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1.
Nat Commun ; 11(1): 4885, 2020 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-32985503

RESUMO

Parkinson's disease (PD) affects millions of patients worldwide and is characterized by alpha-synuclein aggregation in dopamine neurons. Molecular tweezers have shown high potential as anti-aggregation agents targeting positively charged residues of proteins undergoing amyloidogenic processes. Here we report that the molecular tweezer CLR01 decreased aggregation and toxicity in induced pluripotent stem cell-derived dopaminergic cultures treated with PD brain protein extracts. In microfluidic devices CLR01 reduced alpha-synuclein aggregation in cell somas when axonal terminals were exposed to alpha-synuclein oligomers. We then tested CLR01 in vivo in a humanized alpha-synuclein overexpressing mouse model; mice treated at 12 months of age when motor defects are mild exhibited an improvement in motor defects and a decreased oligomeric alpha-synuclein burden. Finally, CLR01 reduced alpha-synuclein-associated pathology in mice injected with alpha-synuclein aggregates into the striatum or substantia nigra. Taken together, these results highlight CLR01 as a disease-modifying therapy for PD and support further clinical investigation.


Assuntos
Hidrocarbonetos Aromáticos com Pontes/administração & dosagem , Neurônios Dopaminérgicos/efeitos dos fármacos , Organofosfatos/administração & dosagem , Doença de Parkinson/tratamento farmacológico , Substâncias Protetoras/administração & dosagem , Animais , Modelos Animais de Doenças , Neurônios Dopaminérgicos/metabolismo , Humanos , Masculino , Camundongos , Doença de Parkinson/genética , Doença de Parkinson/metabolismo , Agregados Proteicos/efeitos dos fármacos , alfa-Sinucleína/química , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismo
2.
Nat Commun ; 11(1): 4183, 2020 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-32826893

RESUMO

We describe a human single-nuclei transcriptomic atlas for the substantia nigra (SN), generated by sequencing approximately 17,000 nuclei from matched cortical and SN samples. We show that the common genetic risk for Parkinson's disease (PD) is associated with dopaminergic neuron (DaN)-specific gene expression, including mitochondrial functioning, protein folding and ubiquitination pathways. We identify a distinct cell type association between PD risk and oligodendrocyte-specific gene expression. Unlike Alzheimer's disease (AD), we find no association between PD risk and microglia or astrocytes, suggesting that neuroinflammation plays a less causal role in PD than AD. Beyond PD, we find associations between SN DaNs and GABAergic neuron gene expression and multiple neuropsychiatric disorders. Conditional analysis reveals that distinct neuropsychiatric disorders associate with distinct sets of neuron-specific genes but converge onto shared loci within oligodendrocytes and oligodendrocyte precursors. This atlas guides our aetiological understanding by associating SN cell type expression profiles with specific disease risk.


Assuntos
Expressão Gênica , Doenças do Sistema Nervoso/genética , Doenças do Sistema Nervoso/metabolismo , Doença de Parkinson/genética , Doença de Parkinson/metabolismo , Substância Negra/metabolismo , Doença de Alzheimer/metabolismo , Astrócitos/metabolismo , Encéfalo , Neurônios Dopaminérgicos/metabolismo , Humanos , Microglia/metabolismo , Mitocôndrias/metabolismo , Doenças do Sistema Nervoso/patologia , Substância Negra/patologia , Transcriptoma
3.
Nat Commun ; 11(1): 3764, 2020 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-32724058

RESUMO

Context can influence reactions to environmental cues and this elemental process has implications for substance use disorder. Using an animal model, we show that an alcohol-associated context elevates entry into a fluid port triggered by a conditioned stimulus (CS) that predicted alcohol (CS-triggered alcohol-seeking). This effect persists across multiple sessions and, after it diminishes in extinction, the alcohol context retains the capacity to augment reinstatement. Systemically administered eticlopride and chemogenetic inhibition of ventral tegmental area (VTA) dopamine neurons reduce CS-triggered alcohol-seeking. Chemogenetically silencing VTA dopamine terminals in the nucleus accumbens (NAc) core reduces CS-triggered alcohol-seeking, irrespective of context, whereas silencing VTA dopamine terminals in the NAc shell selectively reduces the elevation of CS-triggered alcohol-seeking in an alcohol context. This dissociation reveals new roles for divergent mesolimbic dopamine circuits in the control of responding to a discrete cue for alcohol and in the amplification of this behaviour in an alcohol context.


Assuntos
Transtornos Relacionados ao Uso de Álcool/psicologia , Dopamina/metabolismo , Etanol/administração & dosagem , Extinção Psicológica/fisiologia , Área Tegmentar Ventral/fisiologia , Animais , Comportamento Animal/efeitos dos fármacos , Comportamento Animal/fisiologia , Condicionamento Clássico/efeitos dos fármacos , Condicionamento Clássico/fisiologia , Sinais (Psicologia) , Modelos Animais de Doenças , Antagonistas de Dopamina/administração & dosagem , Neurônios Dopaminérgicos/efeitos dos fármacos , Neurônios Dopaminérgicos/metabolismo , Comportamento de Procura de Droga/efeitos dos fármacos , Comportamento de Procura de Droga/fisiologia , Extinção Psicológica/efeitos dos fármacos , Feminino , Humanos , Masculino , Ratos , Salicilamidas/administração & dosagem , Técnicas Estereotáxicas , Área Tegmentar Ventral/citologia
4.
Proc Natl Acad Sci U S A ; 117(29): 17296-17307, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32631998

