Analysis of the MCTP Amino Acid Sequence Reveals the Conservation of Putative Calcium- and Lipid-Binding Pockets Within the C2 Domains In Silico.

MCTPs (Multiple C2 Domains and Transmembrane region Proteins) are evolutionarily and structurally related to other C2 proteins, which are central to exocytosis and membrane trafficking; however, their specific function has been little studied. MCTPs are associated with endosomes and the endoplasmic reticulum and possess three C2 domains (C2A-C2C) and two transmembrane regions (TMRs) well conserved in different species. Here, we generated structural models of the MCTP C2 domains of C. elegans and analyzed their putative function by docking, which revealed that these domains possess Ca2+- and lipid-binding pockets, suggesting that MCTPs play a significant, calcium-dependent role in membrane physiology.

MCTP-1 modulates neurotransmitter release in C. elegans.

Multiple C2 and Transmembrane Domain Proteins (MCTPs) are putative calcium sensors. Proteins that contain C2 domains play essential roles in membrane trafficking and exocytosis; however, MCTPs functions in neurotransmitter release are not known. Here we report that in C. elegans mctp-1 is under the control of two promoters - one active in the nervous system and the second in the spermatheca. We generated and characterized a loss of function amt1 mutant and compared it to a previously published loss of function mutant (av112). Loss of mctp-1 function causes defects in egg-laying, crawling velocity, and thrashing rates. Both amt1 and av112 mutants are hyposensitive to the acetylcholinesterase blocker aldicarb, suggesting that MCTP-1 may play a role in synaptic vesicle release.

MiR-302b-5p enhances the neuroprotective effect of IGF-1 in methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinson's disease by regulating inducible nitric-oxide synthase.

Parkinson's disease (PD) is a neurodegenerative disease which results in damage in neuronal cells. Insulin-like growth factor (IGF)-1 was previously reported to play a role of neuroprotection in some diseases. Nitric oxide (NO) can also regulate neuronal cells. However, the mechanisms underlying IGF-1 and NO in PD still need to be elucidated. In present study, we explored the interaction between IGF-1 and inducible Nitric-Oxide Synthase (iNOS) in PD progression. We firstly constructed PD models by methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) or MPP+ treatment. Then RT-qPCR revealed that IGF-1 expression was downregulated while iNOS expression was upregulated in MPTP model. Moreover, IGF-1 elevation or iNOS depletion enhanced cell viability and blocked cell apoptosis. Rescue assay disclosed iNOS overexpression reversed the effect on viability and apoptosis mediated by IGF-1 upregulation. Furthermore, IGF-1 was identified to positively regulate miR-302b-5p which could target iNOS. MiR-302b-5p could abolish the inhibitory function IGF-1 exerted on cell apoptosis and iNOS could counteract miR-302b-5p upregulation-triggered inhibition on cell apoptosis as well. Besides, we observed the deficiency of miR-302b-5p improved the lesioned neurobehavior of MPTP-treated mice. To sum up, present study proved that miR-302b-5p enhanced the neuroprotective effect of IGF-1 in MPTP-induced PD by regulating iNOS, recommending a novel therapeutic target for PD treatment. SIGNIFICANCE OF THE STUDY: In this study, we mainly explored that IGF-1 was decreased while iNOS was boosted in MPTP-induced PD mice model; IGF-1 suppressed while iNOS promoted MPP+ -induced toxicity and apoptosis in SH-SY5Y cells; miR-302b-5p ehanhced the neuroprotective effect of IGF-1 via targeting Inos; deficiency of miR-302b-5p improved the lesioned neurobehavior of MPTP-treated mice.

Vector-mediated l-3,4-dihydroxyphenylalanine delivery reverses motor impairments in a primate model of Parkinson's disease.

