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

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

Increased vulnerability of dopaminergic neurons in MPTP-lesioned interleukin-6 deficient mice

To test the hypothesis that neuroinflammation contributes to dopaminergic neuron death in the MPTP-lesioned mouse, we compared nigrostriatal degeneration in interleukin (IL)-6 (+/+) with IL-6 (-/-) mice. In the absence of IL-6, a single injection of MPTP (30 mg/kg) resulted in significantly greater striatal dopamine depletion than that measured in IL-6 (+/+) mice. The observed dopamine depletion was MPTP dose dependent. This loss of striatal dopamine and a significantly greater loss of TH+ cells in the substantia nigra pars compacta in IL-6 (-/-) mice as compared with control IL-6 (+/+) mice, suggest that IL-6 is neuroprotective in the MPTP-lesioned nigrostriatal system. Co-localization experiments identified striatal astrocytes as the source of IL-6 in IL-6 (+/+) mice at 1 and 7 days postinjection of MPTP. The increased sensitivity of dopaminergic neurons to neurotoxicant in the absence of IL-6, is compatible with a neuroprotective activity of IL-6 in the injured nigrostriatal system.     

Soluble Epoxide Hydrolase Inhibition Attenuates MPTP-Induced Neurotoxicity in the Nigrostriatal Dopaminergic System: Involvement of α-Synuclein Aggregation and ER Stress

Soluble epoxide hydrolase (sEH) is widely expressed in the mammalian brain and possesses dual enzymatic activities, including C-terminal epoxide hydrolase (C-EH) which degrades epoxyeicosatrienoic acid (EET), a beneficial arachidonic acid metabolite. In the present study, the neuroprotective effect of sEH inhibition on 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced neurodegeneration of nigrostriatal dopaminergic system was investigated using genetic and pharmacological approaches. MPTP (15 mg/kg) was intraperitoneally injected in sEH knockout (KO) mice and C57BL/6J mice as wild-type (WT) mice. Compared with the MPTP-treated WT mice, MPTP-induced reductions in striatal dopamine content and nigral tyrosine hydroxylase level (TH, a biomarker of dopaminergic neurons) were less significant in the treated sEH mice. Furthermore, MPTP-induced HO-1 elevation (a redox-regulated protein), α-synuclein aggregation, and caspase 12 activation (a hallmark of ER stress) were less prominent in sEH KO mice than in WT mice. These data indicate that sEH KO mice are more resistant to MPTP-induced neurotoxicity. The pharmacological effect of N-[1-(1-oxopropyl)-4-piperidinyl]-N0-[4-(trifluoromethoxy)phenyl)-urea (TPPU, an sEH inhibitor) on MPTP-induced neurotoxicity was investigated in WT mice. TPPU (1 mg/kg, i.p.) attenuated MPTP-induced reduction in striatal dopamine content, TH-positive cell numbers, TH, and pro-caspase 9 protein levels (an initiator caspase of apoptosis) in mouse SN. Moreover, TPPU reduced MPTP-induced HO-1 elevation, α-synuclein aggregation and caspase 12 activation, indicating that TPPU is effective in attenuating MPTP-induced oxidative stress, apoptosis, protein aggregation, and ER stress. In conclusion, our study suggests that sEH is a potential target for developing therapies for parkinsonism. Furthermore, sEH inhibitors may be of clinical significance for treating CNS neurodegenerative diseases.     

Inactivation of neuronal forebrain A2A receptors protects dopaminergic neurons in a mouse model of Parkinson’s disease

