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1.
Nat Commun ; 12(1): 5121, 2021 08 25.
Artigo em Inglês | MEDLINE | ID: mdl-34433818

RESUMO

Comparatively little is known about how new instrumental actions are encoded in the brain. Using whole-brain c-Fos mapping, we show that neural activity is increased in the anterior dorsolateral striatum (aDLS) of mice that successfully learn a new lever-press response to earn food rewards. Post-learning chemogenetic inhibition of aDLS disrupts consolidation of the new instrumental response. Similarly, post-learning infusion of the protein synthesis inhibitor anisomycin into the aDLS disrupts consolidation of the new response. Activity of D1 receptor-expressing medium spiny neurons (D1-MSNs) increases and D2-MSNs activity decreases in the aDLS during consolidation. Chemogenetic inhibition of D1-MSNs in aDLS disrupts the consolidation process whereas D2-MSN inhibition strengthens consolidation but blocks the expression of previously learned habit-like responses. These findings suggest that D1-MSNs in the aDLS encode new instrumental actions whereas D2-MSNs oppose this new learning and instead promote expression of habitual actions.


Assuntos
Condicionamento Operante , Corpo Estriado/fisiologia , Neurônios/fisiologia , Animais , Comportamento Animal , Corpo Estriado/citologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ratos , Ratos Long-Evans , Receptores de Dopamina D1/genética , Receptores de Dopamina D1/metabolismo , Receptores de Dopamina D2/genética , Receptores de Dopamina D2/metabolismo
2.
Int J Mol Sci ; 22(15)2021 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-34360638

RESUMO

Perinatal hypoxia-ischemia (HI) is a major cause of striatal injury. Delayed post-treatment with adult-sourced bone marrow-derived mesenchymal stem cells (BMSCs) increased the absolute number of striatal medium-spiny neurons (MSNs) following perinatal HI-induced brain injury. Yet extraction of BMSCs is more invasive and difficult compared to extraction of adipose-derived mesenchymal stem cells (AD-MSCs), which are easily sourced from subcutaneous tissue. Adult-sourced AD-MSCs are also superior to BMSCs in the treatment of adult ischemic stroke. Therefore, we investigated whether delayed post-treatment with adult-sourced AD-MSCs increased the absolute number of striatal MSNs following perinatal HI-induced brain injury. This included investigation of the location of injected AD-MSCs within the brain, which were widespread in the dorsolateral subventricular zone (dlSVZ) at 1 day after their injection. Cells extracted from adult rat tissue were verified to be stem cells by their adherence to tissue culture plastic and their expression of specific 'cluster of differentiation' (CD) markers. They were verified to be AD-MSCs by their ability to differentiate into adipocytes and osteocytes in vitro. Postnatal day (PN) 7/8, male Sprague-Dawley rats were exposed to either HI right-sided brain injury or no HI injury. The HI rats were either untreated (HI + Diluent), single stem cell-treated (HI + MSCs×1), or double stem cell-treated (HI + MSCs×2). Control rats that were matched-for-weight and litter had no HI injury and were treated with diluent (Uninjured + Diluent). Treatment with AD-MSCs or diluent occurred either 7 days, or 7 and 9 days, after HI. There was a significant increase in the absolute number of striatal dopamine and cyclic AMP-regulated phosphoprotein (DARPP-32)-positive MSNs in the double stem cell-treated (HI + MSCs×2) group and the normal control group compared to the HI + Diluent group at PN21. We therefore investigated two potential mechanisms for this effect of double-treatment with AD-MSCs. Specifically, did AD-MSCs: (i) increase the proliferation of cells within the dlSVZ, and (ii) decrease the microglial response in the dlSVZ and striatum? It was found that a primary repair mechanism triggered by double treatment with AD-MSCs involved significantly decreased striatal inflammation. The results may lead to the development of clinically effective and less invasive stem cell therapies for neonatal HI brain injury.


Assuntos
Corpo Estriado/citologia , Hipóxia-Isquemia Encefálica/terapia , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/fisiologia , Células-Tronco Adultas/fisiologia , Animais , Animais Recém-Nascidos , Masculino , Ratos , Ratos Sprague-Dawley , Tempo para o Tratamento
3.
Nat Commun ; 12(1): 4855, 2021 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-34381044

RESUMO

The vertebrate brain consists of diverse neuronal types, classified by distinct anatomy and function, along with divergent transcriptomes and proteomes. Defining the cell-type specific neuroproteomes is important for understanding the development and functional organization of neural circuits. This task remains challenging in complex tissue, due to suboptimal protein isolation techniques that often result in loss of cell-type specific information and incomplete capture of subcellular compartments. Here, we develop a genetically targeted proximity labeling approach to identify cell-type specific subcellular proteomes in the mouse brain, confirmed by imaging, electron microscopy, and mass spectrometry. We virally express subcellular-localized APEX2 to map the proteome of direct and indirect pathway spiny projection neurons in the striatum. The workflow provides sufficient depth to uncover changes in the proteome of striatal neurons following chemogenetic activation of Gαq-coupled signaling cascades. This method enables flexible, cell-type specific quantitative profiling of subcellular proteome snapshots in the mouse brain.