Ever since its introduction 40 years ago l-3,4-dihydroxyphenylalanine (l-DOPA) therapy has retained its role as the leading standard medication for patients with Parkinson's disease. With time, however, the shortcomings of oral l-DOPA treatment have become apparent, particularly the motor fluctuations and troublesome dyskinetic side effects. These side effects, which are caused by the excessive swings in striatal dopamine caused by intermittent oral delivery, can be avoided by delivering l-DOPA in a more continuous manner. Local gene delivery of the l-DOPA synthesizing enzymes, tyrosine hydroxylase and guanosine-tri-phosphate-cyclohydrolase-1, offers a new approach to a more refined dopaminergic therapy where l-DOPA is delivered continuously at the site where it is needed i.e. the striatum. In this study we have explored the therapeutic efficacy of adeno-associated viral vector-mediated l-DOPA delivery to the putamen in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated rhesus monkeys, the standard non-human primate model of Parkinson's disease. Viral vector delivery of the two enzymes, tyrosine hydroxylase and guanosine-5'-tri-phosphate-cyclohydrolase-1, bilaterally into the dopamine-depleted putamen, induced a significant, dose-dependent improvement of motor behaviour up to a level identical to that obtained with the optimal dose of peripheral l-DOPA. Importantly, this improvement in motor function was obtained without any adverse dyskinetic effects. These results provide proof-of-principle for continuous vector-mediated l-DOPA synthesis as a novel therapeutic strategy for Parkinson's disease. The constant, local supply of l-DOPA obtained with this approach holds promise as an efficient one-time treatment that can provide long-lasting clinical improvement and at the same time prevent the appearance of motor fluctuations and dyskinetic side effects associated with standard oral dopaminergic medication.

Genetic suppression of collapsin response mediator protein 2 phosphorylation improves outcome in methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinson's model mice.

Parkinson's disease (PD) is a common neurodegenerative disorder characterized by slow and progressive degeneration of dopaminergic neurons in the substantia nigra pars compacta (SNc). Levodopa (l-Dopa), the current main treatment for PD, supplies dopamine, but it does not prevent neurodegeneration. There is thus no promising remedy for PD. Recent in vitro study showed the increase in the phosphorylation levels of Collapsin Response Mediator Protein 2 (CRMP2) is involved in dopaminergic axon degeneration. In the present study, we report elevation of CRMP2 phosphorylation in dopaminergic neurons in SNc after challenge with the dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a common model for PD. Genetic suppression of CRMP2 phosphorylation by mutation of the obligatory Cyclin-dependent kinase 5 (Cdk5)-targeted serine-522 site prevented axonal degradation in the nigrostriatal pathway of transgenic mice. As a result, the degree of MPTP-induced motor impairment in the rotarod test was suppressed. These results suggest that suppression of CRMP2 phosphorylation may be a novel therapeutic target for PD.

Intestinal Pathology and Gut Microbiota Alterations in a Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) Mouse Model of Parkinson's Disease.

Patients with Parkinson's disease (PD) often have non-motor symptoms related to gastrointestinal (GI) dysfunction, such as constipation and delayed gastric emptying, which manifest prior to the motor symptoms of PD. Increasing evidence indicates that changes in the composition of the gut microbiota may be related to the pathogenesis of PD. However, it is unclear how GI dysfunction occurs and how gut microbial dysbiosis is caused. We investigated whether a neurotoxin model of PD induced by chronic low doses of MPTP is capable of reproducing the clinical intestinal pathology of PD, as well as whether gut microbial dysbiosis accompanies this pathology. C57BL/6 male mice were administered 18 mg/kg MPTP twice per week for 5 weeks via intraperitoneal injection. GI function was assessed by measuring the 1-h stool frequency and fecal water content; motor function was assessed by pole tests; and tyrosine hydroxylase and alpha-synuclein expression were analyzed. Furthermore, the inflammation, intestinal barrier and composition of the gut microbiota were measured. We found that MPTP caused GI dysfunction and intestinal pathology prior to motor dysfunction. The composition of the gut microbiota was changed; in particular, the change in the abundance of Lachnospiraceae, Erysipelotrichaceae, Prevotellaceae, Clostridiales, Erysipelotrichales and Proteobacteria was significant. These results indicate that a chronic low-dose MPTP model can be used to evaluate the progression of intestinal pathology and gut microbiota dysbiosis in the early stage of PD, which may provide new insights into the pathogenesis of PD.