RESUMO

Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common cause of late-onset, autosomal-dominant familial Parkinson's disease (PD). LRRK2 functions as both a kinase and GTPase, and PD-linked mutations are known to influence both enzymatic activities. While PD-linked LRRK2 mutations can commonly induce neuronal damage in culture models, the mechanisms underlying these pathogenic effects remain uncertain. Rodent models containing familial LRRK2 mutations often lack robust PD-like neurodegenerative phenotypes. Here, we develop a robust preclinical model of PD in adult rats induced by the brain delivery of recombinant adenoviral vectors with neuronal-specific expression of human LRRK2 harboring the most common G2019S mutation. In this model, G2019S LRRK2 induces the robust degeneration of substantia nigra dopaminergic neurons, a pathological hallmark of PD. Introduction of a stable kinase-inactive mutation or administration of the selective kinase inhibitor, PF-360, attenuates neurodegeneration induced by G2019S LRRK2. Neuroprotection provided by pharmacological kinase inhibition is mediated by an unusual mechanism involving the robust destabilization of human LRRK2 protein in the brain relative to endogenous LRRK2. Our study further demonstrates that G2019S LRRK2-induced dopaminergic neurodegeneration critically requires normal GTPase activity, as hypothesis-testing mutations that increase GTP hydrolysis or impair GTP-binding activity provide neuroprotection although via distinct mechanisms. Taken together, our data demonstrate that G2019S LRRK2 induces neurodegeneration in vivo via a mechanism that is dependent on kinase and GTPase activity. Our study provides a robust rodent preclinical model of LRRK2-linked PD and nominates kinase inhibition and modulation of GTPase activity as promising disease-modifying therapeutic targets.


Assuntos
Neurônios Dopaminérgicos/metabolismo , GTP Fosfo-Hidrolases/metabolismo , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Doença de Parkinson/metabolismo , Animais , Encéfalo/metabolismo , Encéfalo/patologia , Linhagem Celular , Modelos Animais de Doenças , Dopamina/metabolismo , Feminino , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Camundongos , Camundongos Knockout , Mutação , Doenças Neurodegenerativas/tratamento farmacológico , Doenças Neurodegenerativas/metabolismo , Doença de Parkinson/patologia , Fenótipo , Projetos Piloto , Inibidores de Proteínas Quinases/farmacologia , Ratos , Ratos Wistar , Substância Negra
5.
PLoS One ; 15(7): e0223633, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32701951

RESUMO

BACKGROUND: Small conductance, calcium-activated (SK3) potassium channels control the intrinsic excitability of dopaminergic neurons (DN) in the midbrain and modulate their susceptibility to toxic insults during development. METHODS: We evaluated the age-dependency of the neuroprotective effect of an SK3 agonist, 1-Ethyl-1,3-dihydro-2H-benzimidazol-2-one (1-EBIO), on Amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) excitotoxicity to DN in ventral mesencephalon (VM) organotypic cultures. RESULTS: Most tyrosine hydroxylase (TH)+ neurons were also SK3+; SK3+/TH- cells (DN+) were common at each developmental stage but more prominently at day in vitro (DIV) 8. Young DN+ neurons were small bipolar and fusiform, whereas mature ones were large and multipolar. Exposure of organotypic cultures to AMPA (100 µm, 16 h) had no effect on the survival of DN+ at DIV 8, but caused significant toxicity at DIV 15 (n = 15, p = 0.005) and DIV 22 (n = 15, p<0.001). These results indicate that susceptibility of DN to AMPA excitotoxicity is developmental stage-dependent in embryonic VM organotypic cultures. Immature DN+ (small, bipolar) were increased after AMPA (100 µm, 16 h) at DIV 8, at the expense of the number of differentiated (large, multipolar) DN+ (p = 0.039). This effect was larger at DIV 15 (p<<<0.0001) and at DIV 22 (p<<<0.0001). At DIV 8, 30 µM 1-EBIO resulted in a large increase in DN+. At DIV 15, AMPA toxicity was prevented by exposure to 30 µM, but not 100 µM 1-EBIO. At DIV 22, excitotoxicity was unaffected by 30 µM 1-EBIO, and partially reduced by 100 µM 1-EBIO. CONCLUSION: The effects of the SK3 channel agonist 1-EBIO on the survival of SK3-expressing dopaminergic neurons were concentration-dependent and influenced by neuronal developmental stage.


Assuntos
Neurônios Dopaminérgicos/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Canais de Potássio Ativados por Cálcio de Condutância Baixa/agonistas , Animais , Benzimidazóis/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Neurônios Dopaminérgicos/metabolismo , Neurônios Dopaminérgicos/patologia , Embrião de Mamíferos/citologia , Desenvolvimento Embrionário/efeitos dos fármacos , Feminino , Mesencéfalo/citologia , Mesencéfalo/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de AMPA/metabolismo , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiônico/toxicidade
6.
Nat Commun ; 11(1): 3369, 2020 07 06.
Artigo em Inglês | MEDLINE | ID: mdl-32632153

RESUMO

Induced pluripotent stem cell (iPSC)-derived dopaminergic (DA) neurons are an expected source for cell-based therapies for Parkinson's disease (PD). The regulatory criteria for the clinical application of these therapies, however, have not been established. Here we show the results of our pre-clinical study, in which we evaluate the safety and efficacy of dopaminergic progenitors (DAPs) derived from a clinical-grade human iPSC line. We confirm the characteristics of DAPs by in vitro analyses. We also verify that the DAP population include no residual undifferentiated iPSCs or early neural stem cells and have no genetic aberration in cancer-related genes. Furthermore, in vivo studies using immunodeficient mice reveal no tumorigenicity or toxicity of the cells. When the DAPs are transplanted into the striatum of 6-OHDA-lesioned rats, the animals show behavioral improvement. Based on these results, we started a clinical trial to treat PD patients in 2018.


