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1.
Nat Commun ; 11(1): 504, 2020 01 24.
Artigo em Inglês | MEDLINE | ID: mdl-31980629

RESUMO

Endogenous homeostatic mechanisms can restore normal neuronal function following cocaine-induced neuroadaptations. Such mechanisms may be exploited to develop novel therapies for cocaine addiction, but a molecular target has not yet been identified. Here we profiled mouse gene expression during early and late cocaine abstinence to identify putative regulators of neural homeostasis. Cocaine activated the transcription factor, Nr4a1, and its target gene, Cartpt, a key molecule involved in dopamine metabolism. Sustained activation of Cartpt at late abstinence was coupled with depletion of the repressive histone modification, H3K27me3, and enrichment of activating marks, H3K27ac and H3K4me3. Using both CRISPR-mediated and small molecule Nr4a1 activation, we demonstrated the direct causal role of Nr4a1 in sustained activation of Cartpt and in attenuation of cocaine-evoked behavior. Our findings provide evidence that targeting abstinence-induced homeostatic gene expression is a potential therapeutic target in cocaine addiction.


Assuntos
Comportamento Animal/efeitos dos fármacos , Cocaína/farmacologia , Epigênese Genética , Homeostase/efeitos dos fármacos , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/metabolismo , Animais , Sistemas CRISPR-Cas/genética , Cocaína/administração & dosagem , Epigênese Genética/efeitos dos fármacos , Feminino , Histonas/metabolismo , Homeostase/genética , Masculino , Camundongos Endogâmicos C57BL , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Fenilacetatos/farmacologia , Regiões Promotoras Genéticas/genética , Processamento de Proteína Pós-Traducional , Sinapsinas/metabolismo
2.
PLoS One ; 14(12): e0226368, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31830091

RESUMO

It has been revealed that ß-amyloid (Aß) is generated and released from the presynaptic terminals in activity-dependent manner. However, molecules modulating the presynaptic Aß generation remain elusive. Here we test the hypothesis that Synapsin 1 (Syn1) may acts as a modulator of the Aß production. Using biochemical and Förster resonance energy transfer (FRET)-based imaging approaches we have found that Syn1 knock down decreases, whereas (over)expression of Syn1 in cells increases the Aß levels. Mechanistically, Syn1 does not seem to affect the activity of Presenilin 1 (PS1)/γ-secretase, PS1 conformation, or the proximity between PS1 and amyloid precursor protein (APP). However, we found that Syn1 is involved in up-regulation of the ß-site APP cleaving enzyme 1 (BACE1)/ß-secretase activity and increases the APP/BACE1 interaction. Therefore, we conclude that Syn1 may promote Aß production via the modulation of BACE1.


Assuntos
Secretases da Proteína Precursora do Amiloide/metabolismo , Peptídeos beta-Amiloides/metabolismo , Ácido Aspártico Endopeptidases/metabolismo , Regulação da Expressão Gênica , Presenilina-1/metabolismo , Sinapsinas/metabolismo , Secretases da Proteína Precursora do Amiloide/genética , Animais , Ácido Aspártico Endopeptidases/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Presenilina-1/genética , Sinapsinas/genética
3.
Brain Struct Funct ; 224(9): 3263-3276, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31667576

RESUMO

Most vesicles in the interior of synaptic terminals are clustered in clouds close to active zone regions of the plasma membrane where exocytosis occurs. Electron-dense structures, termed bridges, have been reported between a small minority of pairs of neighboring vesicles within the clouds. Synapsin proteins have been implicated previously, but the existence of the bridges as stable structures in vivo has been questioned. Here we use electron tomography to show that the bridges are present but less frequent in synapsin knockouts compared to wildtype. An analysis of distances between neighbors in wildtype tomograms indicated that the bridges are strong enough to resist centrifugal forces likely induced by fixation with aldehydes. The results confirm that the bridges are stable structures and that synapsin proteins are involved in formation or stabilization.