Behavioral, Biochemical and Molecular Characterization of a Parkinson's Disease Mouse Model Using the Neurotoxin 2'-CH3-MPTP: A Novel Approach.

The neurotoxin MPTP has long been used to create a mouse model of Parkinson's disease (PD). Indeed, several MPTP analogues have been developed, including 2'-CH3-MPTP, which was shown to induce nigrostriatal DA neuronal depletion more potently than MPTP. However, no study on behavioral and molecular alterations in response to 2'-CH3-MPTP has been carried out so far. In the present work, 2'-CH3-MPTP was administered to mice (2.5, 5.0 and 10 mg/kg per injection, once a day, 5 days) and histological, biochemical, molecular and behavioral alterations were evaluated. We show that, despite a dose-dependent-like pattern observed for nigrostriatal dopaminergic neuronal death and dopamine depletion, dose-specific alterations in dopamine metabolism and in the expression of dopaminergic neurotransmission-associated genes could be related to specific motor deficits elicited by the different doses tested. Interestingly, 2'-CH3-MPTP leads to increased DAT and MAO-B transcription, which could explain, respectively, its higher potency and the requirement of higher doses of MAO inhibitors to prevent nigrostriatal neuronal death when compared to MPTP. Also, perturbations in dopamine metabolism as well as possible alterations in dopamine bioavailability in the synaptic cleft were also identified and correlated with strength and ambulation deficits in response to specific doses. Overall, the present work brings new evidence supporting the distinct effects of 2'-CH3-MPTP when compared to its analogue MPTP. Moreover, our data highlight the utmost importance of a precise experimental design, as different administration regimens and doses yield different biochemical, molecular and behavioral alterations, which can be explored to study specific aspects of PD.

Activation and deactivation of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine by cytochrome P450 enzymes and flavin-containing monooxygenases in common marmosets (Callithrix jacchus).

The potential proneurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induces Parkinson-like syndromes in common marmosets, other primates, and humans. MPTP is metabolically activated to 1-methyl-4-phenyl-2,3-dihydropyridinium and 1-methyl-4-phenylpyridinium ions (MPDP(+) and MPP(+), respectively) by desaturation reactions. MPTP is deactivated to 4-phenyl-1,2,3,6-tetrahydropyridine (PTP) by N-demethylation and is also deactivated to MPTP N-oxide. The roles of cytochrome P450 (P450) enzymes and flavin-containing monooxygenases (FMOs) in the oxidative metabolism of MPTP-treated marmosets are not yet fully clarified. This study aimed to elucidate P450- and FMO-dependent MPTP metabolism in marmoset liver and brain. Rates of MPTP N-oxygenation in liver microsomes were similar to those in brain microsomes from 11 individual marmosets (substrate concentration, 50 µM) and were correlated with rates of benzydamine N-oxygenation (r = 0.75, P < 0.05); the reactions were inhibited by methimazole (10 µM). MPTP N-oxygenation was efficiently mediated by recombinantly expressed marmoset FMO3. Rates of PTP formation by MPTP N-demethylation in marmoset liver microsomes were correlated with bufuralol 1'-hydroxylation rates (r = 0.77, P < 0.01) and were suppressed by quinidine (1 µM), thereby indicating the importance of marmoset CYP2D6 in PTP formation. MPTP transformations to MPDP(+) and MPP(+) were efficiently catalyzed by recombinant marmoset CYP2D6 and human CYP1A2. These results indicated the contributions of multiple drug-metabolizing enzymes to MPTP oxidation, especially marmoset FMO3 in deactivation (N-oxygenation) and marmoset CYP2D6 for both MPTP deactivation and MPTP activation to MPDP(+) and MPP(+). These findings provide a foundation for understanding MPTP metabolism and for the successful production of preclinical marmoset models.