Adenosine A_2A receptors antagonists produce neuroprotective effects in animal models of Parkinson’s disease (PD). As neuroinflammation is involved in PD pathogenesis, both neuronal and glial A_2A receptors might participate to neuroprotection. We employed complementary pharmacologic and genetic approaches to A_2A receptor inactivation, in a multiple MPTP mouse model of PD, to investigate the cellular basis of neuroprotection by A_2A antagonism. MPTP·HCl (20 mg/kg daily for 4 days) was administered in mice treated with the A_2A antagonist SCH58261, or in conditional knockout mice lacking A_2A receptors on forebrain neurons (fbnA_2AKO mice). MPTP‐induced partial loss of dopamine neurons in substantia nigra pars compacta (SNc) and striatum (Str), associated with increased astroglial and microglial immunoreactivity in these areas. Astroglia was similarly activated 1, 3, and 7 days after MPTP administration, whereas maximal microglial reactivity was detected on day 1, returning to baseline 7 days after MPTP administration. SCH58261 attenuated dopamine cell loss and gliosis in SNc and Str. Selective depletion of A_2A receptors in fbnA_2AKO mice completely prevented MPTP‐induced dopamine neuron degeneration and gliosis in SNc, and partially counteracted gliosis in Str. Results provide evidence of a primary role played by neuronal A_2A receptors in neuroprotective effects of A_2A antagonists in a multiple MPTP injections model of PD. With the symptomatic antiparkinsonian potential of several A_2A receptor antagonists being pursued in clinical trials, this study adds to the rationale for broader clinical benefit and use of these drugs early in the treatment of PD.     

Reduced MPTP toxicity in noradrenaline transporter knockout mice

The noradrenergic neurons of the locus coeruleus (LC) are damaged in Parkinson's disease (PD). Neurotoxin ablation of the LC noradrenergic neurons has been shown to exacerbate the dopaminergic toxicity of MPTP, suggesting that the noradrenergic system protects dopamine neurons. We utilized mice that exhibit elevated synaptic noradrenaline (NA) by genetically deleting the noradrenaline transporter (NET), a key regulator of the noradrenergic system (NET KO mice). NET KO and wild-type littermates were administered MPTP and striatal dopamine terminal integrity was assessed by HPLC of monoamines, immmunoblotting for dopaminergic markers and tyrosine hydroxylase (TH) immunohistochemistry. MPTP significantly reduced striatal dopamine in wild-type mice, but not in the NET KO mice. To confirm that the protection observed in the NET KO mice was due to the lack of NET, we treated wild-type mice with the specific NET inhibitor, nisoxetine, and then challenged them with MPTP. Nisoxetine conferred protection to the dopaminergic system. These data indicate that NA can modulate MPTP toxicity and suggest that manipulation of the noradrenergic system may have therapeutic value in PD.     

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

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

Preventive role of PD‐1 on MPTP‐induced dopamine depletion in mice

Many current studies of Parkinson's disease (PD) suggest that inflammation is involved in the neurodegenerative process. PD‐1, a traditional Korean medicine, used to treat various brain diseases in Korea. This study was designed to investigate the effect of PD‐1 extract in the Parkinson's model of 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP) lesioned mice. The MPTP administration caused the dopamine neuron loss in the striatum and substantia nigra pars compacta (SNpc), which was demonstrated by a depletion of tyrosine hydroxylase (TH). In addition, a reduction of bcl‐2 expression with elevation of bax expression, caspase‐3 activation, and release of cytochrome c into cytosol in dopaminergic neurons of SNpc were noted. Oral administration of PD‐1 extract (50 and 100 mg kg^?1) attenuated the MPTP‐induced depletion of TH proteins in the striatum and SNpc and prevented the apoptotic effects. These results indicate that PD‐1 extract is able to protect dopaminergic neurons from MPTP‐induced neuronal death, with important implications for the treatment of PD. Copyright © 2010 John Wiley & Sons, Ltd.     

MTH1, an oxidized purine nucleoside triphosphatase, protects the dopamine neurons from oxidative damage in nucleic acids caused by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine

We previously reported that 8-oxoguanine (8-oxoG) accumulates in the cytoplasm of dopamine neurons in the substantia nigra of patients with Parkinson's disease and the expression of MTH1 carrying an oxidized purine nucleoside triphosphatase activity increases in these neurons, thus suggesting that oxidative damage in nucleic acids is involved in dopamine neuron loss. In the present study, we found that levels of 8-oxoG in cellular DNA and RNA increased in the mouse nigrostriatal system during the tyrosine hydroxylase (TH)-positive dopamine neuron loss induced by the administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). MTH1-null mice exhibited a greater accumulation of 8-oxoG in mitochondrial DNA accompanied by a more significant decrease in TH and dopamine transporter immunoreactivities in the striatum after MPTP administration, than in wild-type mice. We thus demonstrated that MTH1 protects the dopamine neurons from oxidative damage in the nucleic acids, especially in the mitochondrial DNA of striatal nerve terminals of dopamine neurons.     