Assuntos
Terminações Pré-Sinápticas/ultraestrutura , Sinapsinas/metabolismo , Vesículas Sinápticas/ultraestrutura , Animais , Camundongos Knockout , Modelos Neurológicos , Terminações Pré-Sinápticas/metabolismo , Sinapsinas/genética , Vesículas Sinápticas/metabolismo
4.
Biol Pharm Bull ; 42(9): 1545-1553, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31474714

RESUMO

The aim of the present study is to construct and characterize a novel three-dimensional culture system for mouse neurons using the functional polymer, FP001. Stereoscopically extended neurites were found in primary mouse cortical neurons cultured in the FP001-containing medium. Neurons cultured with FP001 were distributed throughout the medium of the observation range whereas neurons cultured without FP001 were distributed only on the bottom of the dish. These results demonstrated that neurons can be three-dimensionally cultured using the FP001-containing medium. The mRNA expression of the glutamatergic neuronal marker vesicular glutamate transporter 1 in neurons cultured in the FP001-containing medium were higher than that in neurons cultured in the FP001-free medium. Expression of the matured neuronal marker, microtubule-associated protein 2 (MAP2) a,b, and the synapse formation marker, Synapsin I, in neurons cultured with FP001 was also higher than that in neurons cultured without FP001. The expression pattern of MAP2a,b in neurons cultured with FP001, but not that in neurons cultured without FP001, was similar to that in the embryonic cerebral cortex. Exposure to glutamate significantly increased 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction activity in neurons cultured with FP001 compared to that in neurons cultured without FP001. These results suggested that glutamatergic neurotransmission in neurons three-dimensionally cultured in the FP001-containing medium may be upregulated compared to neurons two-dimensionally cultured in the FP001-free medium. Thus, neurons with the properties close to those in the embryonic brain could be obtained by three-dimensionally culturing neurons using FP001, compared to two-dimensional culture with a conventional adhesion method.


Assuntos
Técnicas de Cultura de Células/métodos , Córtex Cerebral/citologia , Meios de Cultura/química , Neurônios/citologia , Polissacarídeos Bacterianos/química , Animais , Córtex Cerebral/embriologia , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos ICR , Proteínas Associadas aos Microtúbulos/metabolismo , Neuritos/metabolismo , Neurônios/metabolismo , Sinapsinas/metabolismo
5.
Autoimmun Rev ; 18(10): 102367, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31404705

RESUMO

Aggregation of immuno-proteomic data reveals that i) herpesviruses and synaptic proteins -in particular Synapsin-1 and Bassoon - share a large number of hexapeptides that also recur in hundreds of epitopes experimentally validated as immunopositive in the human host, and ii) the shared peptides are also spread among human epilepsy-related proteins. The data indicate that cross-reactive processes may be associated with pathogenetic mechanisms in epilepsy, thus suggesting a role of autoimmunity in etiopathology of epilepsies after herpesvirus-infections.


Assuntos
Autoimunidade/imunologia , Epilepsia/etiologia , Epitopos/imunologia , Herpes Simples/complicações , Herpesviridae/imunologia , Fragmentos de Peptídeos/imunologia , Sinapsinas/imunologia , Animais , Reações Cruzadas , Epilepsia/patologia , Herpes Simples/imunologia , Humanos
6.
Elife ; 82019 07 31.
Artigo em Inglês | MEDLINE | ID: mdl-31364987

RESUMO

The number of neurotransmitter-filled vesicles released into the synaptic cleft with each action potential dictates the reliability of synaptic transmission. Variability of this fundamental property provides diversity of synaptic function across brain regions, but the source of this variability is unclear. The prevailing view is that release of a single (univesicular release, UVR) or multiple vesicles (multivesicular release, MVR) reflects variability in vesicle release probability, a notion that is well-supported by the calcium-dependence of release mode. However, using mouse brain slices, we now demonstrate that the number of vesicles released is regulated by the size of the readily-releasable pool, upstream of vesicle release probability. Our results point to a model wherein protein kinase A and its vesicle-associated target, synapsin, dynamically control release site occupancy to dictate the number of vesicles released without altering release probability. Together these findings define molecular mechanisms that control MVR and functional diversity of synaptic signaling.


Assuntos
Transmissão Sináptica , Vesículas Sinápticas/metabolismo , Animais , Cálcio/metabolismo , Sinalização do Cálcio , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Camundongos , Modelos Biológicos , Sinapsinas/metabolismo
7.
eNeuro ; 6(4)2019.
Artigo em Inglês | MEDLINE | ID: mdl-31387877