Design and synthesis of a MAO-B-selectively activated prodrug based on MPTP: a mitochondria-targeting chemotherapeutic agent for treatment of human malignant gliomas.

Malignant gliomas, including glioblastomas, are extremely difficult to treat. The median survival for glioblastoma patients with optimal therapeutic intervention is 15 months. We developed a novel MAO-B-selectively activated prodrug, N,N-bis(2-chloroethyl)-2-(1-methyl-1,2,3,6-tetrahydropyridin-4-yl)propanamide (MP-MUS), for the treatment of gliomas based on 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The design of neutral MP-MUS involved the use of a seeker molecule capable of binding to mitochondrial MAO-B, which is up-regulated ≥fourfold in glioma cells. Once the binding occurs, MP-MUS is converted into a positively charged moiety, P(+) -MUS, which accumulates inside mitochondria at a theoretical maximal value of 1000:1 gradient. The LD50 of MP-MUS against glioma cells is 75 µM, which is two- to threefold more potent than temozolomide, a primary drug for gliomas. Importantly, MP-MUS was found to be selectively toxic toward glioma cells. In the concentration range of 150-180 µM MP-MUS killed 90-95 % of glioma cells, but stimulated the growth of normal human astrocytes. Moreover, maturation of MP-MUS is highly dependent on MAO-B, and inhibition of MAO-B activity with selegiline protected human glioma cells from apoptosis.

New allometric scaling relationships and applications for dose and toxicity extrapolation.

Allometric scaling between metabolic rate, size, body temperature, and other biological traits has found broad applications in ecology, physiology, and particularly in toxicology and pharmacology. Basal metabolic rate (BMR) was observed to scale with body size and temperature. However, the mass scaling exponent was increasingly debated whether it should be 2/3, 3/4, or neither, and scaling with body temperature also attracted recent attention. Based on thermodynamic principles, this work reports 2 new scaling relationships between BMR, size, temperature, and biological time. Good correlations were found with the new scaling relationships, and no universal scaling exponent can be obtained. The new scaling relationships were successfully validated with external toxicological and pharmacological studies. Results also demonstrated that individual extrapolation models can be built to obtain scaling exponent specific to the interested group, which can be practically applied for dose and toxicity extrapolations.

The role of thioredoxin-1 in suppression of endoplasmic reticulum stress in Parkinson disease.

Endoplasmic reticulum (ER) stress has been implicated in Parkinson disease. We previously reported that thioredoxin 1 (Trx-1) suppressed the ER stress caused by 1-methy-4-phenyl-1,2,3,6-tetrahydropyridine; however, its molecular mechanism remains largely unknown. In the present study, we showed that 1-methyl-4-phenylpyridinium ion (MPP(+)) induced ER stress by activating glucose-regulated protein 78 (GRP78), inositol-requiring enzyme 1α (IRE1α), tumor necrosis factor receptor-associated factor 2 (TRAF2), c-Jun N-terminal kinase (JNK), caspase-12, and C/EBP homologous protein (CHOP) in PC12 cells. The downregulation of Trx-1 aggravated the ER stress and further increased the expression of the above molecules induced by MPP(+). In contrast, overexpression of Trx-1 attenuated the ER stress and repressed the expression of the above molecules induced by MPP(+). More importantly, the overexpression of Trx-1 in transgenic mice suppressed ER stress by inhibiting the activation of these molecules. We present, for the first time, the molecular mechanism of Trx-1 suppression of endoplasmic reticulum stress in Parkinson disease in vitro and in vivo. Based on our findings, we conclude that Trx-1 plays a neuroprotective role in Parkinson disease by suppressing ER stress by regulating the activation of GRP78, IRE1α, TRAF2, JNK, caspase-12, and CHOP.

L-Dihydroxyphenylalanine modulates the steady-state expression of mouse striatal tyrosine hydroxylase, aromatic L-amino acid decarboxylase, dopamine and its metabolites in an MPTP mouse model of Parkinson's disease.