Assuntos
Neurônios Dopaminérgicos/citologia , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Neurais/transplante , Doença de Parkinson/terapia , Transplante de Células-Tronco/métodos , Animais , Diferenciação Celular/genética , Linhagem Celular , Modelos Animais de Doenças , Dopamina/metabolismo , Neurônios Dopaminérgicos/metabolismo , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Macaca fascicularis , Masculino , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Avaliação de Resultados em Cuidados de Saúde/métodos , Avaliação de Resultados em Cuidados de Saúde/estatística & dados numéricos , Ratos Nus , Transplante Heterólogo
7.
Nat Commun ; 11(1): 3460, 2020 07 10.
Artigo em Inglês | MEDLINE | ID: mdl-32651365

RESUMO

The learning of stimulus-outcome associations allows for predictions about the environment. Ventral striatum and dopaminergic midbrain neurons form a larger network for generating reward prediction signals from sensory cues. Yet, the network plasticity mechanisms to generate predictive signals in these distributed circuits have not been entirely clarified. Also, direct evidence of the underlying interregional assembly formation and information transfer is still missing. Here we show that phasic dopamine is sufficient to reinforce the distinctness of stimulus representations in the ventral striatum even in the absence of reward. Upon such reinforcement, striatal stimulus encoding gives rise to interregional assemblies that drive dopaminergic neurons during stimulus-outcome learning. These assemblies dynamically encode the predicted reward value of conditioned stimuli. Together, our data reveal that ventral striatal and midbrain reward networks form a reinforcing loop to generate reward prediction coding.


Assuntos
Dopamina/metabolismo , Neurônios Dopaminérgicos/efeitos dos fármacos , Neurônios Dopaminérgicos/metabolismo , Tubérculo Olfatório/efeitos dos fármacos , Animais , Dopamina/farmacologia , Masculino , Mesencéfalo/citologia , Camundongos , Modelos Teóricos , Estriado Ventral/efeitos dos fármacos , Estriado Ventral/metabolismo
8.
Life Sci ; 257: 118070, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32668327

RESUMO

AIMS: Several studies suggested that ATP-sensitive potassium channels (KATP) are potential therapeutic targets for protection against various neurodegenerative disorders, yet, there is an ongoing controversy regarding their role in Parkinson's disease (PD). Thus, the aim of the current study is to investigate the protective effect of KATP blockade and activation in the mice rotenone model of PD. MAIN METHODS: PD has been induced by 9 subcutaneous injections of rotenone (1.5 mg/kg; 3 times/week) in adult male Swiss albino mice. For 3 consecutive weeks, parkinsonian mice were either untreated or treated with L-dopa (25 mg/kg), the KATP channel blocker glibenclamide (3 mg/kg) or the KATP channel opener nicorandil (6 mg/kg). KEY FINDINGS: Glibenclamide significantly improved motor performance in the wire hanging and stair tests and halted the decline in striatal dopamine content as well as dopaminergic neurons' density. In addition, it reduced the rotenone-induced apoptosis as portrayed in the immunohistopathological examination via increasing Bcl-2 and decreasing caspases-3, -8, -9 contents. Furthermore, through its anti-inflammatory potential, glibenclamide reduced tumor necrosis factor-alpha level. On the other hand, nicorandil failed to mitigate the rotenone-induced neurodegenerative consequences. SIGNIFICANCE: KATP channel blockade by glibenclamide has neuroprotective effect against rotenone-induced neurotoxicity, that was mediated by its anti-inflammatory effect along with hindering apoptosis through extrinsic and intrinsic pathways.


Assuntos
Glibureto/farmacologia , Canais KATP/antagonistas & inibidores , Fármacos Neuroprotetores/farmacologia , Transtornos Parkinsonianos/tratamento farmacológico , Bloqueadores dos Canais de Potássio/farmacologia , Animais , Apoptose/efeitos dos fármacos , Dopamina/metabolismo , Neurônios Dopaminérgicos/metabolismo , Canais KATP/metabolismo , Levodopa/farmacologia , Masculino , Camundongos , Nicorandil/farmacologia , Transtornos Parkinsonianos/fisiopatologia , Rotenona/toxicidade
9.
Proc Natl Acad Sci U S A ; 117(30): 18037-18048, 2020 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-32641508