Melatonin attenuates MPTP-induced neurotoxicity via preventing CDK5-mediated autophagy and SNCA/α-synuclein aggregation

Autophagy is involved in the pathogenesis of neurodegenerative diseases including Parkinson disease (PD). However, little is known about the regulation of autophagy in neurodegenerative process. In this study, we characterized aberrant activation of autophagy induced by neurotoxin 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) and demonstrated that melatonin has a protective effect on neurotoxicity. We found an excessive activation of autophagy in monkey brain tissues and C6 cells, induced by MPTP, which is mediated by CDK5 (cyclin-dependent kinase 5). MPTP treatment significantly reduced total dendritic length and dendritic complexity of cultured primary cortical neurons and melatonin could reverse this effect. Decreased TH (tyrosine hydroxylase)-positive cells and dendrites of dopaminergic neurons in the substantia nigra pars compacta (SNc) were observed in MPTP-treated monkeys and mice. Along with decreased TH protein level, we observed an upregulation of CDK5 and enhanced autophagic activity in the striatum of mice with MPTP injection. These changes could be salvaged by melatonin treatment or knockdown of CDK5. Importantly, melatonin or knockdown of CDK5 reduced MPTP-induced SNCA/α-synuclein aggregation in mice, which is widely thought to trigger the pathogenesis of PD. Finally, melatonin or knockdown of CDK5 counteracted the PD phenotype in mice induced by MPTP. Our findings uncover a potent role of CDK5-mediated autophagy in the pathogenesis of PD, and suggest that control of autophagic pathways may provide an important clue for exploring potential target for novel therapeutics of PD.     

Combination therapy with Coenzyme Q10 and creatine produces additive neuroprotective effects in models of Parkinson's and Huntington's Diseases

Coenzyme Q₁₀ (CoQ₁₀) and creatine are promising agents for neuroprotection in neurodegenerative diseases via their effects on improving mitochondrial function and cellular bioenergetics and their properties as antioxidants. We examined whether a combination of CoQ₁₀ with creatine can exert additive neuroprotective effects in a MPTP mouse model of Parkinson's disease, a 3-NP rat model of Huntington's disease (HD) and the R6/2 transgenic mouse model of HD. The combination of the two agents produced additive neuroprotective effects against dopamine depletion in the striatum and loss of tyrosine hydroxylase neurons in the substantia nigra pars compacta (SNpc) following chronic subcutaneous administration of MPTP. The combination treatment resulted in significant reduction in lipid peroxidation and pathologic α-synuclein accumulation in the SNpc neurons of the MPTP-treated mice. We also observed additive neuroprotective effects in reducing striatal lesion volumes produced by chronic subcutaneous administration of 3-NP to rats. The combination treatment showed significant effects on blocking 3-NP-induced impairment of glutathione homeostasis and reducing lipid peroxidation and DNA oxidative damage in the striatum. Lastly, the combination of CoQ₁₀ and creatine produced additive neuroprotective effects on improving motor performance and extending survival in the transgenic R6/2 HD mice. These findings suggest that combination therapy using CoQ₁₀ and creatine may be useful in the treatment of neurodegenerative diseases such as Parkinson's disease and HD.     