AIMS: l-3,4-Dihydroxyphenylalanine (L-DOPA) is the most effective symptomatic treatment for Parkinson's disease (PD), but PD patients usually experience a successful response to L-DOPA therapy followed by a progressive loss of response. L-DOPA efficacy relies on its decarboxylation by aromatic l-amino acid decarboxylase (AAAD) to form dopamine (DA). So exogenous L-DOPA drives the reaction and AAAD becomes the rate limiting enzyme in the supply of DA. In turn, exogenous L-DOPA regulates the expression and activity of AAAD as well as the synthesis of DA and its metabolites, changes that may be linked to the efficacy and side-effects of L-DOPA. MAIN METHODS: One-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse PD model was utilized to study the effects of L-DOPA on the steady-state level and activity of AAAD, tyrosine hydroxylase (TH), DA and the metabolites of DA. The MPTP and control mice were treated twice daily with PBS or with 100mg/kg of L-DOPA for 14days and the expression and activity of AAAD, the expression of TH and the levels of DA and its metabolites were determined 24h after L-DOPA or PBS treatment, when exogenous L-DOPA is eliminated. KEY FINDINGS: In the MPTP model, L-DOPA reduced the steady-state expression and the activity of striatal AAAD by 52% and 50%, respectively, DA and metabolites were also significantly decreased. SIGNIFICANCE: The outcome shows that while L-DOPA replenishes striatal DA it also down-regulates AAAD and the steady-state synthesis and metabolic capability of the dopaminergic system. These findings are important in the precipitation of L-DOPA induced side effects and the management of L-DOPA therapy.

Polygalae radix inhibits toxin-induced neuronal death in the Parkinson's disease models.

AIM OF THE STUDY: Polygalae radix, the root of Polygala tenuifolia Willd, has commonly been used for the treatment of amnesia and anxiety in traditional Korean medicine. The aim of this study was to investigate its neuroprotective effects and possible mechanisms of action in models of Parkinson's disease. MATERIALS AND METHODS: This study utilized a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, a reactive oxygen species (ROS) assay, a nitric oxide (NO) production assay, and a caspase-3 activity test as measures of cell viability in PC12 cells damaged by 6-hydroxydopamine (6-OHDA). The protective effects of PRE against 1-methyl-4-phenylpyridium (MPP(+))-induced neurotoxicity were assessed in rat primary dopaminergic neurons and in a mouse PD model in which PRE was administered (100mg/kg/day, 3 days, p.o.) before acute 1-mehtyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) toxicity. Finally, TH immunohistochemistry tests were conducted in the substantia nigra pars compacta (SNpc) and striatum (ST). RESULTS AND CONCLUSIONS: PRE significantly inhibited 6-OHDA-induced cell damage at doses of 0.05-1µg/ml with a maximal effect at 0.1µg/ml. Caspase-3 activity and the production of ROS and NO were alleviated at 0.1µg/ml. Also at this dose, PRE protected mesencephalic dopaminergic neurons from MPP(+)-induced toxicity. In an in vivo mouse model of PD, PRE protected dopaminergic neurons and fibers from MPTP-induced toxicity in the SNpc and ST. These results demonstrate that PRE has protective effects on dopaminergic neurons via its anti-oxidant and anti-apoptotic activity.

Imidazolium salt (DBZIM) reduces gliosis in mice treated with neurotoxicant 2'-CH(3) -MPTP.

We have recently identified a class of imidazolium salts (IMSs) with antioxidative property and can function as scavengers for radical oxygen species (ROS) [18]. Here, we investigate one of the IMSs, 1,3-bisbenzylimidazolium bromide (DBZIM), for its possible role in attenuating neurotoxicity and gliosis in the retina and the brain induced by a Parkinsonian neurtoxicant, methyl-4(2'-methylphenyl)-1,2,3,6-tetrahydropyridine (2'-CH(3) -MPTP), which is a free radical generating agent. In this study, we employ a molecular retinal imaging method, which we recently developed in a transgenic mouse model expressing green fluorescent protein (GFP) under the control of glial fibrillary acidic protein (GFAP) promoter [14], to assess the efficacy of DBZIM, since currently no in vitro system with a sufficient complexity is available for accurately assessing a compound's efficacy. The longitudinal imaging results showed DBZIM can effectively suppress the neurotoxicant-induced retinal gliosis. Immunohistochemistry performed on the postmodern mouse brain confirmed that DBZIM also reduced striatal gliosis, and concomitantly attenuated the loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc). These findings suggest that DBZIM could be a useful small molecular compound for studying neurotoxicity and neuroprotection in the retina and the brain.