RESUMO

Axon-axon interactions are essential for axon guidance during nervous system wiring. However, it is unknown whether and how the growth cones communicate with each other while sensing and responding to guidance cues. We found that the Parkinson's disease gene, leucine-rich repeat kinase 2 (LRRK2), has an unexpected role in growth cone-growth cone communication. The LRRK2 protein acts as a scaffold and induces Frizzled3 hyperphosphorylation indirectly by recruiting other kinases and also directly phosphorylates Frizzled3 on threonine 598 (T598). In LRRK1 or LRRK2 single knockout, LRRK1/2 double knockout, and LRRK2 G2019S knockin, the postcrossing spinal cord commissural axons are disorganized and showed anterior-posterior guidance errors after midline crossing. Growth cones from either LRRK2 knockout or G2019S knockin mice showed altered interactions, suggesting impaired communication. Intercellular interaction between Frizzled3 and Vangl2 is essential for planar cell polarity signaling. We show here that this interaction is regulated by phosphorylation of Frizzled3 at T598 and can be regulated by LRRK2 in a kinase activity-dependent way. In the LRRK1/2 double knockout or LRRK2 G2019S knockin, the dopaminergic axon bundle in the midbrain was significantly widened and appeared disorganized, showing aberrant posterior-directed growth. Our findings demonstrate that LRRK2 regulates growth cone-growth cone communication in axon guidance and that both loss-of-function mutation and a gain-of-function mutation (G2019S) cause axon guidance defects in development.


Assuntos
Axônios/metabolismo , Receptores Frizzled/metabolismo , Cones de Crescimento/metabolismo , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Neurogênese/genética , Transdução de Sinais , Animais , Neurônios Dopaminérgicos/citologia , Neurônios Dopaminérgicos/metabolismo , Técnicas de Silenciamento de Genes , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Camundongos , Modelos Biológicos , Mutação , Neurônios/metabolismo , Fosforilação , Medula Espinal/citologia , Medula Espinal/metabolismo
10.
J Toxicol Sci ; 45(7): 391-399, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32612007

RESUMO

This study was aimed at examining propofol- (a known anesthetic) induced emotion-related behavioral disorders in mice, and exploring the possible molecular mechanisms. A total of 60 mice were divided into two groups: control and propofol group. Mice were injected with propofol (150 mg/kg, ip) at 8:00 a.m. (once a day, lasting for 30 days). During the 30 days, loss of righting reflex (LORR) and return of righting reflex (RORR) of mice were recorded every day. At the 1st (T1) and 30th (T2) day of drug discontinuance (T2), 15 mice of each group were selected to perform the open field test; then the mice underwent perfusion fixation, and the midbrain and corpus striatum were separated for immunofluorescence assay with anti-tyrosine hydroxylase (Th) and anti- dopamine transporter (DAT) antibodies. Results showed that after propofol injection, LORR and RORR increased and decreased, respectively. Long-term use of propofol resulted in decreased activities of mice (activity trajectory, line crossing, rearing time, scratching times and defecating frequency). Immunofluorescence assay showed long-term use of propofol induced decrease of Th and DAT. Collectively, our present work suggested long-term abuse of propofol induces neuropsychiatric function impairments, and the possible mechanisms are related to dopamine dyssynthesis via down-regulating tyrosine hydroxylase and dopamine transporter.


Assuntos
Anestésicos/toxicidade , Comportamento Animal/efeitos dos fármacos , Corpo Estriado/metabolismo , Corpo Estriado/patologia , Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo , Neurônios Dopaminérgicos/patologia , Transtornos Mentais/induzido quimicamente , Propofol/toxicidade , Transtornos Relacionados ao Uso de Substâncias/metabolismo , Transtornos Relacionados ao Uso de Substâncias/patologia , Tirosina 3-Mono-Oxigenase/metabolismo , Área Tegmentar Ventral/metabolismo , Área Tegmentar Ventral/patologia , Anestésicos/efeitos adversos , Animais , Neurônios Dopaminérgicos/metabolismo , Emoções/efeitos dos fármacos , Masculino , Transtornos Mentais/metabolismo , Transtornos Mentais/fisiopatologia , Transtornos Mentais/psicologia , Camundongos Endogâmicos C57BL , Propofol/efeitos adversos , Reflexo de Endireitamento/efeitos dos fármacos
11.
Nature ; 582(7813): 550-556, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32581380

RESUMO

Parkinson's disease is characterized by loss of dopamine neurons in the substantia nigra1. Similar to other major neurodegenerative disorders, there are no disease-modifying treatments for Parkinson's disease. While most treatment strategies aim to prevent neuronal loss or protect vulnerable neuronal circuits, a potential alternative is to replace lost neurons to reconstruct disrupted circuits2. Here we report an efficient one-step conversion of isolated mouse and human astrocytes to functional neurons by depleting the RNA-binding protein PTB (also known as PTBP1). Applying this approach to the mouse brain, we demonstrate progressive conversion of astrocytes to new neurons that innervate into and repopulate endogenous neural circuits. Astrocytes from different brain regions are converted to different neuronal subtypes. Using a chemically induced model of Parkinson's disease in mouse, we show conversion of midbrain astrocytes to dopaminergic neurons, which provide axons to reconstruct the nigrostriatal circuit. Notably, re-innervation of striatum is accompanied by restoration of dopamine levels and rescue of motor deficits. A similar reversal of disease phenotype is also accomplished by converting astrocytes to neurons using antisense oligonucleotides to transiently suppress PTB. These findings identify a potentially powerful and clinically feasible approach to treating neurodegeneration by replacing lost neurons.


