MK-801, a non-competitive antagonist of NMDA receptors, has been used as a tool to study the role of excitatory amino acids. In agreement with previous reports, (+)MK-801 did not significantly affect either striatal dopamine (DA) or tyrosine-hydroxylase (TH) activity in MPTP-treated animals. On the contrary (+)MK-801, but not (−)MK-801 significantly reduced the DDC + MPTP-induced fall in striatal DA and TH activity. A similar preventing effect on DA metabolites (DOPAC and HVA) and HVA/DA ratio was observed. The number of TH⁺Mneurons in the substantia nigra (SN) of (+)MK-801-pretreated mice was not significantly different from that of control animals, indicating that this treatment specifically antagonized the extensive DDC-induced lesion of dopaminergic cell bodies in this brain area. (+)MK-801 treatment did not affect the DDC-induced changes of striatal MPP⁺ levels, suggesting that the observed antagonism of MK-801 against DDC is not due to MPP⁺ kinetic modifications. Pretreatment with the MAO-B inhibitor,l-deprenyl, or with the DA uptake blocker, GBR 12909, completely prevented the marked DA depletion elicited by DDC + MPTP within the striatum. Both treatments also protected from the fall in DA metabolites and TH activity as well. This indicates that DDC-induced potentiation is dependent upon MPP⁺ production and its uptake by the dopaminergic nerve terminals. All these findings suggest that NMDA receptors play a crucial role in the DDC-induced enhancement of MPTP toxicity.     

Time course and morphology of dopaminergic neuronal death caused by the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine

Mechanisms responsible for 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced dopamine (DA) neuronal death remain unknown and in mice it is even unclear whether neuronal death does occur. In vitro studies suggest that 1-methyl-4-phenylpyridinium ion (MPP⁺), the active metabolite of MPTP, kills neurons by apoptosis. Herein, we investigated whether MPTP induces DA neuronal death in vivo in mice and whether the mechanism is that of apoptosis. C57/bl Mice received different doses of MPTP administered in four intraperitoneal injections every 2 hours and were sacrificed at different time points for analyses of tyrosine hydroxylase (TH) immunohistochemistry, silver staining, and Nissl staining within the mesencephalon. We found that MPTP induces neuronal destruction in the substantia nigra pars compacta (SNpc) and the ventral tegmental area (VTA). The active phase of degeneration began at 12 h postinjection and continued up to 4 days. During this period, there was a greater decrease in TH-defined neurons than in Nissl-stained neurons suggesting that MPTP can cause a loss in TH without necessarily destroying the neuron. Thereafter, neuronal counts by both techniques equalized and there was no further loss of DA neurons. Dying neurons showed shrunken eosinophilic cytoplasm and shrunken darkly stained nuclei. Double staining revealed degenerating neurons solely among TH positive neurons of SNpc and VTA. At no time point and at no dose of MPTP was apoptosis observed. In addition, in situ labelling revealed no evidence of DNA fragmentation. This study demonstrates that the MPTP mouse model replicates several key features of neurodegeneration of DA neurons in PD and provides no in vivo evidence that, using this specific paradigm of injection, MPTP kills DA neurons by apoptosis.     

Intracerebroventricular administration of 1-methyl-4-phenylpyridinium ion in mice: Effects of simultaneously administered nomifensine,deprenyl, and 1-t-butyl-4,4-diphenylpiperidine

Summary 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) destroys nigrostriatal dopaminergic pathways and thereby produces a syndrome similar to Parkinson's disease. MPTP is oxidized by monoamine oxidase B (MAO B) to the 1-methyl-4-phenylpyridinium ion (MPP⁺), which is taken up in dopaminergic neurons through the dopamine (DA) uptake system, where it develops its toxic effect. Our observations show a new aspect of the MPP⁺ mode of action, in which deprenyl in mice has a partially protective effect against MPP⁺. Furthermore budipine, a therapeutic agent for Parkinsonism, is also able to partially prevent MPP⁺ toxicity. A MAO B-inhibitory component of budipine, as shown in receptor binding studies previously, could contribute to this effect. Comparable experiments with nomifensine do not exclude the possibility of budipine as an effect as a DA uptake inhibitor. An unexplained after effect of budipine leads to a large increase in 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) levels five weeks after the last administration.     