RESUMO

Synapse formation can be promoted by intense activity. At the Drosophila larval neuromuscular junction (NMJ), new synaptic boutons can grow acutely in response to patterned stimulation. We combined confocal imaging with electron microscopy and tomography to investigate the initial stages of growth and differentiation of new presynaptic boutons at the Drosophila NMJ. We found that the new boutons can form rapidly in intact larva in response to intense crawling activity, and we observed two different patterns of bouton formation and maturation. The first pathway involves the growth of filopodia followed by a formation of boutons that are initially devoid of synaptic vesicles (SVs) but filled with filamentous matrix. The second pathway involves rapid budding of synaptic boutons packed with SVs, and these more mature boutons are sometimes capable of exocytosis/endocytosis. We demonstrated that intense activity predominantly promotes the second pathway, i.e., budding of more mature boutons filled with SVs. We also showed that this pathway depends on synapsin (Syn), a neuronal protein which reversibly associates with SVs and mediates their clustering via a protein kinase A (PKA)-dependent mechanism. Finally, we took advantage of the temperature-sensitive mutant sei to demonstrate that seizure activity can promote very rapid budding of new boutons filled with SVs, and this process occurs at scale of minutes. Altogether, these results demonstrate that intense activity acutely and selectively promotes rapid budding of new relatively mature presynaptic boutons filled with SVs, and that this process is regulated via a PKA/Syn-dependent pathway.


Assuntos
Locomoção , Junção Neuromuscular/fisiologia , Terminações Pré-Sinápticas/fisiologia , Vesículas Sinápticas/fisiologia , Animais , Animais Geneticamente Modificados , Diferenciação Celular , Proteínas Quinases Dependentes de AMP Cíclico/fisiologia , Drosophila , Proteínas de Drosophila/fisiologia , Feminino , Masculino , Junção Neuromuscular/citologia , Junção Neuromuscular/crescimento & desenvolvimento , Junção Neuromuscular/ultraestrutura , Terminações Pré-Sinápticas/ultraestrutura , Sinapsinas/fisiologia
8.
J Biol Chem ; 294(37): 13606-13618, 2019 09 13.
Artigo em Inglês | MEDLINE | ID: mdl-31350335

RESUMO

Coronaviruses are enveloped, single-stranded RNA viruses that are distributed worldwide. They include transmissible gastroenteritis virus (TGEV), porcine epidemic diarrhea virus (PEDV), and the human coronaviruses severe acute respiratory syndrome coronavirus (SARS-CoV) and Middle East respiratory syndrome coronavirus (MERS-CoV), many of which seriously endanger human health and well-being. Only alphacoronaviruses and betacoronaviruses harbor nonstructural protein 1 (nsp1), which performs multiple functions in inhibiting antiviral host responses. The role of the C terminus of betacoronavirus nsp1 in virulence has been characterized, but the location of the alphacoronavirus nsp1 region that is important for virulence remains unclear. Here, using TGEV nsp1 as a model to explore the function of this protein in alphacoronaviruses, we demonstrate that alphacoronavirus nsp1 inhibits host gene expression. Solving the crystal structure of full-length TGEV at 1.85-Å resolution and conducting several biochemical analyses, we observed that a specific motif (amino acids 91-95) of alphacoronavirus nsp1 is a conserved region that inhibits host protein synthesis. Using a reverse-genetics system based on CRISPR/Cas9 technology to construct a recombinant TGEV in which this specific nsp1 motif was altered, we found that this mutation does not affect virus replication in cell culture but significantly reduces TGEV pathogenicity in pigs. Taken together, our findings suggest that alphacoronavirus nsp1 is an essential virulence determinant, providing a potential paradigm for the development of a new attenuated vaccine based on modified nsp1.


Assuntos
Alphacoronavirus/genética , Proteínas não Estruturais Virais/metabolismo , Replicação Viral/genética , Alphacoronavirus/patogenicidade , Animais , Betacoronavirus , Interações entre Hospedeiro e Microrganismos/genética , Interações entre Hospedeiro e Microrganismos/fisiologia , Humanos , Coronavírus da Síndrome Respiratória do Oriente Médio/genética , Vírus da Diarreia Epidêmica Suína/genética , Biossíntese de Proteínas , RNA Replicase/genética , RNA Replicase/metabolismo , RNA Replicase/ultraestrutura , Vírus da SARS/genética , Suínos , Sinapsinas/metabolismo , Vírus da Gastroenterite Transmissível/genética , Proteínas não Estruturais Virais/genética , Proteínas não Estruturais Virais/ultraestrutura , Virulência , Replicação Viral/fisiologia
9.
Neuroscience ; 414: 128-140, 2019 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-31283907

RESUMO

Tay-Sachs disease (TSD) is a GM2 gangliosidosis lysosomal storage disease caused by a loss of lysosomal hexosaminidase-A (HEXA) activity and characterized by progressive neurodegeneration due to the massive accumulation of GM2 ganglioside in the brain. Here, we generated iPSCs derived from patients with TSD, and found similar potential for neural differentiation between TSD-iPSCs and normal iPSCs, although neural progenitor cells (NPCs) derived from the TSD-iPSCs exhibited enlarged lysosomes and upregulation of the lysosomal marker, LAMP1, caused by the accumulation of GM2 ganglioside. The NPCs derived from TSD-iPSCs also had an increased incidence of oxidative stress-induced cell death. TSD-iPSC-derived neurons showed a decrease in exocytotic activity with the accumulation of GM2 ganglioside, suggesting deficient neurotransmission in TSD. Our findings demonstrated that NPCs and mature neurons derived from TSD-iPSCs are potentially useful cellular models of TSD and are useful for investigating the efficacy of drug candidates in the future.