Assuntos
Astrócitos/citologia , Modelos Animais de Doenças , Neurônios Dopaminérgicos/citologia , Doença de Parkinson/patologia , Doença de Parkinson/terapia , Substância Negra/citologia , Substância Negra/fisiologia , Animais , Axônios/fisiologia , Dopamina/biossíntese , Dopamina/metabolismo , Neurônios Dopaminérgicos/metabolismo , Feminino , Ribonucleoproteínas Nucleares Heterogêneas/deficiência , Ribonucleoproteínas Nucleares Heterogêneas/genética , Ribonucleoproteínas Nucleares Heterogêneas/metabolismo , Humanos , Técnicas In Vitro , Masculino , Camundongos , Neostriado/citologia , Neostriado/fisiologia , Vias Neurais , Neurogênese , Doença de Parkinson/metabolismo , Fenótipo , Proteína de Ligação a Regiões Ricas em Polipirimidinas/deficiência , Proteína de Ligação a Regiões Ricas em Polipirimidinas/genética , Proteína de Ligação a Regiões Ricas em Polipirimidinas/metabolismo , Substância Negra/metabolismo
12.
Proc Natl Acad Sci U S A ; 117(26): 15209-15220, 2020 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-32541058

RESUMO

Preclinical assessment of the therapeutic potential of dopamine (DA) neuron replacement in Parkinson's disease (PD) has primarily been performed in the 6-hydroxydopamine toxin model. While this is a good model to assess graft function, it does not reflect the pathological features or progressive nature of the disease. In this study, we establish a humanized transplantation model of PD that better recapitulates the main disease features, obtained by coinjection of preformed human α-synuclein (α-syn) fibrils and adeno-associated virus (AAV) expressing human wild-type α-syn unilaterally into the rat substantia nigra (SN). This model gives rise to DA neuron dysfunction and progressive loss of DA neurons from the SN and terminals in the striatum, accompanied by extensive α-syn pathology and a prominent inflammatory response, making it an interesting and relevant model in which to examine long-term function and integrity of transplanted neurons in a PD-like brain. We transplanted DA neurons derived from human embryonic stem cells (hESCs) into the striatum and assessed their survival, growth, and function over 6 to 18 wk. We show that the transplanted cells, even in the presence of ongoing pathology, are capable of innervating the DA-depleted striatum. However, on closer examination of the grafts, we found evidence of α-syn pathology in the form of inclusions of phosphorylated α-syn in a small fraction of the grafted DA neurons, indicating host-to-graft transfer of α-syn pathology, a phenomenon that has previously been observed in PD patients receiving fetal tissue grafts but has not been possible to demonstrate and study in toxin-based animal models.


Assuntos
Células-Tronco Embrionárias/fisiologia , Transplante de Células-Tronco , Sinucleinopatias , alfa-Sinucleína/metabolismo , Animais , Sobrevivência Celular , Neurônios Dopaminérgicos/metabolismo , Regulação para Baixo , Feminino , Humanos , Inflamação , Degeneração Neural , Ratos , Ratos Sprague-Dawley , Substância Negra/citologia
13.
Nature ; 582(7811): 246-252, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32499648

RESUMO

A wealth of specialized neuroendocrine command systems intercalated within the hypothalamus control the most fundamental physiological needs in vertebrates1,2. Nevertheless, we lack a developmental blueprint that integrates the molecular determinants of neuronal and glial diversity along temporal and spatial scales of hypothalamus development3. Here we combine single-cell RNA sequencing of 51,199 mouse cells of ectodermal origin, gene regulatory network (GRN) screens in conjunction with genome-wide association study-based disease phenotyping, and genetic lineage reconstruction to show that nine glial and thirty-three neuronal subtypes are generated by mid-gestation under the control of distinct GRNs. Combinatorial molecular codes that arise from neurotransmitters, neuropeptides and transcription factors are minimally required to decode the taxonomical hierarchy of hypothalamic neurons. The differentiation of γ-aminobutyric acid (GABA) and dopamine neurons, but not glutamate neurons, relies on quasi-stable intermediate states, with a pool of GABA progenitors giving rise to dopamine cells4. We found an unexpected abundance of chemotropic proliferation and guidance cues that are commonly implicated in dorsal (cortical) patterning5 in the hypothalamus. In particular, loss of SLIT-ROBO signalling impaired both the production and positioning of periventricular dopamine neurons. Overall, we identify molecular principles that shape the developmental architecture of the hypothalamus and show how neuronal heterogeneity is transformed into a multimodal neural unit to provide virtually infinite adaptive potential throughout life.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Hipotálamo/citologia , Hipotálamo/embriologia , Morfogênese , Animais , Diferenciação Celular , Linhagem da Célula , Dopamina/metabolismo , Neurônios Dopaminérgicos/citologia , Neurônios Dopaminérgicos/metabolismo , Ectoderma/citologia , Ectoderma/metabolismo , Feminino , Neurônios GABAérgicos/citologia , Neurônios GABAérgicos/metabolismo , Redes Reguladoras de Genes , Estudo de Associação Genômica Ampla , Ácido Glutâmico/metabolismo , Hipotálamo/metabolismo , Masculino , Camundongos , Morfogênese/genética , Proteínas do Tecido Nervoso/metabolismo , Neuroglia/citologia , Neuroglia/metabolismo , Neuropeptídeos/metabolismo , Neurotransmissores/metabolismo , Receptores Imunológicos/metabolismo , Regulon/genética , Transdução de Sinais , Fatores de Transcrição/metabolismo , Ácido gama-Aminobutírico/metabolismo
14.
Life Sci ; 253: 117745, 2020 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-32376269