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

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

Therapeutic Effects of Rapamycin on MPTP-Induced Parkinsonism in Mice

In neurodegenerative disorders such as Parkinson’s disease (PD), autophagy is implicated in the process of dopaminergic neuron cell death. The α-synuclein protein is a major component of Lewy bodies and Lewy neurites, and mutations in α-synuclein have been implicated in the etiology of familial PD. The current work investigates the mechanisms underlying the therapeutic effects of the autophagy-stimulating antibiotic rapamycin in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD. Male C57BL/6 mice were treated with intravenous rapamycin or saline control for 7 days following MPTP administration. Immunohistochemistry and western blotting were used to detect alterations in the expression of PD biomarkers, including tyrosine hydroxylase (TH), and the level of autophagy was evaluated by the detection of both microtubule-associated protein light chain 3 (LC3) and α-Synuclein cleavage. In addition, levels of monoamine neurotransmitters were measured in the striatum using high performance liquid chromatography (HPLC). Immunohistochemistry using antibodies against TH indicated that the number of dopaminergic neurons in the substantia nigra following MPTP treatment was significantly higher in rapamycin-treated mice compared with saline-treated controls (p < 0.01). Levels of TH expression in the striatum were similar between the groups. α-synuclein Immunoreactivity was significantly decreased in rapamycin-treated mice compared with controls (p < 0.01). Immunoreactivity for LC3, however, was significantly higher in the rapamycin-treated animals than controls (p < 0.01). The concentrations of both striatal dopamine, and the dopamine metabolite DOPAC, were significantly decreased in both MPTP-treated groups compared with untreated controls. The loss of DOPAC was less severe in rapamycin-treated mice compared with saline-treated mice (p < 0.01) following MPTP treatment. These results demonstrate that treatment with rapamycin is able to prevent the loss of TH-positive neurons and to ameliorate the loss of DOPAC following MPTP treatment, likely via activation of autophagy/lysosome pathways. Thus, further investigation into the effectiveness of rapamycin administration in the treatment of PD is warranted.     

Compensatory regulation of dopamine after ablation of the tyrosine hydroxylase gene in the nigrostriatal projection

The tyrosine hydroxylase (TH; EC 1.14.16.2) is a rate-limiting enzyme in the dopamine synthesis and important for the central dopaminergic system, which controls voluntary movements and reward-dependent behaviors. Here, to further explore the regulatory mechanism of dopamine levels by TH in adult mouse brains, we employed a genetic method to inactivate the Th gene in the nigrostriatal projection using the Cre-loxP system. Stereotaxic injection of adeno-associated virus expressing Cre recombinase (AAV-Cre) into the substantia nigra pars compacta (SNc), where dopaminergic cell bodies locate, specifically inactivated the Th gene. Whereas the number of TH-expressing cells decreased to less than 40% in the SNc 2 weeks after the AAV-Cre injection, the striatal TH protein level decreased to 75%, 50%, and 39% at 2, 4, and 8 weeks, respectively, after the injection. Thus, unexpectedly, the reduction of TH protein in the striatum, where SNc dopaminergic axons innervate densely, was slower than in the SNc. Moreover, despite the essential requirement of TH for dopamine synthesis, the striatal dopamine contents were only moderately decreased, to 70% even 8 weeks after AAV-Cre injection. Concurrently, in vivo synthesis activity of l-dihydroxyphenylalanine, the dopamine precursor, per TH protein level was augmented, suggesting up-regulation of dopamine synthesis activity in the intact nigrostriatal axons. Collectively, our conditional Th gene targeting method demonstrates two regulatory mechanisms of TH in axon terminals for dopamine homeostasis in vivo: local regulation of TH protein amount independent of soma and trans-axonal regulation of apparent L-dihydroxyphenylalanine synthesis activity per TH protein.     

Time-course of nigrostriatal degeneration in a progressive MPTP-lesioned macaque model of parkinson’s disease

Parkinson’s disease (PD) is characterized by a progressive loss of substantia nigra pars compacta (SNc) neurons. The onset of clinical symptoms only occurs after the degeneration has exceeded a certain threshold. In most of the current 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) nonhuman primate models, nigrostriatal lesions and the onset of PD symptoms are the result of an immediate neuronal degeneration in the SNc caused by acute injection of the toxin. In order to develop a model that more closely mimics the degeneration pattern of human PD, we eventually established a protocol that produces a progressive parkinsonian state by treating monkeys repeatedly with MPTP for 15 ± 2 d. Mean onset of parkinsonian symptoms occurred after 13.2 d of treatment. At this time, 56.8 ± 6.3% of tyrosine hydroxylase immunoreactive neurons and 75.2 ± 6.2% of Nissl-stained cells remained in the SNc. Striatal dopamine transporter (DAT) binding and dopamine (DA) content decreased to 19.7 ± 4.9% and 18.2 ± 5.6% of untreated monkeys. Parallel ¹²³I-PEI single-photon emission computed tomography (SPECT) imaging in living animals showed a similar decrease in striatal DAT binding. In this article, we examine how this and other chronic MPTP models fit with human pathology.     