Assuntos
Células-Tronco Pluripotentes Induzidas/fisiologia , Neurogênese/fisiologia , Neurônios/fisiologia , Terminações Pré-Sinápticas/fisiologia , Doença de Tay-Sachs/fisiopatologia , Humanos , Proteína 1 de Membrana Associada ao Lisossomo/metabolismo , Células-Tronco Neurais/fisiologia , Neuritos/fisiologia , Sinapsinas/metabolismo , Doença de Tay-Sachs/metabolismo , Regulação para Cima/fisiologia
10.
Invest Ophthalmol Vis Sci ; 60(7): 2676-2684, 2019 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-31242289

RESUMO

Purpose: To explore the role of synapsin-Ia/b in visual cortical plasticity, the dynamic changes in total protein expression (T-) and conventional protein kinase C (cPKC)γ-modulated phosphorylation (P-) levels of synapsin-Ia/b were observed in the developing visual cortex of mice. Methods: The Western blot analysis was used to determine the levels of T- and P-synapsin-Ia/b at site of Ser9, 549, and 603; the cPKCγ gene wild-type (cPKCγ+/+) and knockout (cPKCγ-/-) mice were applied to explore the modulation of cPKCγ on synapsin-Ia/b phosphorylation status in visual cortex of mice at postnatal 7 to 60 days (P7-P60, n = 6 per group). Results: The results showed that T-synapsin-Ia/b protein levels significantly increased at P14 to P35 and peaked at P42 to 60 (P < 0.001) in visual cortex when compared with that of P7 cPKCγ+/+ mice, and cPKCγ-/- did not affect this pattern of T-synapsin-Ia/b protein expressions. For synapsin-Ia/b phosphorylation status, the levels of P-Ser9 and 603 synapsin-Ia/b significantly elevated at P21 to P28 (P < 0.05 or 0.001), and then went down and maintained at lower levels at P35 to P60 (P < 0.05 or 0.001) compared with P7 cPKCγ+/+ mice. In addition, the cPKCγ gene knockout could significantly (P < 0.001) inhibit both the increase and decrease of P-Ser9 and 603 synapsin-Ia/b levels when compared with cPKCγ+/+ mice at P7 to P60. However, there were no significant changes of P-Ser549 synapsin-Ia/b in the developing visual cortex of both cPKCγ+/+ and cPKCγ-/- mice at P7 to P60. Conclusions: These results suggested that both protein expression levels and cPKCγ-modulated phosphorylation status at Ser9 and 603 of synapsin-Ia/b may play important role in developing visual cortex of mice.


Assuntos
Proteína Quinase C/farmacologia , Sinapsinas/metabolismo , Córtex Visual/efeitos dos fármacos , Animais , Western Blotting , Eletroforese em Gel de Poliacrilamida , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fosforilação , Isoformas de Proteínas , Córtex Visual/metabolismo
11.
J Neurochem ; 150(3): 264-281, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31148170

RESUMO

Leucine-rich repeat kinase 2 (LRRK2) is a large multidomain scaffolding protein with kinase and GTPase activities involved in synaptic vesicle (SV) dynamics. While its role in Parkinson's disease has been largely investigated, little is known about LRRK2 physiological role and until now few proteins have been described as substrates. We have previously demonstrated that LRRK2 through its WD40 domain interacts with synapsin I, an important SV-associated phosphoprotein involved in neuronal development and in the regulation of neurotransmitter release. To test whether synapsin I is substrate for LRRK2 and characterize the properties of its phosphorylation, we used in vitro kinase and binding assays as well as cellular model and site-direct mutagenesis. Using synaptosomes in superfusion, patch-clamp recordings in autaptic WT and synapsin I KO cortical neurons and SypHy assay on primary cortical culture from wild-type and BAC human LRRK2 G2019S mice we characterized the role of LRRK2 kinase activity on glutamate release and SV trafficking. Here we reported that synapsin I is phosphorylated by LRRK2 and demonstrated that the interaction between LRRK2 WD40 domain and synapsin I is crucial for this phosphorylation. Moreover, we showed that LRRK2 phosphorylation of synapsin I at threonine 337 and 339 significantly reduces synapsin I-SV/actin interactions. Using complementary experimental approaches, we demonstrated that LRRK2 controls glutamate release and SV dynamics in a kinase activity and synapsin I-dependent manner. Our findings show that synapsin I is a LRRK2 substrate and describe a novel mechanisms of regulation of glutamate release by LRRK2 kinase activity.