Neuroprotective effects of an herbal medicine, Yi-Gan San on MPP+/MPTP-induced cytotoxicity in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: A traditional herb, Yi-Gan San, has been widely used for the management of neurodegenerative disorders in traditional East Asian Medicine. AIM OF THE STUDY: The present study investigated the neuroprotective effects of Yi-Gan San in 1-methyl-4-phenylpyridine/1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced cytotoxicity in vitro and in vivo and sought to clarify its underlying mechanisms. MATERIALS AND METHODS: The effect of Yi-Gan San on 1-methyl-4-phenylpyridine was measured in terms of 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assays, caspase-3 activity, and western blot analysis of phosphorylated Akt, one of the survival-related signaling proteins in SH-SY5Y cells. The effects of Yi-Gan San were also confirmed in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinsonian mouse model using a rotarod test and tyrosine hydroxylase-immunohistochemistry. RESULTS: Pretreatment of Yi-Gan San with 1-methyl-4-phenylpyridine showed a significant protective effect on SH-SY5Y cells and significantly decreased the level of caspase-3 activity compared to the values for the 1-methyl-4-phenylpyridine-treated cells. This process increased the protein expressions of phosphorylated Akt, and an inhibitor of phosphatidylinositol 3-kinase (PI3K)/Akt, LY294002, significantly decreased this protective effect of Yi-Gan San. In the mouse Parkinson's disease model, treatment with Yi-Gan San also significantly improved motor functioning and prevented dopaminergic loss related to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine challenge. CONCLUSION: Using both in vitro and in vivo methods, this study revealed that Yi-Gan San has neuroprotective effects and rescues dopaminergic neurons from 1-methyl-4-phenylpyridine/1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine toxicity, possibly via the PI3K/Akt pathway.

Protective effects of Chunghyuldan against ROS-mediated neuronal cell death in models of Parkinson's disease.

Previous reports have suggested that the herbal medicine Chunghyuldan (CHD, Qingxue-dan in Chinese and Daio-Orengedokuto in Japanese) has wide-ranging biological effects, including anti-hyperlipidaemic, anti-ischaemic, anti-inflammatory and antioxidant activities. Reactive oxygen species (ROS)-mediated mitochondrial dysfunction is thought to be one of the major pathological mechanisms responsible for Parkinson's disease (PD) and may underlie the selective loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc) that is a hallmark of this disease. In this study, we examined the neuroprotective effects of CHD in PD models produced by treatment with neurotoxins that act via ROS-mediated mitochondrial dysfunction. In an in vitro PD model using 6-hydroxydopamine, CHD applied at concentrations of 10 and 100 µg/ml exhibited significant protective effects in PC12 cells by inhibiting intracellular ROS generation. CHD applied at 10 and 100 µg/ml also prevented 6-hydroxydopamine-induced mitochondrial depolarization and elevation of caspase-3 activity. At the same doses, CHD showed regulatory effects on the haem oxygenase-1 and gp91 phagocytic oxidase which have critical roles in generating ROS. In addition, CHD protected dopaminergic neurons in a primary mesencephalic culture against MPP+ neurotoxicity. In an in vivo PD model produced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine treatment (20 mg/kg, 4 times, i.p.), co-administration of CHD (50 mg/kg, 5 days, p.o.) ameliorated PD-like behavioural symptoms (bradykinesia) and reduced dopaminergic neuronal damage in the SNpc and striatum as measured by immunocytochemistry. These results demonstrate the neuroprotective effects of CHD in PD models that are mediated through inhibition of ROS generation and associated mitochondrial dysfunction.