Assuntos
Ascorbato Peroxidases/metabolismo , Núcleo Celular/metabolismo , Corpo Estriado/metabolismo , Proteoma/metabolismo , Animais , Ascorbato Peroxidases/genética , Corpo Estriado/citologia , Citosol/metabolismo , Espectrometria de Massas , Camundongos , Vias Neurais , Neurônios/citologia , Neurônios/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Transdução de Sinais , Coloração e Rotulagem , Fluxo de Trabalho
4.
FASEB J ; 35(8): e21791, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34320240

RESUMO

Chemical neurotransmission typically occurs through synapses. Previous ultrastructural examinations of monoamine neuron axon terminals often failed to identify a pre- and postsynaptic coupling, leading to the concept of "volume" transmission. Whether this results from intrinsic properties of these neurons remains undefined. We find that dopaminergic neurons in vitro establish a distinctive axonal arbor compared to glutamatergic or GABAergic neurons in both size and propensity of terminals to avoid direct contact with target neurons. While most dopaminergic varicosities are active and contain exocytosis proteins like synaptotagmin 1, only ~20% of these are synaptic. The active zone protein bassoon was found to be enriched in dopaminergic terminals that are in proximity to a target cell. Finally, we found that the proteins neurexin-1αSS4- and neuroligin-1A+B play a critical role in the formation of synapses by dopamine (DA) neurons. Our findings suggest that DA neurons are endowed with a distinctive developmental connectivity program.


Assuntos
Axônios/fisiologia , Proteínas de Ligação ao Cálcio/metabolismo , Moléculas de Adesão Celular Neuronais/metabolismo , Corpo Estriado/citologia , Dopamina/metabolismo , Neurônios Dopaminérgicos/fisiologia , Moléculas de Adesão de Célula Nervosa/metabolismo , Animais , Proteínas de Ligação ao Cálcio/genética , Moléculas de Adesão Celular Neuronais/genética , Diferenciação Celular , Técnicas de Cocultura/métodos , Dopamina/genética , Regulação da Expressão Gênica , Proteínas de Fluorescência Verde , Imuno-Histoquímica , Camundongos Transgênicos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Moléculas de Adesão de Célula Nervosa/genética , Tirosina 3-Mono-Oxigenase/genética , Tirosina 3-Mono-Oxigenase/metabolismo
5.
RNA ; 27(10): 1220-1240, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34244459

RESUMO

Metabotropic glutamate receptor 4 (mGlu4) is one of eight mGlu receptors within the Class C G protein-coupled receptor superfamily. mGlu4 is primarily localized to the presynaptic membrane of neurons where it functions as an auto and heteroreceptor controlling synaptic release of neurotransmitter. mGlu4 is implicated in numerous disorders and is a promising drug target; however, more remains to be understood about its regulation and pharmacology. Using high-throughput sequencing, we have validated and quantified an adenosine-to-inosine (A-to-I) RNA editing event that converts glutamine 124 to arginine in mGlu4; additionally, we have identified a rare but novel K129R site. Using an in vitro editing assay, we then validated the pre-mRNA duplex that allows for editing by ADAR enzymes and predicted its conservation across the mammalian species. Structural modeling of the mGlu4 protein predicts the Q124R substitution to occur in the B helix of the receptor that is critical for receptor dimerization and activation. Interestingly, editing of a receptor homodimer does not disrupt G protein activation in response to the endogenous agonist, glutamate. Using an assay designed to specifically measure heterodimer populations at the surface, however, we found that Q124R substitution decreased the propensity of mGlu4 to heterodimerize with mGlu2 and mGlu7 Our study is the first to extensively describe the extent and regulatory factors of RNA editing of mGlu4 mRNA transcripts. In addition, we have proposed a novel functional consequence of this editing event that provides insights regarding its effects in vivo and expands the regulatory capacity for mGlu receptors.