Assuntos
Ácido Glutâmico/metabolismo , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Sinapsinas/metabolismo , Transmissão Sináptica/fisiologia , Animais , Encéfalo/metabolismo , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neurônios/metabolismo , Fosforilação , Vesículas Sinápticas/metabolismo
12.
J Nat Med ; 73(4): 717-726, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31190266

RESUMO

Tau oligomers are the etiologic molecules of Alzheimer's disease, and correlate strongly with neuronal loss and exhibit neurotoxicity. Recent evidence indicates that small tau oligomers are the most relevant toxic aggregate species. The aim of the present study was to investigate the mechanisms of cornel iridoid glycoside (CIG) on tau oligomers and cognitive functions. We injected wortmannin and GF-109203X (WM/GFX, 200 µM each) into the lateral ventricles to induce tau oligomer and memory impairment in rats. When orally administered with CIG at 60 and 120 mg/kg/day for 14 days, CIG decreased the escape latency in the Morris water maze test. We also found that CIG restored the expression of presynaptic p-synapsin, synaptophysin, and postsynaptic density-95 (PSD-95) decreased by WM/GFX in rat cortex. CIG reduced the accumulation of tau oligomers in the brain of WM/GFX rats and in cells transfected with wild type glycogen synthase kinase-3ß (wtGSK-3ß). In addition, CIG up-regulated the levels of ATG7, ATG12, Beclin-1, and LC3II in vivo and in vitro, suggesting the restoration of autophagy function. These results suggest that CIG could ameliorate memory deficits and regulate memory-associated synaptic proteins through the clearance of tau oligomers accumulation. Moreover, CIG clears tau oligomers by restoring autophagy function.


Assuntos
Doença de Alzheimer/patologia , Glicogênio Sintase Quinase 3 beta/metabolismo , Glicosídeos Iridoides/farmacologia , Transtornos da Memória/patologia , Proteínas tau/toxicidade , Animais , Autofagia/efeitos dos fármacos , Encéfalo/metabolismo , Linhagem Celular Tumoral , Proteína 4 Homóloga a Disks-Large/metabolismo , Indóis/toxicidade , Masculino , Maleimidas/toxicidade , Aprendizagem em Labirinto/efeitos dos fármacos , Memória/efeitos dos fármacos , Camundongos , Fosforilação , Substâncias Protetoras , Ratos , Ratos Wistar , Sinapsinas/metabolismo , Sinaptofisina/metabolismo , Wortmanina/toxicidade
13.
Cells ; 8(6)2019 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-31212628

RESUMO

Direct conversion of non-neural cells into induced neurons holds great promise for brain repair. As the most common malignant tumor in the central nervous system, glioma is currently incurable due to its exponential growth and invasive behavior. Given that neurons are irreversible postmitotic cells, reprogramming glioma cells into terminally differentiated neuron-like cells represents a potential approach to inhibit brain tumor development. We here show that human glioma cells can be directly, rapidly and efficiently reprogrammed into terminally differentiated neuron-like cells by the single transcription factor ASCL1 (Achaete-scute complex-like 1, also known as MASH1). These induced cells exhibit typical neuron-like morphology and express multiple neuron-specific markers. Importantly, ASCL1-mediated neuronal reprogramming drives human glioma cells to exit the cell cycle and results in dramatic inhibition of proliferation, both in vitro and in vivo. Taken together, this proof-of-principle study demonstrates a potential strategy for impeding brain tumor development by ASCL1-induced direct neuronal reprogramming.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Reprogramação Celular , Glioma/patologia , Neurônios/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Linhagem Celular Tumoral , Proliferação de Células , Regulação da Expressão Gênica , Glioma/metabolismo , Glioma/mortalidade , Humanos , Estimativa de Kaplan-Meier , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Neurogênese , Neurônios/citologia , Neuropeptídeos/genética , Neuropeptídeos/metabolismo , Sinapsinas/metabolismo , Transplante Heterólogo , Tubulina (Proteína)/metabolismo
14.
Int J Neuropsychopharmacol ; 22(7): 453-465, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-31188434