Hydrogen in drinking water reduces dopaminergic neuronal loss in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine mouse model of Parkinson's disease.

It has been shown that molecular hydrogen (H(2)) acts as a therapeutic antioxidant and suppresses brain injury by buffering the effects of oxidative stress. Chronic oxidative stress causes neurodegenerative diseases such as Parkinson's disease (PD). Here, we show that drinking H(2)-containing water significantly reduced the loss of dopaminergic neurons in PD model mice using both acute and chronic administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The concentration-dependency of H(2) showed that H(2) as low as 0.08 ppm had almost the same effect as saturated H(2) water (1.5 ppm). MPTP-induced accumulation of cellular 8-oxoguanine (8-oxoG), a marker of DNA damage, and 4-hydroxynonenal (4-HNE), a marker of lipid peroxidation were significantly decreased in the nigro-striatal dopaminergic pathway in mice drinking H(2)-containing water, whereas production of superoxide (O(2)*(-)) detected by intravascular injection of dihydroethidium (DHE) was not reduced significantly. Our results indicated that low concentration of H(2) in drinking water can reduce oxidative stress in the brain. Thus, drinking H(2)-containing water may be useful in daily life to prevent or minimize the risk of life style-related oxidative stress and neurodegeneration.

Cortical serotonin and norepinephrine denervation in parkinsonism: preferential loss of the beaded serotonin innervation.

Parkinson's Disease (PD) is marked by prominent motor symptoms that reflect striatal dopamine insufficiency. However, non-motor symptoms, including depression, are common in PD. It has been suggested that these changes reflect pathological involvement of non-dopaminergic systems. We examined regional changes in serotonin (5-HT) and norepinephrine (NE) systems in mice treated with two different 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treatment paradigms, at survival times of 3 or 16 weeks after the last MPTP injection. MPTP caused a decrease in striatal dopamine concentration, the magnitude of which depended on the treatment regimen and survival interval after MPTP treatment. There was significant involvement of other subcortical areas receiving a dopamine innervation, but no consistent changes in 5-HT or NE levels in subcortical sites. In contrast, we observed an enduring decrease in 5-HT and NE concentrations in both the somatosensory cortex and medial prefrontal cortex (PFC). Immunohistochemical studies also revealed a decrease in the density of PFC NE and 5-HT axons. The decrease in the cortical serotonergic innervation preferentially involved the thick beaded but not smooth fine 5-HT axons. Similar changes in the 5-HT innervation of post-mortem samples of the PFC from idiopathic PD cases were seen. Our findings point to a major loss of the 5-HT and NE innervations of the cortex in MPTP-induced parkinsonism, and suggest that loss of the beaded cortical 5-HT innervation is associated with a predisposition to the development of depression in PD.

Zocor Forte (simvastatin) has a neuroprotective effect against LPS striatal dopaminergic terminals injury, whereas against MPP+ does not.

Due to their potential role in preventing further deterioration of Parkinson's disease, anti-inflammatory strategies have attracted great interest. In this context, some studies point out the possible protective effect of anti-inflammatory compounds against the in vivo degeneration of dopaminergic neurons produced by lipopolysaccharide (LPS)-induced inflammatory processes and others. We have investigated the effect of the treatment of Zocor Forte (simvastatin) in LPS and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) neurodegenerative models to identify neuroprotective drugs for Parkinson's disease. We have perfused different concentrations of LPS or 1 mM 1-methyl-4-phenylpyridinium ion (MPP+) in the rat's striatum, 24 h after implanting a brain microdialysis probe, both with and without Zocor Forte (simvastatin) treatment. Results show that LPS perfusion produced a decrease in the basal release of dopamine. Forty-eight hours after implanting the probe, we have perfused 1 mM MPP+ to check the integrity of the dopaminergic terminals present around the cannula. Our model to study toxicity in the striatal dopaminergic terminals suggests that Zocor Forte (simvastatin) could prevent the neurotoxic damage produced by LPS, but not that produced by MPP+.