Assuntos
Edição de RNA , RNA Mensageiro/genética , Receptores de Glutamato Metabotrópico/genética , Adenosina Desaminase/genética , Adenosina Desaminase/metabolismo , Sequência de Aminoácidos , Animais , Pareamento de Bases , Sequência de Bases , Aves , Córtex Cerebral/citologia , Córtex Cerebral/metabolismo , Corpo Estriado/citologia , Corpo Estriado/metabolismo , Células HEK293 , Hipocampo/citologia , Hipocampo/metabolismo , Humanos , Modelos Moleculares , Neurônios/citologia , Neurônios/metabolismo , Conformação de Ácido Nucleico , Mutação Puntual , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , RNA Mensageiro/química , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de Glutamato Metabotrópico/química , Receptores de Glutamato Metabotrópico/metabolismo , Répteis , Homologia de Sequência de Aminoácidos
6.
Science ; 372(6542)2021 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-33958447

RESUMO

Deciphering how the human striatum develops is necessary for understanding the diseases that affect this region. To decode the transcriptional modules that regulate this structure during development, we compiled a catalog of 1116 long intergenic noncoding RNAs (lincRNAs) identified de novo and then profiled 96,789 single cells from the early human fetal striatum. We found that D1 and D2 medium spiny neurons (D1- and D2-MSNs) arise from a common progenitor and that lineage commitment is established during the postmitotic transition, across a pre-MSN phase that exhibits a continuous spectrum of fate determinants. We then uncovered cell type-specific gene regulatory networks that we validated through in silico perturbation. Finally, we identified human-specific lincRNAs that contribute to the phylogenetic divergence of this structure in humans. This work delineates the cellular hierarchies governing MSN lineage commitment.


Assuntos
Atlas como Assunto , Corpo Estriado/citologia , Corpo Estriado/embriologia , Neurogênese/genética , RNA Longo não Codificante/genética , Análise de Célula Única , Fatores de Transcrição/genética , Feto , Neurônios GABAérgicos/metabolismo , Humanos , RNA-Seq , Transcrição Genética
7.
Science ; 372(6540)2021 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-33888613

RESUMO

The integrated stress response (ISR) maintains proteostasis by modulating protein synthesis and is important in synaptic plasticity, learning, and memory. We developed a reporter, SPOTlight, for brainwide imaging of ISR state with cellular resolution. Unexpectedly, we found a class of neurons in mouse brain, striatal cholinergic interneurons (CINs), in which the ISR was activated at steady state. Genetic and pharmacological manipulations revealed that ISR signaling was necessary in CINs for normal type 2 dopamine receptor (D2R) modulation. Inhibiting the ISR inverted the sign of D2R modulation of CIN firing and evoked dopamine release and altered skill learning. Thus, a noncanonical, steady-state mode of ISR activation is found in CINs, revealing a neuromodulatory role for the ISR in learning.


Assuntos
Neurônios Colinérgicos/metabolismo , Dopamina/metabolismo , Interneurônios/fisiologia , Aprendizagem/fisiologia , Estresse Fisiológico , Potenciais de Ação , Animais , Corpo Estriado/citologia , Corpo Estriado/fisiologia , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Destreza Motora , Plasticidade Neuronal , Técnicas de Patch-Clamp , Biossíntese de Proteínas , Receptores de Dopamina D2/metabolismo
8.
Nat Commun ; 12(1): 1994, 2021 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-33790281

RESUMO

The medial prefrontal cortex (mPFC) steers goal-directed actions and withholds inappropriate behavior. Dorsal and ventral mPFC (dmPFC/vmPFC) circuits have distinct roles in cognitive control, but underlying mechanisms are poorly understood. Here we use neuroanatomical tracing techniques, in vitro electrophysiology, chemogenetics and fiber photometry in rats engaged in a 5-choice serial reaction time task to characterize dmPFC and vmPFC outputs to distinct thalamic and striatal subdomains. We identify four spatially segregated projection neuron populations in the mPFC. Using fiber photometry we show that these projections distinctly encode behavior. Postsynaptic striatal and thalamic neurons differentially process synaptic inputs from dmPFC and vmPFC, highlighting mechanisms that potentially amplify distinct pathways underlying cognitive control of behavior. Chemogenetic silencing of dmPFC and vmPFC projections to lateral and medial mediodorsal thalamus subregions oppositely regulate cognitive control. In addition, dmPFC neurons projecting to striatum and thalamus divergently regulate cognitive control. Collectively, we show that mPFC output pathways targeting anatomically and functionally distinct striatal and thalamic subregions encode bi-directional command of cognitive control.