Pharmacological effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) on striatal dopamine receptor system

In mice, chronic administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) produces an increase in the maximum number of [³H]spiperone binding sites in the striatum. The sensitivity of striatal protein phosphorylation to calcium plus calmodulin is also potentiated in MPTP-treated mice. These observations are associated with an enhancement of apomorphine-induced climbing behavior in the drug-treated animals. The results of this study suggest that in an animal model for Parkinson's disease, MPTP interrupts the dopamine (DA) transmission by chemically denervating the nigrostriatal neurons and through a compensatory mechanism, it increases the number of DA receptors as well as the sensitivity of protein phosphorylation to calcium plus calmodulin in mouse striatum. The latter two events may contribute to the development of DA receptor supersensitivity.     

7-Nitroindazole prevents 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced ATP loss in the mouse striatum

The neurotoxicity of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is dependent upon the MAO-B (monoamine oxidase type B)-catalyzed production of 1-methyl-4-phenylpyridinium ion (MPP(+)) and is likely to involve a perturbation of energy metabolism. Protection against MPTP neurotoxicity has been shown by treating mice with 7-nitroindazole (7-NI), a reversible inhibitor of both MAO-B and neuronal nitric oxide synthase (nNOS) activity. The objective of the present study was to evaluate (i) the relationship between the neuroprotective effect of 7-NI and MPTP-induced energy deficiency, and (ii) the role of nitric oxide production as a potential mechanism for energy perturbation after MPTP exposure. Maximum protection against striatal dopamine depletion and nigral neuronal loss was achieved when 7-NI (50 mg/kg, i.p.) was administered to C57BL/6 mice immediately before and after MPTP (50 mg/kg, s.c.). This short-term regimen of 7-NI administration parallels the time when MPTP exposure causes energy failure. 7-NI also completely prevented the loss of striatal ATP that occurs in mice during the initial hours after MPTP administration. In contrast, N(G)-nitro-L-arginine (two injections of 50 mg/kg each, given i.p. 20 and 4 h prior to MPTP), another NOS inhibitor, failed to affect MPTP-induced ATP depletion. Taken together, data indicate that (i) a temporal and causal relationship exists between the neuroprotective effect of 7-NI and its ability to counteract ATP reduction, and (ii) MAO-B rather than NOS inhibition is the mechanism by which 7-NI counteracts MPTP-induced ATP depletion.     

Effects of monoamine oxidase inhibitors on the diethyldithiocarbamate-induced enhancement of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine toxicity in C57BL/6 mice

Summary. Diethyldithiocarbamate (DDC) is known to potentiate the neurotoxicity of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The aims of the present study were to provide biochemical, pathological and behavioral evidence for the degeneration of dopamine (DA) neurons in C57BL/6 strain mice treated simultaneously with DDC and MPTP, and to evaluate the effects of monoamine oxidase (MAO) inhibitors on DDC-enhanced MPTP toxicity. DDC (400mg/kg)+ MPTP (30mg/kg) treatment decreased significantly the levels of striatal DA and its metabolites and induced bradykinesia. In mice treated with DDC+MPTP, degenerative areas were found in striatum, substantia nigra and tuberculum olfactorium by assessment of the binding of [¹²⁵I]RTI-121, a DA transporter ligand. Pretreatment with a MAO-B inhibitor selegiline prior to the administration of DDC and MPTP completely inhibited the decrease in the levels of DA and its metabolites, bradykinesia and degeneration of dopaminergic nerve terminals. In contrast, the protective action of clorgyline was not clearly observed in this model system.Received December 9, 2002; accepted March 12, 2003 Published online June 10, 2003     

Neuro-protective effects of bee venom by suppression of neuroinflammatory responses in a mouse model of Parkinson’s disease: Role of regulatory T cells

In the present study, we sought to determine whether bee venom (BV) promotes the survival of dopaminergic (DA) neurons in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of Parkinson’s disease (PD). Treatment with BV prevented degeneration of DA neurons in the substantia nigra (SN). This neuro-protective effect of BV was associated with microglial deactivation and reduction of CD4 T cell infiltration. Additionally, BV treatment significantly increased the proportion of CD4⁺CD25⁺Foxp3⁺ Tregs in vivo and in vitro. The increased proportion of Tregs by BV treatment remained suppressive ex vivo. Interestingly, BV treatment did not prevent MPTP neurotoxicity in mice depleted of Tregs by anti-CD25 antibody injection. Therefore, our present studies suggest that modulation of peripheral immune tolerance by Treg may contribute to the neuroprotective effect of BV in the MPTP model of Parkinson’s disease.     

