scholarly journals β-synuclein promotes synaptic vesicle dopamine uptake and rescues dopaminergic neurons from MPTP-induced death

2020 ◽  
Author(s):  
Natalia Ninkina ◽  
Steven J. Millership ◽  
Owen M. Peters ◽  
Natalie Connor-Robson ◽  
Kirill Chaprov ◽  
...  
Keyword(s):  
Dopaminergic Neurons ◽  
Synaptic Vesicles ◽  
Dopamine Uptake ◽  
Substantia Nigra Pars Compacta ◽  
Dopamine Synthesis ◽  
Acid Decarboxylase ◽  
Toxic Metabolite ◽  
Per Se ◽  
Mptp Toxicity

Abstract Background. Previous studies demonstrated that dopaminergic neurons in the substantia nigra pars compacta (SNpc) of mice with null mutations for genes encoding α-synuclein and/or γ-synuclein are resistant to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) toxicity. An original straightforward interpretation of these results was that these proteins are directly involved in the mechanism of MPTP-induced degeneration and this view has become commonly accepted. Here we provide evidence that a plausible alternative explanation of this resistance is not the absence of these synucleins per se but their substitution on the membrane of synaptic vesicles by the third member of the family, β-synuclein. Methods. An effect of sub-chronic MPTP regimen on dopaminergic neurons of SNpc was studied in mice lacking members of the synuclein family in all possible combinations. Dopamine uptake was assessed in synaptic vesicles isolated from synuclein null mutant mice. Protein composition of synaptic vesicles was studied by mass spectrometry. Results. Dopaminergic neurons of mice lacking β-synuclein singularly or in combination with the loss of other synucleins, were sensitive to the toxic effect of MPTP. Dopamine uptake by synaptic vesicles isolated from the striatum of triple α/β/γ-synuclein deficient mice was significantly reduced, while reintroduction of β-synuclein either in vivo or in vitro reversed this effect. Proteomic analysis of complexes formed on the surface of synuclein-free synaptic vesicles after addition of recombinant β-synuclein identified multiple integral constituents of these vesicles as well as typically cytosolic proteins, including key enzymes involved in dopamine synthesis, tyrosine hydroxylase (TH) and aromatic L-amino acid decarboxylase (AADC). Conclusions. Of the three members of the synuclein family, only β-synuclein can play a scaffolding role for the assembly of molecular complexes that potentiate the ability of synaptic vesicles to uptake and sequester dopamine and other structurally similar molecules, including 1-methyl-4-phenylpyridinium (MPP+), a toxic metabolite of MPTP. The increased presence and activity of β-synuclein at the synaptic vesicles, and not the absence of other synucleins per se, explains the decreased sensitivity to MPTP toxicity of SNpc dopaminergic neurons in mice lacking α-synuclein and/or γ-synuclein.

scholarly journals Roles of Transcription Factors in the Development and Reprogramming of the Dopaminergic Neurons

2022 ◽  
Vol 23 (2) ◽  
pp. 845
Author(s):  
Lulu Tian ◽  
Murad Al-Nusaif ◽  
Xi Chen ◽  
Song Li ◽  
Weidong Le
Keyword(s):  
Transcription Factors ◽  
Cell Therapy ◽  
Dopaminergic Neurons ◽  
Neuronal Loss ◽  
Human Movement ◽  
Dopamine Synthesis ◽  
Acid Decarboxylase ◽  
Wide Range

The meso-diencephalic dopaminergic (mdDA) neurons regulate various critical processes in the mammalian nervous system, including voluntary movement and a wide range of behaviors such as mood, reward, addiction, and stress. mdDA neuronal loss is linked with one of the most prominent human movement neurological disorders, Parkinson’s disease (PD). How these cells die and regenerate are two of the most hotly debated PD research topics. As for the latter, it has been long known that a series of transcription factors (TFs) involves the development of mdDA neurons, specifying cell types and controlling developmental patterns. In vitro and in vivo, TFs regulate the expression of tyrosine hydroxylase, a dopamine transporter, vesicular monoamine transporter 2, and L-aromatic amino acid decarboxylase, all of which are critical for dopamine synthesis and transport in dopaminergic neurons (DA neurons). In this review, we encapsulate the molecular mechanism of TFs underlying embryonic growth and maturation of mdDA neurons and update achievements on dopaminergic cell therapy dependent on knowledge of TFs in mdDA neuronal development. We believe that a deeper understanding of the extrinsic and intrinsic factors that influence DA neurons’ fate and development in the midbrain could lead to a better strategy for PD cell therapy.