RESUMO

BACKGROUND: Synapsins are encoded by SYN I, SYN II, and SYN III, and they regulate neurotransmitter release by maintaining a reserve pool of synaptic vesicles. METHODS: Presynaptic dopamine responses to cocaine were examined by microdialysis, and postsynaptic responses were evaluated to various dopamine receptor agonists in the open field with SynI/SynII/SynIII triple knockout mice. RESULTS: Triple knockout mice showed enhanced spontaneous locomotion in a novel environment and were hyper-responsive to indirect and direct D1 and D2 dopamine agonists. Triple knockout animals appeared sensitized to cocaine upon first open field exposure; sensitization developed across days in wild-type controls. When mutants were preexposed to a novel environment before injection, cocaine-stimulated locomotion was reduced and behavioral sensitization retarded. Baseline dopamine turnover was enhanced in mutants and novel open field exposure increased their striatal dopamine synthesis rates. As KCl-depolarization stimulated comparable dopamine release in both genotypes, their readily releasable pools appeared indistinguishable. Similarly, cocaine-induced hyperlocomotion was indifferent to blockade of newly synthesized dopamine and depletion of releasable dopamine pools. Extracellular dopamine release was similar in wild-type and triple knockout mice preexposed to the open field and given cocaine or placed immediately into the arena following injection. Since motor effects to novelty and psychostimulants depend upon frontocortical-striatal inputs, we inhibited triple knockout medial frontal cortex with GABA agonists. Locomotion was transiently increased in cocaine-injected mutants, while their supersensitive cocaine response to novelty was lost. CONCLUSIONS: These results reveal presynaptic dopamine release is not indicative of agonist-induced triple knockout hyperlocomotion. Instead, their novelty response occurs primarily through postsynaptic mechanisms and network effects.


Assuntos
Estimulantes do Sistema Nervoso Central/farmacologia , Agonistas de Dopamina/farmacologia , Dopamina/metabolismo , Atividade Motora/efeitos dos fármacos , Sinapses/metabolismo , Sinapsinas/deficiência , Animais , Cocaína/farmacologia , Inibidores da Captação de Dopamina/farmacologia , Espaço Extracelular/efeitos dos fármacos , Espaço Extracelular/metabolismo , Feminino , Lobo Frontal/efeitos dos fármacos , Lobo Frontal/metabolismo , Agonistas GABAérgicos/farmacologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microdiálise , Atividade Motora/fisiologia , Sinapsinas/genética
15.
Eur J Histochem ; 63(2)2019 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-31060348

RESUMO

Cognitive impairment in Alzheimer's disease (AD) is usually accompanied by synaptic loss in both the hippocampus and neocortex. In the early stage of AD, amyloid ß-induced synapse changes is the main reason, while in the later stage, the accumulation of Tau protein promotes synapse degeneration as the key factor leading to dementia. MicroRNA (miRNA) is closely related to the expression changes of many AD-related genes. One of the most abundant brain-enriched miRNAs is miR-132, which has been shown to regulate both neuron morphogenesis and plasticity. It has been reported that miR-132 is significantly reduced in the brains of Alzheimer's patients. Genetic deletion of miR-132 in mice promotes Aß deposition, leading to impaired memory and enhanced Tau pathology, but how the miRNA-mediated gene expression dysregulation contributes to AD pathology remains unclear. Here we found the possible downstream target of miR-132 by in silico analysis, namely C1q. C1q is the primary protein of classical complement cascade, which is highly expressed in the synaptic regions of the central nervous system in Alzheimer's patients. However, it is not clear whether miR-132 plays a role in AD through regulating C1q. To address this question, the APP/PS1 transgenic mice were transfected with miR-132 and given C1 inhibitors. Behavior tests were conducted to assess memory and cognitive abilities seven days after administration. In addition, we analyzed the expression of PSD95, Synapsin-1 and phosphorylated (p)-Synapsin. We found that the expression levels of the synaptic proteins treated with miR-132 or C1INH were significantly increased compared with the AD group. Further RT-qPCR result suggested that miR-132 might regulate C1q expression in AD.