Assuntos
Cognição/fisiologia , Corpo Estriado/fisiologia , Neurônios/fisiologia , Córtex Pré-Frontal/fisiologia , Tálamo/fisiologia , Animais , Corpo Estriado/citologia , Fenômenos Eletrofisiológicos , Masculino , Modelos Neurológicos , Vias Neurais/fisiologia , Córtex Pré-Frontal/citologia , Ratos Long-Evans , Tálamo/citologia
9.
Molecules ; 26(5)2021 Mar 07.
Artigo em Inglês | MEDLINE | ID: mdl-33800024

RESUMO

The endocannabinoid system (ECS) is involved in the modulation of several basic biological processes, having widespread roles in neurodevelopment, neuromodulation, immune response, energy homeostasis and reproduction. In the adult central nervous system (CNS) the ECS mainly modulates neurotransmitter release, however, a substantial body of evidence has revealed a central role in regulating neurogenesis in developing and adult CNS, also under pathological conditions. Due to the complexity of investigating ECS functions in neural progenitors in vivo, we tested the suitability of the ST14A striatal neural progenitor cell line as a simplified in vitro model to dissect the role and the mechanisms of ECS-regulated neurogenesis, as well as to perform ECS-targeted pharmacological approaches. We report that ST14A cells express various ECS components, supporting the presence of an active ECS. While CB1 and CB2 receptor blockade did not affect ST14A cell number, exogenous administration of the endocannabinoid 2-AG and the synthetic CB2 agonist JWH133 increased ST14A cell proliferation. Phospholipase C (PLC), but not PI3K pharmacological blockade negatively modulated CB2-induced ST14A cell proliferation, suggesting that a PLC pathway is involved in the steps downstream to CB2 activation. On the basis of our results, we propose ST14A neural progenitor cells as a useful in vitro model for studying ECS modulation of neurogenesis, also in prospective in vivo pharmacological studies.


Assuntos
Moduladores de Receptores de Canabinoides/farmacologia , Células-Tronco Neurais/efeitos dos fármacos , Neurogênese/fisiologia , Receptores de Canabinoides/metabolismo , Animais , Canabinoides/farmacologia , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Corpo Estriado/citologia , Estrenos/farmacologia , Células-Tronco Neurais/fisiologia , Neurogênese/efeitos dos fármacos , Pirrolidinonas/farmacologia , Ratos , Receptor CB1 de Canabinoide/agonistas , Receptor CB1 de Canabinoide/antagonistas & inibidores , Receptor CB1 de Canabinoide/genética , Receptor CB2 de Canabinoide/agonistas , Receptor CB2 de Canabinoide/antagonistas & inibidores , Receptor CB2 de Canabinoide/genética , Receptores de Canabinoides/genética , Fosfolipases Tipo C/antagonistas & inibidores
10.
Nat Commun ; 12(1): 364, 2021 01 13.
Artigo em Inglês | MEDLINE | ID: mdl-33441541

RESUMO

Temporal dynamics and mechanisms underlying epigenetic changes in Huntington's disease (HD), a neurodegenerative disease primarily affecting the striatum, remain unclear. Using a slowly progressing knockin mouse model, we profile the HD striatal chromatin landscape at two early disease stages. Data integration with cell type-specific striatal enhancer and transcriptomic databases demonstrates acceleration of age-related epigenetic remodelling and transcriptional changes at neuronal- and glial-specific genes from prodromal stage, before the onset of motor deficits. We also find that 3D chromatin architecture, while generally preserved at neuronal enhancers, is altered at the disease locus. Specifically, we find that the HD mutation, a CAG expansion in the Htt gene, locally impairs the spatial chromatin organization and proximal gene regulation. Thus, our data provide evidence for two early and distinct mechanisms underlying chromatin structure changes in the HD striatum, correlating with transcriptional changes: the HD mutation globally accelerates age-dependent epigenetic and transcriptional reprogramming of brain cell identities, and locally affects 3D chromatin organization.


Assuntos
Envelhecimento , Montagem e Desmontagem da Cromatina/genética , Corpo Estriado/metabolismo , Modelos Animais de Doenças , Doença de Huntington/genética , Doenças Neurodegenerativas/genética , Animais , Comportamento Animal/fisiologia , Cromatina/genética , Corpo Estriado/citologia , Corpo Estriado/fisiopatologia , Epigenômica/métodos , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica , Humanos , Proteína Huntingtina/genética , Doença de Huntington/diagnóstico , Doença de Huntington/fisiopatologia , Camundongos Endogâmicos C57BL , Doenças Neurodegenerativas/diagnóstico , Doenças Neurodegenerativas/fisiopatologia , Neurônios/metabolismo , Expansão das Repetições de Trinucleotídeos/genética
11.
Nature ; 591(7850): 420-425, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33473213