Zonisamide for the treatment of Parkinson's disease

Zonisamide is an antiepileptic drug widely used to treat seizures worldwide. In addition to epilepsy, zonisamide may have beneficial efficacy in various neurological or psychiatric diseases. This article reviews the structure, mechanism of action, pharmacokinetics and possible antiparkinsonian action of zonisamide. A multicentered, randomized, double-blind, placebo-controlled study conducted in Japan provided data suggesting that zonisamide, as an add-on treatment, has efficacy in treating motor symptoms in patients with Parkinson's disease (PD). Zonisamide may be effective in reducing the duration of 'off' time in patients with PD treated with L-DOPA. The therapeutic doses of zonisamide for the treatment of PD are 50-100 mg/day, considerably lower than those for the treatment of epilepsy (200-400 mg/day). It is expected that zonisamide will be safe and tolerated in patients with PD, as it has been used as an antiepileptic for more than 15 years; however, further studies are required to evaluate its safety and tolerability in the treatment of PD. The pharmacological mechanisms of the beneficial actions of zonisamide in PD remain unclear. Various hypotheses have been proposed, but the supporting data are not yet sufficient to draw any conclusions. Further basic research is required to advance our understanding of the antiparkinsonian mechanism of zonisamide.     

Transplacentally-transported 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) affects the catecholamine and indoleamine levels in the fetal mouse brain

Summary The effects of a dopaminergic neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) on the amounts of dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) were examined in the whole brains of fetal mice and maternal mice after its administration to pregnant mice. DA and DOPAC concentrations were decreased significantly in both the fetal and maternal brains. At 3 hr after injection, reduction of the DOPAC concentration was more marked than that of DA in both the fetal and maternal brains. Increase of 5-HT concentration was observed until 12 hr after injection in the fetal brains and 6 hr in the maternal brains. These results indicate that 1-methyl-4-phenyl-pyridinium ion (MPP⁺) and MPTP affect the levels of catechol- and indoleamines in the brain of premature stage as well as in the mature brain.     

1,2,3,4-Tetrahydro-2-methyl-4,6,7-isoquinolinetriol inhibits tyrosine hydroxylase activity in rat striatal synaptosomes

Summary 1,2,3,4-tetrahydro-2-methyl-4,6,7-isoquinolinetriol (TMIQ), a tetrahydroisoquinoline derivate of adrenaline, was tested for potency as an analog of the dopamine depleting agent 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in assays of tyrosine hydroxylase (TH) activity in the striatal synaptosome preparation. TMIQ inhibited TH activity with an IC₅₀ (4 × 10^–6M) similar to that found for MPTP (IC₅₀ 1 × 10^–6M). TH inhibitions produced by IC₅₀ concentrations of TMIQ were reversed by monoamine oxidase (MAO)-A or MAO-B inhibitors (clorgyline or deprenyl), or the dopamine reuptake blocker nomifensine, or excess cofactor (6R)-5,6,7,8-tetrahydro-L-biopterin. TMIQ did not appear to act at the presynaptic D₂ sulpiride sensitive autoreceptor for dopamine synthesis modulation. Thesein vitro data are consistent with earlier findings that TMIQ acts as a dopamine depleting agent, and with the possibility that TMIQ may have a degree of MPTP-like activityin vivo.     