Assuntos
Precursor de Proteína beta-Amiloide/genética , Complemento C1q/metabolismo , MicroRNAs/metabolismo , Presenilina-1/genética , Doença de Alzheimer/metabolismo , Doença de Alzheimer/fisiopatologia , Animais , Cognição/efeitos dos fármacos , Complemento C1q/antagonistas & inibidores , Complemento C1q/genética , Proteína 4 Homóloga a Disks-Large/genética , Proteína 4 Homóloga a Disks-Large/metabolismo , Reposicionamento de Medicamentos , Regulação da Expressão Gênica , Masculino , Memória/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Sinapsinas/genética , Sinapsinas/metabolismo
16.
Proc Natl Acad Sci U S A ; 116(23): 11116-11118, 2019 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-31110014

RESUMO

The normal function of α-synuclein (α-syn) remains elusive. Although recent studies suggest α-syn as a physiologic attenuator of synaptic vesicle (SV) recycling, mechanisms are unclear. Here, we show that synapsin-a cytosolic protein with known roles in SV mobilization and clustering-is required for presynaptic functions of α-syn. Our data offer a critical missing link and advocate a model where α-syn and synapsin cooperate to cluster SVs and attenuate recycling.


Assuntos
Sinapsinas/metabolismo , Vesículas Sinápticas/metabolismo , alfa-Sinucleína/metabolismo , Animais , Linhagem Celular , Camundongos , Neurônios/metabolismo , Sinapses/metabolismo
17.
Elife ; 82019 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-31081751

RESUMO

Information processing by cerebellar molecular layer interneurons (MLIs) plays a crucial role in motor behavior. MLI recruitment is tightly controlled by the profile of short-term plasticity (STP) at granule cell (GC)-MLI synapses. While GCs are the most numerous neurons in the brain, STP diversity at GC-MLI synapses is poorly documented. Here, we studied how single MLIs are recruited by their distinct GC inputs during burst firing. Using slice recordings at individual GC-MLI synapses of mice, we revealed four classes of connections segregated by their STP profile. Each class differentially drives MLI recruitment. We show that GC synaptic diversity is underlain by heterogeneous expression of synapsin II, a key actor of STP and that GC terminals devoid of synapsin II are associated with slow MLI recruitment. Our study reveals that molecular, structural and functional diversity across GC terminals provides a mechanism to expand the coding range of MLIs.


Assuntos
Cerebelo/citologia , Cerebelo/fisiologia , Rede Nervosa/fisiologia , Plasticidade Neuronal/fisiologia , Neurônios/fisiologia , Animais , Camundongos , Sinapsinas/metabolismo
18.
Neuroreport ; 30(8): 544-549, 2019 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-30964765

RESUMO

Early exposure to anesthetics may interfere with synaptic development and lead to cognitive deficits. We previously demonstrated a decrease in vesicles docked at and within 100 nm from the presynaptic membrane in hippocampal nerve terminals of neonatal rats after anesthesia. Hence, we designed this study to assess the effects of neonatal anesthesia on synapsin 1 (Syn1) and synaptotagmin 1 (Syt1), two key regulators of vesicle docking and fusion. To test the link between changes in Syn1 and Syt1 and behavioral deficits observed after neonatal anesthesia, we also assessed retention memory and fear conditioning in adolescent rats after neonatal anesthesia. Pups received a combination of clinical anesthetics, then Syn1 and Syt1 mRNA and protein expression were determined at the peak (postnatal day 8, P8), part-way through (P12) and end of synaptogenesis (P24) in the CA1-subiculum by qPCR and western blotting. Anesthesia decreased Syn1 and Syt1 mRNA expression at P8 (P<0.01 and <0.001) and P12 (P=0.001 and 0.017), but not P24 (P=0.538 and 0.671), and impaired Syn1, p-Syn1, and Syt1 protein levels at P8 (P=0.038, 0.041, and 0.004, respectively), P12 (P<0.001, P=0.001, and P<0.0001), and P24 (P=0.025, 0.031, and 0.001). Anesthetic-challenged rats displayed deficient long-term retention memory (P=0.019) and hippocampus-dependent fear conditioning (P<0.001). These results suggest that anesthetics alter Syn1 and Syt1 during synapse assembly and maturation, raising the possibility that anesthetic interference with Syn1 and Syt1 could initiate changes in synaptic function that contribute to the cognitive deficits observed after neonatal anesthesia.