scholarly journals Establishment of an in vitro model for analyzing mitochondrial ultrastructure in PRKN-mutated patient iPSC-derived dopaminergic neurons

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Mutsumi Yokota ◽  
Soichiro Kakuta ◽  
Takahiro Shiga ◽  
Kei-ichi Ishikawa ◽  
Hideyuki Okano ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Dopaminergic Neurons ◽  
Structural Changes ◽  
In Vitro Model ◽  
Induced Pluripotent Stem Cell ◽  
Substantia Nigra Pars Compacta ◽  
Ultrastructural Changes ◽  
Mitochondrial Morphology ◽  
Vitro Model

AbstractMitochondrial structural changes are associated with the regulation of mitochondrial function, apoptosis, and neurodegenerative diseases. PRKN is known to be involved with various mechanisms of mitochondrial quality control including mitochondrial structural changes. Parkinson’s disease (PD) with PRKN mutations is characterized by the preferential degeneration of dopaminergic neurons in the substantia nigra pars compacta, which has been suggested to result from the accumulation of damaged mitochondria. However, ultrastructural changes of mitochondria specifically in dopaminergic neurons derived from iPSC have rarely been analyzed. The main reason for this would be that the dopaminergic neurons cannot be distinguished directly among a mixture of iPSC-derived differentiated cells under electron microscopy. To selectively label dopaminergic neurons and analyze mitochondrial morphology at the ultrastructural level, we generated control and PRKN-mutated patient tyrosine hydroxylase reporter (TH-GFP) induced pluripotent stem cell (iPSC) lines. Correlative light-electron microscopy analysis and live cell imaging of GFP-expressing dopaminergic neurons indicated that iPSC-derived dopaminergic neurons had smaller and less functional mitochondria than those in non-dopaminergic neurons. Furthermore, the formation of spheroid-shaped mitochondria, which was induced in control dopaminergic neurons by a mitochondrial uncoupler, was inhibited in the PRKN-mutated dopaminergic neurons. These results indicate that our established TH-GFP iPSC lines are useful for characterizing mitochondrial morphology, such as spheroid-shaped mitochondria, in dopaminergic neurons among a mixture of various cell types. Our in vitro model would provide insights into the vulnerability of dopaminergic neurons and the processes leading to the preferential loss of dopaminergic neurons in patients with PRKN mutations.

scholarly journals Apelin-13 Protects Dopaminergic Neurons against Rotenone—Induced Neurotoxicity through the AMPK/mTOR/ULK-1 Mediated Autophagy Activation

2020 ◽  
Vol 21 (21) ◽  
pp. 8376
Author(s):  
Peng Chen ◽  
Youcui Wang ◽  
Leilei Chen ◽  
Ning Song ◽  
Junxia Xie
Keyword(s):  
Dopaminergic Neurons ◽  
Substantia Nigra Pars Compacta ◽  
Protective Effects ◽  
Gut Peptides ◽  
Neuroprotective Effects ◽  
Compound C ◽  
Apelin Receptor ◽  
Signaling Activation