RESUMO

The cortex projects to the dorsal striatum topographically1,2 to regulate behaviour3-5, but spiking activity in the two structures has previously been reported to have markedly different relations to sensorimotor events6-9. Here we show that the relationship between activity in the cortex and striatum is spatiotemporally precise, topographic, causal and invariant to behaviour. We simultaneously recorded activity across large regions of the cortex and across the width of the dorsal striatum in mice that performed a visually guided task. Striatal activity followed a mediolateral gradient in which behavioural correlates progressed from visual cue to response movement to reward licking. The summed activity in each part of the striatum closely and specifically mirrored activity in topographically associated cortical regions, regardless of task engagement. This relationship held for medium spiny neurons and fast-spiking interneurons, whereas the activity of tonically active neurons differed from cortical activity with stereotypical responses to sensory or reward events. Inactivation of the visual cortex abolished striatal responses to visual stimuli, supporting a causal role of cortical inputs in driving the striatum. Striatal visual responses were larger in trained mice than untrained mice, with no corresponding change in overall activity in the visual cortex. Striatal activity therefore reflects a consistent, causal and scalable topographical mapping of cortical activity.


Assuntos
Córtex Cerebral/citologia , Córtex Cerebral/fisiologia , Corpo Estriado/citologia , Corpo Estriado/fisiologia , Animais , Feminino , Interneurônios/metabolismo , Aprendizagem , Masculino , Camundongos , Neurônios/metabolismo , Estimulação Luminosa , Desempenho Psicomotor , Recompensa , Córtex Sensório-Motor/fisiologia , Córtex Visual/fisiologia
12.
J Med Chem ; 64(2): 941-957, 2021 01 28.
Artigo em Inglês | MEDLINE | ID: mdl-33185430

RESUMO

GPR52 is an orphan G protein-coupled receptor (GPCR) that has been recently implicated as a potential drug target of Huntington's disease (HD), an incurable monogenic neurodegenerative disorder. In this research, we found that striatal knockdown of GPR52 reduces mHTT levels in adult HdhQ140 mice, validating GPR52 as an HD target. In addition, we discovered a highly potent and specific GPR52 antagonist Comp-43 with an IC50 value of 0.63 µM by a structure-activity relationship (SAR) study. Further studies showed that Comp-43 reduces mHTT levels by targeting GPR52 and promotes survival of mouse primary striatal neurons. Moreover, in vivo study showed that Comp-43 not only reduces mHTT levels but also rescues HD-related phenotypes in HdhQ140 mice. Taken together, our study confirms that inhibition of GPR52 is a promising strategy for HD therapy, and the GPR52 antagonist Comp-43 might serve as a lead compound for further investigation.


Assuntos
Proteína Huntingtina/biossíntese , Doença de Huntington/tratamento farmacológico , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Animais , Comportamento Animal/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Corpo Estriado/citologia , Corpo Estriado/efeitos dos fármacos , Desenho de Fármacos , Humanos , Doença de Huntington/fisiopatologia , Doença de Huntington/psicologia , Camundongos , Modelos Moleculares , Neurônios/efeitos dos fármacos , Relação Estrutura-Atividade
13.
Methods Mol Biol ; 2239: 77-100, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33226614

RESUMO

MicroRNAs (miRNAs), miR-9/9*, and miR-124 (miR-9/9*-124) display fate-reprogramming activities when ectopically expressed in human fibroblasts by erasing the fibroblast identity and evoking a pan-neuronal state. In contrast to induced pluripotent stem cell-derived neurons, miRNA-induced neurons (miNs) retain the biological age of the starting fibroblasts through direct fate conversion and thus provide a human neuron-based platform to study cellular properties inherent in aged neurons and model adult-onset neurodegenerative disorders using patient-derived cells. Furthermore, expression of neuronal subtype-specific transcription factors in conjunction with miR-9/9*-124 guides the miNs to distinct neuronal fates, a feature critical for modeling disorders that affect specific neuronal subtypes. Here, we describe the miR-9/9*-124-based neuronal reprogramming protocols for the generation of several disease-relevant neuronal subtypes: striatal medium spiny neurons, cortical neurons, and spinal cord motor neurons.