The interaction of 1-alkyl-4, 4-diphenylpiperidines with the 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine receptor binding site

Summary 1-Methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) is a selective neurotoxin which produces degeneration of the nigrostriatal bundles in the central nervous system of man and animals. In these areas of the brain are concentrated the receptor binding sites for [³H]MPTP. 1-Alkyl-4, 4-diphenylpiperidines displace [³H]MPTP from these binding sites with K₁ values in the micromolar range. The t-butyl analogue in this class of substances, budipine, is a novel therapeutic agent for Parkinsonism whose mechanism has not yet been fully clarified. The affinity of budipine for the MPTP receptor binding site was determined as a K₁ value of 2.2M. Other 4, 4-diphenylpiperidine derivatives such as 1-methyl-4, 4-diphenylpiperidine and 1-i-propyl-4, 4-diphenylpiperidine have substantially lower affinities. Monoamine oxidase inhibitors such as deprenyl, pargyline and harmaline have affinities to the MPTP receptors which parallel their affinity for the B type of monoamine oxidase (MAO B). This supports the theory that the MPTP receptor binding sites is identical with membrane bound MAO B.     

Dopamine-releasing action of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and 1-methyl-4-phenylpyridine (MPP) in the neostriatum of the rat as demonstrated in vivo by the push-pull perfusion technique: dependence on sodium but not calcium ions

This study examined the effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and its metabolite, 1-methyl-4-phenylpyridine (MPP+) on the levels of dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) in push-pull perfusates of the striatum in chloral hydrate-anaesthetized rats. In control animals the levels of DA and DOPAC remained stable for at least 6 h and responded rapidly to a depolarizing stimulus of 25 mM K+. This K+-induced DA release was Ca2+-dependent since no stimulation was observed when the striatal sites were perfused with high K+ in a Ca2+-free medium containing 2 mM EGTA thus verifying that the striatal sites were functionally active. MPTP (0.025 and 0.05 microgram/microliter) stimulated DA release and inhibited DOPAC output in a dose-related manner. MPP+ (0.01, 0.025 and 0.05 microgram/microliter) produced a more robust dose-dependent increase in DA levels in the perfusates; however, the level of suppression of DOPAC was similar to that in response to MPTP. The effect of MPP+ on DA release was attenuated by 10(-6) M benztropine, the DA re-uptake blocker and completely inhibited by 10 micrograms/kg i.p. benztropine and 10(-4) M ouabain, the Na+, K+-ATPase (Na pump) inhibitor. However, although these substances prevented the MPP+-induced release of DA, the levels of DOPAC in the perfusates did not recover and remained completely suppressed suggesting that MPP+ may inhibit extraneuronal rather than intraneuronal monoamine oxidase (MAO). Perfusion of the striatal sites with a Ca2+-free medium containing 2 mM EGTA did not prevent the MPP+-induced DA release indicating that MPP+ does not release DA from the striatal DA terminals by the Ca2+-dependent process of exocytosis. The responses of DA and DOPAC to 25 mM K+ were markedly suppressed in animals treated with MPTP and MPP+, these effects being most severe with the highest dose of MPP+. Moreover, this suppression of the K+-induced responses persisted in animals perfused with MPP+ in the presence of benztropine or ouabain, thus suggesting that MPP+ may have potent deleterious membrane effects. These studies have provided the first direct in vivo demonstration of the action of MPTP and MPP+ and the neuropharmacological basis of this action on DA metabolism in the rat striatum. The results show that the elevated levels of DA in the striatal perfusates are due to a direct action of MPTP and MPP+ on the nigrostriatal DA terminals and cannot be fully accounted for solely by their inhibition of MAO activity and/or inhibition of DA re-uptake.(ABSTRACT TRUNCATED AT 250 WORDS)     

Neuroprotective effect of thalidomide on MPTP-induced toxicity

Thalidomide is a sedative with unique pharmacological properties; studies on epilepsy and brain ischemia have shown intense neuroprotective effects. We analyzed the effect of thalidomide treatment on the neurotoxicity caused by the administration of 1-methyl-4-phenyl-1,2,3,6-tetrahidropyridine (MPTP) in mice. Thalidomide was administered at two times; before and after the exposure to MPTP. In both circumstances thalidomide improved the neurotoxicity induced by MPTP as seen by a significant raise of the striatal contents of dopamine and simultaneous decrease of monoamine-oxidase-B (MAO-B). These results indicate that in the experimental model of Parkinson's disease the administration of thalidomide improves the functional damage on the nigrostriatal cell substratum as seen by the production of dopamine. This neuroprotective effect seems to be mediated by inhibition of excitotoxicity. Our results suggest that thalidomide could be investigated as potential adjuvant therapy for Parkinson's disease.     