Parkinson’s disease (PD) is characterized by the progressive loss of dopaminergic neurons in the substantia nigra pars compacta. Several brain–gut peptides are able to exert neuroprotective effects on the nigrostriatal dopaminergic system. Apelin-13 is a neuropeptide, conveying potential neuroprotective activities. However, whether, and how, apelin-13 could antagonize rotenone-induced neurotoxicity has not yet been elucidated. In the present study, rotenone-treated SH-SY5Y cells and rats were used to clarify whether apelin-13 has protective effects on dopaminergic neurons, both in vivo and in vitro. The results showed that apelin-13 could protect SH-SY5Y cells from rotenone-induced injury and apoptosis. Apelin-13 was able to activate autophagy, and restore rotenone induced autophagy impairment in SH-SY5Y cells, which could be blocked by the autophagy inhibitor 3-Methyladenine. Apelin-13 activated AMPK/mTOR/ULK-1 signaling, AMPKα inhibitor compound C, as well as apelin receptor blockage via siRNA, which could block apelin-13-induced signaling activation, autophagy activation, and protective effects, in rotenone-treated SH-SY5Y cells. These results indicated that apelin-13 exerted neuroprotective properties against rotenone by stimulating AMPK/mTOR/ULK-1 signaling-mediated autophagy via the apelin receptor. We also observed that intracerebroventricular injection of apelin-13 could alleviate nigrostriatal dopaminergic neuron degeneration in rotenone-treated rats. Our findings provide new insights into the mechanism by which apelin-13 might attenuate neurotoxicity in PD.

scholarly journals Morphometric analysis of dopaminergic neurons (substantia nigra) in the brain of MPTP-treated alpha-synuclein knockout mice

2021 ◽  
pp. 32-37
Author(s):  
В.В. Голоборщева ◽  
Н.А. Воронина ◽  
Р.К. Овчинников ◽  
В.Г. Кучеряну ◽  
С.Г. Морозов
Keyword(s):  
Substantia Nigra ◽  
Morphometric Analysis ◽  
Knockout Mice ◽  
Dopaminergic Neurons ◽  
Water Solution ◽  
Neuronal Population ◽  
Alpha Synuclein ◽  
Substantia Nigra Pars Compacta ◽  
Mptp Toxicity ◽  
The Brain

Целью данной работы являлась оценка выживаемости популяции зрелых дофаминергических (ДА-ергических) нейронов чёрной субстанции двух альфа-синуклеин нокаутных линий мышей Abel-KO и ΔFlox-KO, а также бессинуклеиновых животных abg-КО в условиях МФТП-токсического моделирования паркинсонического синдрома. Методы исследования: Водный раствор нейротоксина МФТП вводили 3-месячным мышам внутрибрюшинно в дозе 30 мг/кг ежедневно в течение 5 дней по субхроническому протоколу. Через 21 день после последней инъекции МФТП у животных извлекали головной мозг, фиксировали в холодном растворе Карнуа и парафинизировали для последующего приготовления гистологических препаратов на ротационном микротоме Leica RM2265 (Leica Biosystems, Германия). Иммуногистохимическое окрашивание проводили антителами против тирозингидроксилазы (моноклональные антитела мыши, Sigma, разведение 1:2000). Сравнительный морфометрический анализ популяции ДА-ергических нейронов чёрной субстанции выполнен с учётом поправки Аберкромби. Результаты: Установлено, что в условиях дефицита альфа-синуклеина мыши устойчивы к потере ДА-ергических нейронов в компактной части ЧС после введения МФТП. При генетической делеции всех трёх синуклеинов чувствительность ДА-ергических нейронов ЧС к токсическому действию МФТП не отличается от таковой у животных с немодифицированным геномом. Заключение. На основании проведённого морфометрического анализа предполагается, что особенности чувствительности к нейротоксину МФТП у альфа-синуклеин нокаутных линий мышей обусловлены повышением функциональной активности (замещением) бета-синуклеина, оптимизирующего захват ДА синаптическими везикулами. The aim of this study was to assess survival of mature dopaminergic (DAergic) neuronal population in the substantia nigra pars compacta (SNpc) of two alpha-synuclein knockout mice strains (Abel-KO and ΔFlox-KO) and of non-synuclein animals (abg-KO) in MPTP-induced parkinsonism. Material and methods: MPTP water solution was administered to 3-month-old mice intraperitoneally (30 mg/kg daily for 5 days) according to a subchronic protocol. On the 21st day after the last MPTP injection, the brain was excised, fixed in cold Carnoy’s solution and paraffined for the subsequent preparation of histological samples on a Leica RM2265 rotary microtome (Leica Biosystems, Germany). Immunohistochemical staining was performed with antibodies against tyrosine hydroxylase (mouse monoclonal antibodies, Sigma, dilution 1:2000). A comparative morphometric analysis of substantia nigra dopaminergic neurons was performed using the Abercrombie correction. Results: MPTP-treated alpha-synuclein deficient mice were resistant to the loss of DAergic neurons in the SNpc. Genetic deletion of all three synucleins restored the sensitivity of SNpc DAergic neurons to the MPTP toxicity, which did not differ from the sensitivity of wild type animals. Conclusion: Based on the morphometric analysis, it was assumed that the specific features of MPTP sensitivity in alpha-synuclein knockout mice are due to an increased functional activity (substitution) of beta-synuclein, which optimizes the capture of DA by synaptic vesicles.