Primate model of parkinsonism: selective lesion of nigrostriatal neurons by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine produces an extrapyramidal syndrome in rhesus monkeys

Systemic administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to rhesus monkeys (1.0-2.5 mg/kg i.v.) produces irreversible damage to nigrostriatal neurons. Dopaminergic neurons in the dorsolateral part of striatum were the most vulnerable. The major clinical signs of an extrapyramidal syndrome, but not resting tremor, appeared only in MPTP-treated monkeys suffering from more than 80% reduction in striatal dopamine. No chronic changes in the mesolimbic dopaminergic system were observed. Immunocytochemical staining of the mid-brain with a tyrosine hydroxylase antiserum indicated that MPTP produced a significant decrease of dopaminergic cell bodies in the A9, but not in the A10 ventrotegmental area. Despite greater than 80% decrease in A9 nigral cell bodies, the dopamine content decreased only by 50%. Sprouting of the surviving nigral A9 neurons was observed histologically and neurochemically in the area above substantia nigra. The present behavioral, neurochemical and histological results indicate that MPTP produces an ideal primate model for studying parkinsonism. Selective lesion of more than 80% of the nigrostrial neurons by MPTP is sufficient to produce the major clinical signs of the extrapyramidal syndrome in idiopathic parkinsonism.     

环加氧酶-2对帕金森病小鼠黑质多巴胺能神经元的影响

目的和方法 :选用C5 7BL种系环加氧酶 2 (cyclooxygenase 2 ,COX 2 )缺陷小鼠 ,腹腔注射 1 甲基 4 苯基 1,2 ,3,6 四氢吡啶 (MPTP)制备帕金森病小鼠模型 ,用免疫组织化学方法观察COX 2对帕金森病小鼠黑质多巴胺能神经元的影响。结果 :行为学及免疫组织化学观察显示 ,野生型帕金森病小鼠的死亡率明显高于COX 2缺陷杂合子帕金森病小鼠 (P <0 .0 1) ,野生型帕金森病小鼠黑质致密部酪氨酸羟化酶 (tyrosinehydroxylase,TH)免疫反应阳性神经元数目较杂合子帕金森病小鼠明显减少 (P <0 .0 1)。结论 :COX 2可能与帕金森病时黑质多巴胺能神经元的损伤有关     

Effects of R- and S-apomorphine on MPTP-induced nigro-striatal dopamine neuronal loss

In order to establish whether the antioxidant and iron-chelating activities of R-apomorphine (R-APO), a D(1)-D(2) receptor agonist, may contribute to its neuroprotective property, its S-isomer, which is not a dopamine agonist, was studied. The neuroprotective property of R- and S-APO has been studied in the MPTP model of Parkinson's disease (PD). Both S-APO (0.5-1 mg/kg, subcutaneous) and R-APO (10 mg/kg) pretreatment of C57-BL mice, protected against MPTP (24 mg/kg, intraperitoneally) induced dopamine (DA) depletion and reduction in tyrosine hydroxylase (TH) activity. However, only R-APO prevented nigro-striatal neuronal cell degeneration, as indicated by the immunohistochemistry of TH positive neurones in substantia nigra and by western analysis of striatal TH content. R-APO prevented the reduction of striatal-GSH and the increase in the ratio of GSSG over total glutathione, caused by MPTP treatment. In vitro both R-APO and S-APO inhibited monoamine oxidase A and B activity at relatively high concentrations (100 and 300 micromol/L, respectively). The elevated activity of TH induced by the two enantiomers may contribute to the maintenance of normal DA levels, suggesting that one of the targets of these molecules may involve upregulation of TH activity. It is suggested that the antioxidant and iron-chelating properties, possible monoamine oxidase inhibitory actions, together with activation of DA receptors, may participate in the mechanism of neuroprotection by APO enantiomers against MPTP.     

The neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine induces apoptosis in mouse nigrostriatal glia. Relevance to nigral neuronal death and striatal neurochemical changes

Swiss mice were given 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), 25 mg/kg/day, for 5 consecutive days and killed at different days after MPTP discontinuance. Decreases in striatal tyrosine hydroxylase activity and levels of dopamine and its metabolites were observed 1 day after MPTP discontinuance. Ascorbic acid and glutamate levels had increased, dehydroascorbic acid and GSH decreased, whereas catabolites of high-energy phosphates (inosine, hypoxanthine, xanthine, and uric acid) were unchanged. In addition, gliosis was observed in both striatum and substantia nigra compacta (SNc). Sections of SNc showed some terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate nick end labeling (TUNEL)-positive cells. Neurochemical parameters of dopaminergic activity showed a trend toward recovery 3 days after MPTP discontinuance. At this time point, TUNEL-positive cells were detected in SNc; some of them showed nuclei with neuronal morphology. A late (days 6-11) increase in striatal dopamine oxidative metabolism, ascorbic acid oxidative status, and catabolites of high-energy phosphates were observed concomitant with nigral neuron and nigrostriatal glial cell apoptotic death, as revealed by TUNEL, acridine orange, and Hoechst staining, and transmission electron microscopy. These data suggest that MPTP-induced activation/apoptotic death of glial cells plays a key role in the sequential linkage of neurochemical and cellular events leading to dopaminergic nigral neuron apoptotic death.     

Angiotensin type 1 receptor antagonist losartan, reduces MPTP-induced degeneration of dopaminergic neurons in substantia nigra

Background

Recent attention has focused on understanding the role of the brain-renin-angiotensin-system (RAS) in stroke and neurodegenerative diseases. Direct evidence of a role for the brain-RAS in Parkinson's disease (PD) comes from studies demonstrating the neuroprotective effect of RAS inhibitors in several neurotoxin based PD models. In this study, we show that an antagonist of the angiotensin II (Ang II) type 1 (AT₁) receptor, losartan, protects dopaminergic (DA) neurons against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) toxicity both in primary ventral mesencephalic (VM) cultures as well as in the substantia nigra pars compacta (SNpc) of C57BL/6 mice (Fig. 1).

Results

In the presence of exogenous Ang II, losartan reduced MPP⁺ (5 μM) induced DA neuronal loss by 72% in vitro. Mice challenged with MPTP showed a 62% reduction in the number of DA neurons in the SNpc and a 71% decrease in tyrosine hydroxylase (TH) immunostaining of the striatum, whereas daily treatment with losartan lessened MPTP-induced loss of DA neurons to 25% and reduced the decrease in striatal TH⁺ immunostaining to 34% of control.

Conclusion

Our study demonstrates that the brain-RAS plays an important neuroprotective role in the MPTP model of PD and points to AT₁ receptor as a potential novel target for neuroprotection.     

Electroconvulsive shock does not modify striatal contents of dopamine in MPTP-treated mice

It has been suggested that the therapeutic response to electroconvulsive therapy in depressed patients could be mediated by functional changes in the dopaminergic pathways; a favorable response to electroconvulsive therapy was also observed recently in patients with Parkinson's disease. To study a possible interference of electroconvulsive shock in the course of MPTP-induced parkinsonism in rodents, we measured the striatal content of dopamine in MPTP-treated mice that received electroconvulsive shock at various intervals in the course of MPTP neurotoxicity. Our results showed no immediate or delayed differences in striatal dopamine content of animals that received MPTP and electroconvulsive shock when compared with animals that received only MPTP, thus suggesting that the strong biological effects of MPTP and electroconvulsive shock on the brain may follow different biochemical mechanisms.