Assuntos
Reprogramação Celular/genética , MicroRNAs/metabolismo , Neurônios Motores/citologia , Neurogênese/genética , Fatores de Transcrição/metabolismo , Linhagem Celular , Células Cultivadas , Senescência Celular/genética , Corpo Estriado/citologia , Corpo Estriado/metabolismo , Meios de Cultura/química , Fibroblastos/citologia , Fibroblastos/metabolismo , Vetores Genéticos , Humanos , Lentivirus/genética , MicroRNAs/genética , Neurônios Motores/metabolismo , Neurônios/citologia , Neurônios/metabolismo , Medula Espinal/citologia , Medula Espinal/metabolismo , Fatores de Transcrição/genética
14.
J Neurosci ; 41(7): 1455-1469, 2021 02 17.
Artigo em Inglês | MEDLINE | ID: mdl-33376156

RESUMO

In the macaque brain, projections from distant, interconnected cortical areas converge in specific zones of the striatum. For example, specific zones of the motor putamen are targets of projections from frontal motor, inferior parietal, and ventrolateral prefrontal hand-related areas and thus are integral part of the so-called "lateral grasping network." In the present study, we analyzed the laminar distribution of corticostriatal neurons projecting to different parts of the motor putamen. Retrograde neural tracers were injected in different parts of the putamen in 3 Macaca mulatta (one male) and the laminar distribution of the labeled corticostriatal neurons was analyzed quantitatively. In frontal motor areas and frontal operculum, where most labeled cells were located, almost everywhere the proportion of corticostriatal labeled neurons in layers III and/or VI was comparable or even stronger than in layer V. Furthermore, within these regions, the laminar distribution pattern of corticostriatal labeled neurons largely varied independently from their density and from the projecting area/sector, but likely according to the target striatal zone. Accordingly, the present data show that cortical areas may project in different ways to different striatal zones, which can be targets of specific combinations of signals originating from the various cortical layers of the areas of a given network. These observations extend current models of corticostriatal interactions, suggesting more complex modes of information processing in the basal ganglia for different motor and nonmotor functions and opening new questions on the architecture of the corticostriatal circuitry.SIGNIFICANCE STATEMENT Projections from the ipsilateral cerebral cortex are the major source of input to the striatum. Previous studies have provided evidence for distinct zones of the putamen specified by converging projections from specific sets of interconnected cortical areas. The present study shows that the distribution of corticostriatal neurons in the various layers of the primary motor and premotor areas varies depending on the target striatal zone. Accordingly, different striatal zones collect specific combinations of signals from the various cortical layers of their input areas, possibly differing in terms of coding, timing, and direction of information flow (e.g., feed-forward, or feed-back).


Assuntos
Córtex Cerebral/fisiologia , Corpo Estriado/fisiologia , Vias Neurais/fisiologia , Putamen/fisiologia , Animais , Mapeamento Encefálico , Córtex Cerebral/citologia , Corpo Estriado/citologia , Retroalimentação Fisiológica/fisiologia , Feminino , Lobo Frontal/fisiologia , Macaca mulatta , Masculino , Córtex Motor/fisiologia , Vias Neurais/citologia , Neurônios/fisiologia , Lobo Parietal/fisiologia , Putamen/citologia
15.
Nat Biotechnol ; 38(12): 1421-1430, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33273741

RESUMO

Cortico-striatal projections are critical components of forebrain circuitry that regulate motivated behaviors. To enable the study of the human cortico-striatal pathway and how its dysfunction leads to neuropsychiatric disease, we developed a method to convert human pluripotent stem cells into region-specific brain organoids that resemble the developing human striatum and include electrically active medium spiny neurons. We then assembled these organoids with cerebral cortical organoids in three-dimensional cultures to form cortico-striatal assembloids. Using viral tracing and functional assays in intact or sliced assembloids, we show that cortical neurons send axonal projections into striatal organoids and form synaptic connections. Medium spiny neurons mature electrophysiologically following assembly and display calcium activity after optogenetic stimulation of cortical neurons. Moreover, we derive cortico-striatal assembloids from patients with a neurodevelopmental disorder caused by a deletion on chromosome 22q13.3 and capture disease-associated defects in calcium activity, showing that this approach will allow investigation of the development and functional assembly of cortico-striatal connectivity using patient-derived cells.


Assuntos
Córtex Cerebral/citologia , Corpo Estriado/citologia , Organoides/citologia , Células-Tronco Pluripotentes/citologia , Cálcio/metabolismo , Feminino , Humanos , Modelos Biológicos , Rede Nervosa/fisiologia , Optogenética , Fenótipo , Gravidez
16.
Mol Brain ; 13(1): 170, 2020 12 14.
Artigo em Inglês | MEDLINE | ID: mdl-33317605