Assuntos
Anestésicos Inalatórios/administração & dosagem , Região CA1 Hipocampal/efeitos dos fármacos , Isoflurano/administração & dosagem , Sinapsinas/metabolismo , Vesículas Sinápticas/efeitos dos fármacos , Sinaptotagmina I/metabolismo , Animais , Animais Recém-Nascidos , Região CA1 Hipocampal/metabolismo , Feminino , Masculino , Ratos Sprague-Dawley , Memória Espacial/efeitos dos fármacos , Transmissão Sináptica/efeitos dos fármacos , Vesículas Sinápticas/metabolismo
19.
Mol Autism ; 10: 19, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31011411

RESUMO

Background: Autism spectrum disorders (ASD) affect around 1.5% of people worldwide. Symptoms start around age 2, when children fail to maintain eye contact and to develop speech and other forms of communication. Disturbances in glutamatergic and GABAergic signaling that lead to synaptic changes and alter the balance between excitation and inhibition in the developing brain are consistently found in ASD. One of the hallmarks of these disorders is hypersensitivity to sensory stimuli; however, little is known about its underlying causes. Since the retina is the part of the CNS that converts light into a neuronal signal, we set out to study how it is affected in adolescent mice prenatally exposed to valproic acid (VPA), a useful tool to study ASD endophenotypes. Methods: Pregnant female mice received VPA (600 mg/kg, ip) or saline at gestational day 11. Their male adolescent pups (P29-35) were behaviorally tested for anxiety and social interaction. Proteins known to be related with ASD were quantified and visualized in their retinas by immunoassays, and retinal function was assessed by full-field scotopic electroretinograms (ERGs). Results: Early adolescent mice prenatally exposed to VPA displayed impaired social interest and increased anxiety-like behaviors consistent with an ASD phenotype. The expression of GABA, GAD, synapsin-1, and FMRP proteins were reduced in their retinas, while mGluR5 was increased. The a-wave amplitudes of VPA-exposed were smaller than those of CTR animals, whereas the b-wave and oscillatory potentials were normal. Conclusions: This study establishes that adolescent male mice of the VPA-induced ASD model have alterations in retinal function and protein expression compatible with those found in several brain areas of other autism models. These results support the view that synaptic disturbances with excitatory/inhibitory imbalance early in life are associated with ASD and point to the retina as a window to understand their subjacent mechanisms.


Assuntos
Transtorno do Espectro Autista/fisiopatologia , Retina/metabolismo , Potenciais de Ação , Animais , Transtorno do Espectro Autista/etiologia , Transtorno do Espectro Autista/metabolismo , Endofenótipos , Feminino , Proteína do X Frágil de Retardo Mental/genética , Proteína do X Frágil de Retardo Mental/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Gravidez , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Efeitos Tardios da Exposição Pré-Natal/fisiopatologia , Receptores de GABA/genética , Receptores de GABA/metabolismo , Receptores de Glutamato/genética , Receptores de Glutamato/metabolismo , Retina/fisiopatologia , Comportamento Social , Sinapsinas/genética , Sinapsinas/metabolismo , Ácido Valproico/toxicidade
20.
J Agric Food Chem ; 67(10): 3006-3017, 2019 Mar 13.
Artigo em Inglês | MEDLINE | ID: mdl-30816709

RESUMO

Alzheimer's disease (AD) is closely related to gut microbial alteration. Prebiotic fructooligosaccharides (FOS) play major roles by regulating gut microbiota. The present study aimed to explore the effect and mechanism of FOS protection against AD via regulating gut microbiota. Male Apse/PSEN 1dE9 (APP/PS1) transgenic (Tg) mice were administrated with FOS for 6 weeks. Cognitive deficits and amyloid deposition were evaluated. The levels of synaptic plasticity markers including postsynaptic density protein 95 (PSD-95) and synapsin I, as well as phosphorylation of c-Jun N-terminal kinase (JNK), were determined. The intestinal microbial constituent was detected by 16S rRNA sequencing. Moreover, the levels of glucagon-like peptide-1 (GLP-1) in the gut and GLP-1 receptor (GLP-1R) in the brain were measured. The results indicated that FOS treatment ameliorated cognitive deficits and pathological changes in the Tg mice. FOS significantly upregulated the expression levels of synapsin I and PSD-95, as well as decreased phosphorylated level of JNK. The sequencing results showed that FOS reversed the altered microbial composition. Furthermore, FOS increased the level of GLP-1 and decreased the level of GLP-1R in the Tg mice. These findings indicated that FOS exerted beneficial effects against AD via regulating the gut microbiota-GLP-1/GLP-1R pathway.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/microbiologia , Microbioma Gastrointestinal/efeitos dos fármacos , Oligossacarídeos/administração & dosagem , Prebióticos/administração & dosagem , Doença de Alzheimer/metabolismo , Doença de Alzheimer/psicologia , Peptídeos beta-Amiloides/genética , Peptídeos beta-Amiloides/metabolismo , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Cognição , Modelos Animais de Doenças , Proteína 4 Homóloga a Disks-Large/metabolismo , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Plasticidade Neuronal , Presenilina-1/genética , Presenilina-1/metabolismo , Sinapsinas/metabolismo
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