scholarly journals Neuroprotective and Neurorestorative Effects of Holothuria scabra Extract in the MPTP/MPP+-Induced Mouse and Cellular Models of Parkinson’s Disease

2020 ◽  
Vol 14 ◽  
Author(s):  
Kunwadee Noonong ◽  
Prasert Sobhon ◽  
Morakot Sroyraya ◽  
Kulathida Chaithirayanon
Keyword(s):  
Dopaminergic Neurons ◽  
Substantia Nigra Pars Compacta ◽  
Motor Deficits ◽  
Holothuria Scabra ◽  
Cellular Models ◽  
Anti Oxidant ◽  
Pre Treatment ◽  
Induced Apoptosis

Extracts from Holothuria scabra (HS) have been shown to possess anti-inflammation, anti-oxidant and anti-cancer activities. More recently, it was shown to have neuroprotective potential in Caenorhabditis elegans PD model. Here, we assessed whether HS has neuroprotective and neurorestorative effects on dopaminergic neurons in both mouse and cellular models of PD. We found that both pre-treatment and post-treatment with HS improved motor deficits in PD mouse model induced with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) as determined by grid walk test. This was likely mediated by HS protective and restorative effects on maintaining the numbers of dopaminergic neurons and fibers in both substantia nigra pars compacta (SNpc) and striatum. In a cellular model of PD, HS significantly attenuated 1-methyl-4-phenylpyridinium (MPP+)-induced apoptosis of DAergic-like neurons differentiated from SH-SY5Y cells by enhancing the expression of Bcl-2, suppressing the expression of cleaved Caspase 3 and preventing depolarization of mitochondrial membrane. In addition, HS could stimulate the expression of tyrosine hydroxylase (TH) and suppressed the formation of α-synuclein protein. Taken together, our in vivo and in vitro findings suggested that HS is an attractive candidate for the neuroprotection rather than neurorestoration in PD.

scholarly journals Protective Mechanisms of Flavonoids in Parkinson’s Disease

2015 ◽  
Vol 2015 ◽  
pp. 1-14 ◽  
Author(s):  
Kasthuri Bai Magalingam ◽  
Ammu Kutty Radhakrishnan ◽  
Nagaraja Haleagrahara
Keyword(s):  
Oxidative Stress ◽  
Parkinson’S Disease ◽  
Parkinson's Disease ◽  
Movement Disorder ◽  
Dopaminergic Neurons ◽  
Target Therapy ◽  
Cellular Damage ◽  
Substantia Nigra Pars Compacta ◽  
Protective Mechanisms

Parkinson’s disease is a chronic, debilitating neurodegenerative movement disorder characterized by progressive degeneration of dopaminergic neurons in thesubstantia nigra pars compactaregion in human midbrain. To date, oxidative stress is the well accepted concept in the etiology and progression of Parkinson’s disease. Hence, the therapeutic agent is targeted against suppressing and alleviating the oxidative stress-induced cellular damage. Within the past decades, an explosion of research discoveries has reported on the protective mechanisms of flavonoids, which are plant-based polyphenols, in the treatment of neurodegenerative disease using bothin vitroandin vivomodels. In this paper, we have reviewed the literature on the neuroprotective mechanisms of flavonoids in protecting the dopaminergic neurons hence reducing the symptoms of this movement disorder. The mechanism reviewed includes effect of flavonoids in activation of endogenous antioxidant enzymes, suppressing the lipid peroxidation, inhibition of inflammatory mediators, flavonoids as a mitochondrial target therapy, and modulation of gene expression in neuronal cells.