RESUMO

BACKGROUND: Immune molecules, such as cytokines, complement, and major histocompatibility complex (MHC) proteins, in the central nervous system are often associated with neuropsychiatric disorders. Neuronal MHC class I (MHCI), such as H-2D, regulate neurite outgrowth, the establishment and function of cortical connections, and activity-dependent refinement in mice. We previously established mice expressing MHCI specifically in astrocytes of the media prefrontal cortex (mPFC) using the adeno-associated virus (AAV) vector under the control of the GfaABC1D promoter. Mice expressing the soluble form of H-2D (sH-2D) in the mPFC (sH-2D-expressing mice) showed abnormal behaviors, including social interaction deficits and cognitive dysfunctions. However, the pathophysiological significance of astroglial MHCI on higher brain functions, such as learning, memory, and behavioral flexibility, remains unclear. Therefore, cognitive function in mice expressing sH-2D in astrocytes of the mPFC was tested using the visual discrimination (VD) task. METHODS: sH-2D-expressing mice were subjected to the VD and reversal learning tasks, and morphological analysis. RESULTS: In the pretraining, sH-2D-expressing mice required significantly more trials to reach the learning criterion than control mice. The total number of sessions, trials, normal trials, and correction trials to reach the VD criterion were also significantly higher in sH-2D-expressing mice than in control mice. A morphological study showed that dendritic complexity and spine density were significantly reduced in the dorsal striatum of sH-2D-expressing mice. CONCLUSION: Collectively, the present results suggest that the overexpression of astroglial MHCI in the mPFC results in impaired VD learning, which may be accompanied by decreased dendritic complexity in the dorsal striatum and mPFC.


Assuntos
Astrócitos/metabolismo , Aprendizagem por Discriminação , Discriminação Psicológica , Complexo Principal de Histocompatibilidade , Córtex Pré-Frontal/metabolismo , Percepção Visual , Animais , Corpo Estriado/citologia , Espinhas Dendríticas/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Neurônios/metabolismo , Reversão de Aprendizagem , Solubilidade , Análise e Desempenho de Tarefas
17.
Elife ; 92020 11 20.
Artigo em Inglês | MEDLINE | ID: mdl-33215609

RESUMO

The posterior dorsomedial striatum (pDMS) is necessary for goal-directed action; however, the role of the direct (dSPN) and indirect (iSPN) spiny projection neurons in the pDMS in such actions remains unclear. In this series of experiments, we examined the role of pDMS SPNs in goal-directed action in rats and found that whereas dSPNs were critical for goal-directed learning and for energizing the learned response, iSPNs were involved in updating that learning to support response flexibility. Instrumental training elevated expression of the plasticity marker Zif268 in dSPNs only, and chemogenetic suppression of dSPN activity during training prevented goal-directed learning. Unilateral optogenetic inhibition of dSPNs induced an ipsilateral response bias in goal-directed action performance. In contrast, although initial goal-directed learning was unaffected by iSPN manipulations, optogenetic inhibition of iSPNs, but not dSPNs, impaired the updating of this learning and attenuated response flexibility after changes in the action-outcome contingency.


Assuntos
Corpo Estriado/fisiologia , Objetivos , Aprendizagem/fisiologia , Vias Neurais/fisiologia , Neurônios/fisiologia , Animais , Clozapina/análogos & derivados , Clozapina/farmacologia , Corpo Estriado/citologia , Corpo Estriado/efeitos dos fármacos , Proteína 1 de Resposta de Crescimento Precoce/genética , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , Comportamento Alimentar , Feminino , Corantes Fluorescentes , Antagonistas GABAérgicos/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/fisiologia , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Optogenética , Desempenho Psicomotor/fisiologia , Ratos , Ratos Long-Evans
18.
Cells ; 9(11)2020 11 09.
Artigo em Inglês | MEDLINE | ID: mdl-33182316

RESUMO

The classical motor symptoms of Parkinson's disease (PD) are caused by degeneration of dopaminergic neurons in the substantia nigra, which is followed by secondary dendritic pruning and spine loss at striatal medium spiny neurons (MSN). We hypothesize that these morphological changes at MSN underlie at least in part long-term motor complications in PD patients. In order to define the potential benefits and limitations of dopamine substitution, we tested in a mouse model whether dendritic pruning and spine loss can be reversible when dopaminergic axon terminals regenerate. In order to induce degeneration of nigrostriatal dopaminergic neurons we used the toxicity of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in C57BL/6J mice; 30 mg/kg MPTP was applied i.p. on five consecutive days. In order to assess the consequences of dopamine depletion, mice were analyzed 21 days after the last injection. In order to test reversibility of MSN changes we exploited the property of this model that striatal axon terminals regenerate by sprouting within 90 days and analyzed a second cohort 90 days after MPTP. Degeneration of dopaminergic neurons was confirmed by counting TH-positive neurons in the substantia nigra and by analyzing striatal catecholamines. Striatal catecholamine recovered 90 days after MPTP. MSN morphology was visualized by Golgi staining and quantified as total dendritic length, number of dendritic branch points, and density of dendritic spines. All morphological parameters of striatal MSN were reduced 21 days after MPTP. Statistical analysis indicated that dendritic pruning and the reduction of spine density represent two distinct responses to dopamine depletion. Ninety days after MPTP, all morphological changes recovered. Our findings demonstrate that morphological changes in striatal MSN resulting from dopamine depletion are reversible. They suggest that under optimal conditions, symptomatic dopaminergic therapy might be able to prevent maladaptive plasticity and long-term motor complications in PD patients.