RESUMO

AIMS: Neuroinflammation and apoptosis play a crucial role in Parkinson's disease (PD) pathogenesis. Eupatilin is a lipophilic flavonoid isolated from Artemisia species and exerts anti-apoptotic and anti-inflammatory activities. In this study, we investigated the effects of Eupatilin on a mouse model of PD induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). MAIN METHODS: The rotarod test and traction test were constructed to examine the motor function. Immunofluorescent staining was performed to detect the expression of TH, Iba-1 and GFAP. Apoptosis was examined by the TUNEL assay. Real-time PCR was used to determine the mRNA expression and Western blot and ELISA were used to determine the protein expression. KEY FINDINGS: Eupatilin improved behavioral impairment caused by MPTP. A loss of TH positive neurons was observed in the substantia nigra pars compacta of MPTP-lesioned brain, while it was rescued by Eupatilin. Moreover, MPTP administration increased the cell number of microglia and astrocytes and the expression of inflammatory factors TNF-α, IL-1ß, and IL-6. Whereas Eupatilin suppressed the activation of neuroinflammation. Eupatilin also decreased cell apoptosis enhanced by MPTP/MPP+ exposure in vivo and in vitro. We further revealed that Eupatilin abolished MPTP-induced downregulation of IκBα expression and accumulation of p65 in the nuclear compartment. Besides, MPTP administration led to dephosphorylation of Akt and GSK-3ß, but it was restored by Eupatilin. SIGNIFICANCE: We demonstrate that Eupatilin alleviates behavioral impairment and dopaminergic neuron loss induced by MPTP through inhibition of neuroinflammation and apoptosis. Our research provides more evidence for Eupatilin as a potential preventative drug for PD.


Assuntos
Neurônios Dopaminérgicos/efeitos dos fármacos , Flavonoides/farmacologia , Degeneração Neural/tratamento farmacológico , Transtornos Parkinsonianos/tratamento farmacológico , 1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina , Animais , Apoptose/efeitos dos fármacos , Astrócitos/metabolismo , Comportamento Animal/efeitos dos fármacos , Neurônios Dopaminérgicos/metabolismo , Inflamação/tratamento farmacológico , Inflamação/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microglia/metabolismo , Transtornos Parkinsonianos/fisiopatologia , Parte Compacta da Substância Negra/efeitos dos fármacos , Parte Compacta da Substância Negra/patologia , Teste de Desempenho do Rota-Rod
15.
N Engl J Med ; 382(20): 1926-1932, 2020 05 14.
Artigo em Inglês | MEDLINE | ID: mdl-32402162

RESUMO

We report the implantation of patient-derived midbrain dopaminergic progenitor cells, differentiated in vitro from autologous induced pluripotent stem cells (iPSCs), in a patient with idiopathic Parkinson's disease. The patient-specific progenitor cells were produced under Good Manufacturing Practice conditions and characterized as having the phenotypic properties of substantia nigra pars compacta neurons; testing in a humanized mouse model (involving peripheral-blood mononuclear cells) indicated an absence of immunogenicity to these cells. The cells were implanted into the putamen (left hemisphere followed by right hemisphere, 6 months apart) of a patient with Parkinson's disease, without the need for immunosuppression. Positron-emission tomography with the use of fluorine-18-L-dihydroxyphenylalanine suggested graft survival. Clinical measures of symptoms of Parkinson's disease after surgery stabilized or improved at 18 to 24 months after implantation. (Funded by the National Institutes of Health and others.).


Assuntos
Neurônios Dopaminérgicos/citologia , Células-Tronco Pluripotentes Induzidas/transplante , Doença de Parkinson/terapia , Parte Compacta da Substância Negra/citologia , Idoso , Animais , Gânglios da Base/diagnóstico por imagem , Gânglios da Base/metabolismo , Diferenciação Celular , Modelos Animais de Doenças , Neurônios Dopaminérgicos/metabolismo , Neurônios Dopaminérgicos/transplante , Seguimentos , Humanos , Células-Tronco Pluripotentes Induzidas/imunologia , Masculino , Camundongos , Camundongos SCID , Doença de Parkinson/diagnóstico por imagem , Tomografia por Emissão de Pósitrons , Putamen/diagnóstico por imagem , Tomografia Computadorizada por Raios X , Transplante Autólogo , Transplante Homólogo
16.
Anesthesiology ; 133(2): 377-392, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32412932