Subthalamic Stimulation-Induced Synaptic Responses in Substantia Nigra Pars Compacta Dopaminergic Neurons In Vitro

1999 ◽  
Vol 82 (2) ◽  
pp. 925-933 ◽  
Author(s):  
Yuji Iribe ◽  
Kevin Moore ◽  
Kevin C. H. Pang ◽  
James M. Tepper
Keyword(s):  
Substantia Nigra ◽  
Glutamate Receptor ◽  
Dopaminergic Neurons ◽  
Reversal Potential ◽  
Substantia Nigra Pars Compacta ◽  
Inhibitory Effects ◽  
Synaptic Potentials ◽  
Stimulation Of ◽  
Synaptic Responses

The subthalamic nucleus (STN) is one of the principal sources of excitatory glutamatergic input to dopaminergic neurons of the substantia nigra, yet stimulation of the STN produces both excitatory and inhibitory effects on nigral dopaminergic neurons recorded extracellularly in vivo. The present experiments were designed to determine the sources of the excitatory and inhibitory effects. Synaptic potentials were recorded intracellularly from substantia nigra pars compacta dopaminergic neurons in parasagittal slices in response to stimulation of the STN. Synaptic potentials were analyzed for onset latency, amplitude, duration, and reversal potential in the presence and absence of GABA and glutamate receptor antagonists. STN-evoked depolarizing synaptic responses in dopaminergic neurons reversed at approximately −31 mV, intermediate between the expected reversal potential for an excitatory and an inhibitory postsynaptic potential (EPSP and IPSP). Blockade of GABAA receptors with bicuculline caused a positive shift in the reversal potential to near 0 mV, suggesting that STN stimulation evoked a near simultaneous EPSP and IPSP. Both synaptic responses were blocked by application of the glutamate receptor antagonist, 6-cyano-7-nitroquinoxalene-2,3-dione. The confounding influence of inhibitory fibers of passage from globus pallidus and/or striatum by STN stimulation was eliminated by unilaterally transecting striatonigral and pallidonigral fibers 3 days before recording. The reversal potential of STN-evoked synaptic responses in dopaminergic neurons in slices from transected animals was approximately −30 mV. Bath application of bicuculline shifted the reversal potential to ∼5 mV as it did in intact animals, suggesting that the source of the IPSP was within substantia nigra. These data indicate that electrical stimulation of the STN elicits a mixed EPSP-IPSP in nigral dopaminergic neurons due to the coactivation of an excitatory monosynaptic and an inhibitory polysynaptic connection between the STN and the dopaminergic neurons of substantia nigra pars compacta. The EPSP arises from a direct monosynaptic excitatory glutamatergic input from the STN. The IPSP arises polysynaptically, most likely through STN-evoked excitation of GABAergic neurons in substantia nigra pars reticulata, which produces feed-forward GABAA-mediated inhibition of dopaminergic neurons through inhibitory intranigral axon collaterals.

scholarly journals Characterization of substantia nigra neurogenesis in homeostasis and dopaminergic degeneration: beneficial effects of the microneurotrophin BNN-20

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Theodora Mourtzi ◽  
Dimitrios Dimitrakopoulos ◽  
Dimitrios Kakogiannis ◽  
Charalampos Salodimitris ◽  
Konstantinos Botsakis ◽  
...  
Keyword(s):  
Stem Cell ◽  
Substantia Nigra ◽  
Mouse Model ◽  
Dopaminergic Neurons ◽  
Stem Cell Niche ◽  
Substantia Nigra Pars Compacta ◽  
Dopaminergic Cell ◽  
Cell Niche