Assuntos
Corpo Estriado/citologia , Dopamina/deficiência , Neurônios/citologia , 1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina , Animais , Axônios/metabolismo , Espinhas Dendríticas/metabolismo , Modelos Animais de Doenças , Modelos Lineares , Camundongos Endogâmicos C57BL , Fenótipo
19.
Elife ; 92020 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-33016873

RESUMO

Huntington's disease (HD) is a neurological disorder characterized by motor disturbances. HD pathology is most prominent in the striatum, the central hub of the basal ganglia. The cerebral cortex is the main striatal afferent, and progressive cortico-striatal disconnection characterizes HD. We mapped striatal network dysfunction in HD mice to ultimately modulate the activity of a specific cortico-striatal circuit to ameliorate motor symptoms and recover synaptic plasticity. Multimodal MRI in vivo indicates cortico-striatal and thalamo-striatal functional network deficits and reduced glutamate/glutamine ratio in the striatum of HD mice. Moreover, optogenetically-induced glutamate release from M2 cortex terminals in the dorsolateral striatum (DLS) was undetectable in HD mice and striatal neurons show blunted electrophysiological responses. Remarkably, repeated M2-DLS optogenetic stimulation normalized motor behavior in HD mice and evoked a sustained increase of synaptic plasticity. Overall, these results reveal that selective stimulation of the M2-DLS pathway can become an effective therapeutic strategy in HD.


Assuntos
Córtex Cerebral , Corpo Estriado , Estimulação Elétrica , Doença de Huntington/fisiopatologia , Animais , Córtex Cerebral/citologia , Córtex Cerebral/fisiologia , Córtex Cerebral/efeitos da radiação , Corpo Estriado/citologia , Corpo Estriado/fisiologia , Corpo Estriado/efeitos da radiação , Ácido Glutâmico/metabolismo , Camundongos , Atividade Motora/fisiologia , Plasticidade Neuronal/fisiologia , Neurônios/metabolismo , Neurônios/fisiologia , Optogenética
20.
Nature ; 586(7829): 417-423, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32999463

RESUMO

Microglia, the brain's resident macrophages, help to regulate brain function by removing dying neurons, pruning non-functional synapses, and producing ligands that support neuronal survival1. Here we show that microglia are also critical modulators of neuronal activity and associated behavioural responses in mice. Microglia respond to neuronal activation by suppressing neuronal activity, and ablation of microglia amplifies and synchronizes the activity of neurons, leading to seizures. Suppression of neuronal activation by microglia occurs in a highly region-specific fashion and depends on the ability of microglia to sense and catabolize extracellular ATP, which is released upon neuronal activation by neurons and astrocytes. ATP triggers the recruitment of microglial protrusions and is converted by the microglial ATP/ADP hydrolysing ectoenzyme CD39 into AMP; AMP is then converted into adenosine by CD73, which is expressed on microglia as well as other brain cells. Microglial sensing of ATP, the ensuing microglia-dependent production of adenosine, and the adenosine-mediated suppression of neuronal responses via the adenosine receptor A1R are essential for the regulation of neuronal activity and animal behaviour. Our findings suggest that this microglia-driven negative feedback mechanism operates similarly to inhibitory neurons and is essential for protecting the brain from excessive activation in health and disease.


Assuntos
Retroalimentação Fisiológica , Microglia/fisiologia , Inibição Neural , Neurônios/fisiologia , 5'-Nucleotidase/metabolismo , Potenciais de Ação , Adenosina/metabolismo , Monofosfato de Adenosina/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Antígenos CD/metabolismo , Apirase/metabolismo , Cálcio/metabolismo , Corpo Estriado/citologia , Corpo Estriado/fisiologia , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microglia/citologia , Inibição Neural/genética , Receptor A1 de Adenosina/metabolismo , Receptor Muscarínico M3/genética , Receptor Muscarínico M3/metabolismo , Fatores de Tempo
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