RESUMO

BACKGROUND: Dexmedetomidine induces a sedative response that is associated with rapid arousal. To elucidate the underlying mechanisms, the authors hypothesized that dexmedetomidine increases the activity of dopaminergic neurons in the ventral tegmental area, and that this action contributes to the unique sedative properties of dexmedetomidine. METHODS: Only male mice were used. The activity of ventral tegmental area dopamine neurons was measured by a genetically encoded Ca indicator and patch-clamp recording. Dopamine neurotransmitter dynamics in the medial prefrontal cortex and nucleus accumbens were measured by a genetically encoded dopamine sensor. Ventral tegmental area dopamine neurons were inhibited or activated by a chemogenetic approach, and the depth of sedation was estimated by electroencephalography. RESULTS: Ca signals in dopamine neurons in the ventral tegmental area increased after intraperitoneal injection of dexmedetomidine (40 µg/kg; dexmedetomidine, 16.917 [14.882; 21.748], median [25%; 75%], vs. saline, -0.745 [-1.547; 0.359], normalized data, P = 0.001; n = 6 mice). Dopamine transmission increased in the medial prefrontal cortex after intraperitoneal injection of dexmedetomidine (40 µg/kg; dexmedetomidine, 10.812 [9.713; 15.104], median [25%; 75%], vs. saline, -0.498 [-0.664; -0.355], normalized data, P = 0.001; n = 6 mice) and in the nucleus accumbens (dexmedetomidine, 8.543 [7.135; 11.828], median [25%; 75%], vs. saline, -0.329 [-1.220; -0.047], normalized data, P = 0.001; n = 6 mice). Chemogenetic inhibition or activation of ventral tegmental area dopamine neurons increased or decreased slow waves, respectively, after intraperitoneal injection of dexmedetomidine (40 µg/kg; delta wave: two-way repeated measures ANOVA, F[2, 33] = 8.016, P = 0.002; n = 12 mice; theta wave: two-way repeated measures ANOVA, F[2, 33] = 22.800, P < 0.0001; n = 12 mice). CONCLUSIONS: Dexmedetomidine activates dopamine neurons in the ventral tegmental area and increases dopamine concentrations in the related forebrain projection areas. This mechanism may explain rapid arousability upon dexmedetomidine sedation.


Assuntos
Dexmedetomidina/farmacologia , Dopamina/metabolismo , Neurônios Dopaminérgicos/metabolismo , Hipnóticos e Sedativos/farmacologia , Área Tegmentar Ventral/metabolismo , Animais , Neurônios Dopaminérgicos/química , Neurônios Dopaminérgicos/efeitos dos fármacos , Eletroencefalografia/efeitos dos fármacos , Eletroencefalografia/métodos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Técnicas de Cultura de Órgãos , Fotometria/métodos , Área Tegmentar Ventral/química , Área Tegmentar Ventral/efeitos dos fármacos
17.
Int J Mol Sci ; 21(7)2020 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-32276415

RESUMO

Numerous studies highlighted the beneficial effects of the Mediterranean diet (MD) in maintaining health, especially during ageing. Even neurodegeneration, which is part of the natural ageing process, as well as the foundation of ageing-related neurodegenerative disorders like Alzheimer's and Parkinson's disease (PD), was successfully targeted by MD. In this regard, olive oil and its polyphenolic constituents have received increasing attention in the last years. Thus, this study focuses on two main olive oil polyphenols, hydroxytyrosol (HT) and oleuropein aglycone (OLE), and their effects on ageing symptoms with special attention to PD. In order to avoid long-lasting, expensive, and ethically controversial experiments, the established invertebrate model organism Caenorhabditis elegans was used to test HT and OLE treatments. Interestingly, both polyphenols were able to increase the survival after heat stress, but only HT could prolong the lifespan in unstressed conditions. Furthermore, in aged worms, HT and OLE caused improvements of locomotive behavior and the attenuation of autofluorescence as a marker for ageing. In addition, by using three different C. elegans PD models, HT and OLE were shown i) to enhance locomotion in worms suffering from α-synuclein-expression in muscles or rotenone exposure, ii) to reduce α-synuclein accumulation in muscles cells, and iii) to prevent neurodegeneration in α-synuclein-containing dopaminergic neurons. Hormesis, antioxidative capacities and an activity-boost of the proteasome & phase II detoxifying enzymes are discussed as potential underlying causes for these beneficial effects. Further biological and medical trials are indicated to assess the full potential of HT and OLE and to uncover their mode of action.


Assuntos
Acetatos/uso terapêutico , Monoterpenos Ciclopentânicos/uso terapêutico , Neurônios Dopaminérgicos/metabolismo , Doença de Parkinson/prevenção & controle , Álcool Feniletílico/análogos & derivados , Piranos/uso terapêutico , alfa-Sinucleína , Acetatos/farmacologia , Animais , Animais Geneticamente Modificados , Caenorhabditis elegans/efeitos dos fármacos , Monoterpenos Ciclopentânicos/farmacologia , Modelos Animais de Doenças , Neurônios Dopaminérgicos/fisiologia , Álcool Feniletílico/farmacologia , Álcool Feniletílico/uso terapêutico , Polifenóis/farmacologia , Piranos/farmacologia , Resultado do Tratamento
18.
J Neurosci ; 40(18): 3533-3548, 2020 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-32253360