Dopamine Terminal Loss and Onset of Motor Symptoms in MPTP-Treated Monkeys: A Positron Emission Tomography Study with 11C-CFT

We studied the time course of dopamine (DA) terminal loss in three macaca fascicularis injected with MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) intravenously every 10-14 days for up to 389 days. Striatal DA terminal loss was monitored in vivo by positron emission tomography using ¹¹ C-CFT (WIN 35,428), a cocaine derivative that labels the DA transporter. The ¹¹C-CFT uptake rate constant in the striatum of MPTP-treated monkeys decreased exponentially over time, with the putamen significantly more affected than the caudate. Spontaneous locomotor activity decreased in parallel with the decline of the ¹¹C-CFT uptake rate; however, overt parkinsonian signs appeared only after the ¹¹C-CFT uptake rate had declined to about 30% of the pretreatment values. We conclude that a long-term intermittent mode of administration of MPTP can lead to a pattern of terminal loss that closely resembles idiopathic Parkinson disease.     

Melatonin fails to protect against long-term MPTP-induced dopamine depletion in mouse striatum

Several laboratories recently have reported that melatonin may possess neuroprotective properties. The present paper presents the results of our studies on the long term in vivo neuroprotective effects of melatonin in a well-defined neurotoxicity model using 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in the C57BL/6 mouse. MPTP is bioactivated by brain monoamine oxidase B (MAO-B) to its neurotoxic pyridinium metabolite 1-methyl-4-phenylpyridinium (MPP(+)) which destroys dopaminergic nerve terminals leading to the depletion of neostriatal dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC). Our initial study compared striatal DA and DOPAC levels in MPTP-only-treated animals and animals treated with melatonin 30 min prior to and 3 times hourly post-MPTP. DA/DOPAC levels measured 7 days after MPTP were similar in both groups. A second study was designed to address the possibility that melatonin cleared from the brain prior to MPP(+). Animals, that had been administered the same regimen of melatonin as in the first study plus a fourth post-MPTP melatonin dose, were maintained on melatonin in drinking water until 5 days post-MPTP. Striatal DA/DOPAC levels of these melatonin-plus-MPTP treated animals also were the same as the MPTP-only-treated animals. In vitro studies confirmed that melatonin is not an inhibitor of MAO-B. These data demonstrate that melatonin does not have any significant protective effects against the long-term striatal DA and DOPAC depletion induced by MPTP in the C57BL/6 mouse.     

Nitric oxide and MPP+-induced hydroxyl radical generation

Summary. Although neuroprotective effect of nitric oxide (NO) is discussed, NO has a role of pathogenesis of cellular injury. NO is synthesized from L-arginine by NO synthase (NOS). NO contributes to the extracellular potassium-ion concentration ([K⁺]_o)-induced hydroxyl radical (^•OH) generation. Cytotoxic free radicals such as peroxinitrite (ONOO⁻) and ^•OH may also be implicated in NO-mediated cell injury. NO activation was induced by K⁺ depolarization. NO may react with superoxide anion (O₂ ⁻) to form ONOO⁻ and its decomposition generates ^•OH. 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) metabolite 1-methyl-4-phenylpyridinium ion (MPP⁺) involve toxicity induced by NO. Intraneuronal Ca²⁺ triggered by MPP⁺ may be detrimental to the functioning of dopaminergic nerve terminals in the striatum. Although the [K⁺]_o-induced depolarization enhances the formation of ^•OH product due to MPP⁺, the ^•OH generation via NOS activation may be unrelated the dopamine (DA)-induced ^•OH generation. Depolarization enhances the MPP⁺-induced ^•OH formation via NOS activation. NOS inhibition is associated with a protective effect due to suppression of depolarization-induced ^•OH generation. ONOO⁻ has been implicated as a causative factor under conditions in which DA neurons are damaged. These findings may be useful in elucidating the actual mechanism of free radical formation in the pathogenesis of neurodegenerative brain disorders, including Parkinson’s disease and traumatic brain injuries.