Abstract Background Loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc) underlines much of the pathology of Parkinson’s disease (PD), but the existence of an endogenous neurogenic system that could be targeted as a therapeutic strategy has been controversial. BNN-20 is a synthetic, BDNF-mimicking, microneurotrophin that we previously showed to exhibit a pleiotropic neuroprotective effect on the dopaminergic neurons of the SNpc in the “weaver” mouse model of PD. Here, we assessed its potential effects on neurogenesis. Methods We quantified total numbers of dopaminergic neurons in the SNpc of wild-type and “weaver” mice, with or without administration of BNN-20, and we employed BrdU labelling and intracerebroventricular injections of DiI to evaluate the existence of dopaminergic neurogenesis in the SNpc and to assess the origin of newborn dopaminergic neurons. The in vivo experiments were complemented by in vitro proliferation/differentiation assays of adult neural stem cells (NSCs) isolated from the substantia nigra and the subependymal zone (SEZ) stem cell niche to further characterize the effects of BNN-20. Results Our analysis revealed the existence of a low-rate turnover of dopaminergic neurons in the normal SNpc and showed, using three independent lines of experiments (stereologic cell counts, BrdU and DiI tracing), that the administration of BNN-20 leads to increased neurogenesis in the SNpc and to partial reversal of dopaminergic cell loss. The newly born dopaminergic neurons, that are partially originated from the SEZ, follow the typical nigral maturation pathway, expressing the transcription factor FoxA2. Importantly, the pro-cytogenic effects of BNN-20 were very strong in the SNpc, but were absent in other brain areas such as the cortex or the stem cell niche of the hippocampus. Moreover, although the in vitro assays showed that BNN-20 enhances the differentiation of NSCs towards glia and neurons, its in vivo administration stimulated only neurogenesis. Conclusions Our results demonstrate the existence of a neurogenic system in the SNpc that can be manipulated in order to regenerate the depleted dopaminergic cell population in the “weaver” PD mouse model. Microneurotrophin BNN-20 emerges as an excellent candidate for future PD cell replacement therapies, due to its area-specific, pro-neurogenic effects.

scholarly journals Enhanced Hexokinase 2 Expression and Lactate Production Promote The Apoptosis of Dopaminergic Neurons in Parkinson’s Disease

2021 ◽  
Author(s):  
Jingyi Li ◽  
Longmin Chen ◽  
Qixiong Qin ◽  
Danlei Wang ◽  
Jingwei Zhao ◽  
...  
Keyword(s):  
Parkinson’S Disease ◽  
Parkinson's Disease ◽  
Mouse Model ◽  
Dopaminergic Neurons ◽  
Lactate Production ◽  
Substantia Nigra Pars Compacta ◽  
Hexokinase 2 ◽  
Lactate Levels

Abstract Background: Parkinson’s disease (PD) is characterized by impaired mitochondrial function and decreased ATP levels. Glycolysis is upregulated and lactate production is enhanced in PD. Since lactate promotes apoptosis and α-synuclein accumulation in neurons, we hypothesized that the increased lactate resulted from upregulated glycolysis is involved in the apoptosis of dopaminergic neurons in PD.Methods: We examined the expression of hexokinase 2 (HK2) and lactate dehydrogenase (LDH), the key enzymes in glycolysis, and lactate levels in the substantia nigra pars compacta (SNpc) of MPTP-induced mouse model of PD and in MPP+-treated SH-SY5Y cells. We investigated the role of HK2, lactate and AMPK pathway in the apoptosis of dopaminergic neurons by intervened with 3-Brpa, the HK2 inhibitor, in in vivo and in vitro systems.Results: We found that the expression of HK2 and LDHA, and lactate levels were markedly increased in brain SNpc of MPTP-treated mouse and in MPP+-treated SH-SY5Y cells. Meanwhile, the apoptosis of dopaminergic neurons in the mouse model and the apoptosis of the SH-SY5Y in vitro system were increased. Intriguingly, using HK2 inhibitor or siRNA can decrease the lactate levels and suppressed the apoptosis of dopaminergic neurons both in vivo and in vitro. Mechanistically, lactate increased the activity of adenosine monophosphate activated protein kinase (AMPK), and suppressed the phosphorylation of serine/threonine kinase 1 (Akt) and mammalian target of rapamycin (mTOR). Conclusion：Inhibition of HK2 ameliorate the apoptosis of dopaminergic neurons through downregulating the lactate production and AMPK/ Akt/ mTOR pathway activation in PD.

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