RESUMO

Dopaminergic neurons innervate extensive areas of the brain and release dopamine (DA) onto a wide range of target neurons. However, DA release is also precisely regulated. In Drosophila melanogaster brain explant preparations, DA is released specifically onto α3/α'3 compartments of mushroom body (MB) neurons that have been coincidentally activated by cholinergic and glutamatergic inputs. The mechanism for this precise release has been unclear. Here we found that coincidentally activated MB neurons generate carbon monoxide (CO), which functions as a retrograde signal evoking local DA release from presynaptic terminals. CO production depends on activity of heme oxygenase in postsynaptic MB neurons, and CO-evoked DA release requires Ca2+ efflux through ryanodine receptors in DA terminals. CO is only produced in MB areas receiving coincident activation, and removal of CO using scavengers blocks DA release. We propose that DA neurons use two distinct modes of transmission to produce global and local DA signaling.SIGNIFICANCE STATEMENT Dopamine (DA) is needed for various higher brain functions, including memory formation. However, DA neurons form extensive synaptic connections, while memory formation requires highly specific and localized DA release. Here we identify a mechanism through which DA release from presynaptic terminals is controlled by postsynaptic activity. Postsynaptic neurons activated by cholinergic and glutamatergic inputs generate carbon monoxide, which acts as a retrograde messenger inducing presynaptic DA release. Released DA is required for memory-associated plasticity. Our work identifies a novel mechanism that restricts DA release to the specific postsynaptic sites that require DA during memory formation.


Assuntos
Monóxido de Carbono/metabolismo , Dopamina/metabolismo , Neurônios Dopaminérgicos/metabolismo , Corpos Pedunculados/metabolismo , Terminações Pré-Sinápticas/metabolismo , Animais , Animais Geneticamente Modificados , Aprendizagem da Esquiva/fisiologia , Drosophila melanogaster , Feminino , Masculino , Olfato/fisiologia , Transmissão Sináptica/fisiologia
19.
Science ; 368(6487): 197-201, 2020 04 10.
Artigo em Inglês | MEDLINE | ID: mdl-32273471

RESUMO

Vulnerability to relapse during periods of attempted abstinence from cocaine use is hypothesized to result from the rewiring of brain reward circuitries, particularly ventral tegmental area (VTA) dopamine neurons. How cocaine exposures act on midbrain dopamine neurons to precipitate addiction-relevant changes in gene expression is unclear. We found that histone H3 glutamine 5 dopaminylation (H3Q5dop) plays a critical role in cocaine-induced transcriptional plasticity in the midbrain. Rats undergoing withdrawal from cocaine showed an accumulation of H3Q5dop in the VTA. By reducing H3Q5dop in the VTA during withdrawal, we reversed cocaine-mediated gene expression changes, attenuated dopamine release in the nucleus accumbens, and reduced cocaine-seeking behavior. These findings establish a neurotransmission-independent role for nuclear dopamine in relapse-related transcriptional plasticity in the VTA.


Assuntos
Transtornos Relacionados ao Uso de Cocaína/metabolismo , Transtornos Relacionados ao Uso de Cocaína/psicologia , Cocaína/efeitos adversos , Dopamina/metabolismo , Neurônios Dopaminérgicos/metabolismo , Comportamento de Procura de Droga , Histonas/metabolismo , Área Tegmentar Ventral/metabolismo , Animais , Transtornos Relacionados ao Uso de Cocaína/genética , Regulação da Expressão Gênica , Glutamina/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Plasticidade Neuronal , Núcleo Accumbens/metabolismo , Ratos , Ratos Sprague-Dawley , Transmissão Sináptica
20.
Proc Natl Acad Sci U S A ; 117(15): 8611-8615, 2020 04 14.
Artigo em Inglês | MEDLINE | ID: mdl-32229573

RESUMO

Electrical or optogenetic stimulation of lateral hypothalamic (LH) GABA neurons induces rapid vigorous eating in sated animals. The dopamine system has been implicated in the regulation of feeding. Previous work has suggested that a subset of LH GABA neurons projects to the ventral tegmental area (VTA) and targets GABA neurons, inhibiting them and thereby disinhibiting dopaminergic activity and release. Furthermore, stimulation-induced eating is attenuated by dopamine lesions or receptor antagonists. Here we explored the involvement of dopamine in LH stimulation-induced eating. LH stimulation caused sated mice to pick up pellets of standard chow with latencies that varied based on stimulation intensity; once food was picked up, animals ate for the remainder of the 60-s stimulation period. However, lesion of VTA GABA neurons failed to disrupt this effect. Moreover, direct stimulation of VTA or substantia nigra dopamine cell bodies failed to induce food approach or eating. Looking further, we found that some LH GABA fibers pass through the VTA to more caudal sites, where they synapse onto neurons near the locus coeruleus (LC). Similar eating was induced by stimulation of LH GABA terminals or GABA cell bodies in this peri-LC region. Lesion of peri-LC GABA neurons blocked LH stimulation-induced eating, establishing them as a critical downstream circuit element for LH neurons. Surprisingly, lesions did not alter body weight, suggesting that this system is not involved in the hunger or satiety mechanisms that govern normal feeding. Thus, we present a characterization of brain circuitry that may promote overeating and contribute to obesity.


Assuntos
Neurônios Dopaminérgicos/metabolismo , Ingestão de Alimentos/fisiologia , Comportamento Alimentar/fisiologia , Neurônios GABAérgicos/metabolismo , Região Hipotalâmica Lateral/fisiologia , Área Tegmentar Ventral/fisiologia , Animais , Comportamento Animal , Dopamina/metabolismo , Neurônios Dopaminérgicos/citologia , Feminino , Neurônios GABAérgicos/citologia , Região Hipotalâmica Lateral/citologia , Masculino , Camundongos , Vias Neurais , Receptores de GABA-A/metabolismo , Recompensa , Área Tegmentar Ventral/citologia , Ácido gama-Aminobutírico/metabolismo
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