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1.
Nat Commun ; 11(1): 4958, 2020 10 02.
Artigo em Inglês | MEDLINE | ID: mdl-33009395

RESUMO

Striatal dopamine (DA) is critical for action and learning. Recent data show that DA release is under tonic inhibition by striatal GABA. Ambient striatal GABA tone on striatal projection neurons can be determined by plasma membrane GABA uptake transporters (GATs) located on astrocytes and neurons. However, whether striatal GATs and astrocytes determine DA output are unknown. We reveal that DA release in mouse dorsolateral striatum, but not nucleus accumbens core, is governed by GAT-1 and GAT-3. These GATs are partly localized to astrocytes, and are enriched in dorsolateral striatum compared to accumbens core. In a mouse model of early parkinsonism, GATs are downregulated, tonic GABAergic inhibition of DA release augmented, and nigrostriatal GABA co-release attenuated. These data define previously unappreciated and important roles for GATs and astrocytes in supporting DA release in striatum, and reveal a maladaptive plasticity in early parkinsonism that impairs DA output in vulnerable striatal regions.


Assuntos
Corpo Estriado/metabolismo , Dopamina/metabolismo , Regulação para Baixo , Proteínas da Membrana Plasmática de Transporte de GABA/metabolismo , Transtornos Parkinsonianos/metabolismo , Ácido gama-Aminobutírico/metabolismo , Animais , Astrócitos/metabolismo , Membrana Celular/metabolismo , Modelos Animais de Doenças , Glutamato Descarboxilase/metabolismo , Camundongos Endogâmicos C57BL , Modelos Biológicos , Núcleo Accumbens/metabolismo
2.
Nat Commun ; 11(1): 5073, 2020 10 08.
Artigo em Inglês | MEDLINE | ID: mdl-33033238

RESUMO

Brain cells continuously produce and release protons into the extracellular space, with the rate of acid production corresponding to the levels of neuronal activity and metabolism. Efficient buffering and removal of excess H+ is essential for brain function, not least because all the electrogenic and biochemical machinery of synaptic transmission is highly sensitive to changes in pH. Here, we describe an astroglial mechanism that contributes to the protection of the brain milieu from acidification. In vivo and in vitro experiments conducted in rodent models show that at least one third of all astrocytes release bicarbonate to buffer extracellular H+ loads associated with increases in neuronal activity. The underlying signalling mechanism involves activity-dependent release of ATP triggering bicarbonate secretion by astrocytes via activation of metabotropic P2Y1 receptors, recruitment of phospholipase C, release of Ca2+ from the internal stores, and facilitated outward HCO3- transport by the electrogenic sodium bicarbonate cotransporter 1, NBCe1. These results show that astrocytes maintain local brain extracellular pH homeostasis via a neuronal activity-dependent release of bicarbonate. The data provide evidence of another important metabolic housekeeping function of these glial cells.


Assuntos
Astrócitos/metabolismo , Bicarbonatos/metabolismo , Encéfalo/metabolismo , Espaço Extracelular/metabolismo , Acetazolamida/farmacologia , Trifosfato de Adenosina/metabolismo , Animais , Astrócitos/efeitos dos fármacos , Anidrases Carbônicas/metabolismo , Células Cultivadas , Estimulação Elétrica , Fluorescência , Hipocampo/metabolismo , Concentração de Íons de Hidrogênio , Camundongos Endogâmicos C57BL , Modelos Biológicos , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Antagonistas Purinérgicos/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Sprague-Dawley , Receptores Purinérgicos/metabolismo , Transdução de Sinais , Simportadores de Sódio-Bicarbonato/metabolismo
3.
Annu Int Conf IEEE Eng Med Biol Soc ; 2020: 2245-2248, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-33018454

RESUMO

Recently, the study of communication in an 'Astrocyte Network' has been suggested to be of equal importance to that of the traditional 'Neural Network'. In this paper, for the first time, we use nanosecond laser stimulation to stimulate the central cell in an organized grid network of connected human astrocytes in order to observe calcium wave propagation at the single-cell level. We show that the calcium waves indeed propagate from the central astrocyte to the outer periphery of the organized astrocyte network. We observe also, like astrocytes in standard in vitro petri dishes, that the calcium wave propagates through specific connections to the outer periphery of cells rather than in a uniform radial manner predicted by mathematical theory. The results show that such a platform provides an excellent environment to perform repeatable, controlled studies of calcium wave signal propagation through an organized grid network of human astrocytes at single-cell resolution.


Assuntos
Astrócitos , Sinalização do Cálcio , Astrócitos/metabolismo , Cálcio/metabolismo , Humanos , Lasers
4.
J Toxicol Sci ; 45(9): 559-567, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32879255

RESUMO

Lead is a main threat to human health due to its neurotoxicity and the astrocyte is known to be a common deposit site of lead in vivo. However, the detailed mechanisms related to lead exposure in the astrocytes were unclear. In order to deeply investigate this issue, we used Sprague-Dawley (SD) rats and astrocytes isolated from the hippocampus of SD rats to establish the lead-exposed animal and cell models through treating with lead acetate. The expression levels of GFAP, LC3, and p62 in the rat hippocampus were detected by immunofluorescence and Western blot after lead exposure. The effects of autophagy on lead-exposed astrocytes were studied by further autophagy inhibitor 3-methyladenine (3-MA) induction. Transmission electron microscopy was used to observe autophagosomes in astrocytes after lead acetate treatment, followed by assessing related autophagy protein markers. In addition, some inflammatory cytokines and oxidative stress markers were also evaluated after lead exposure and 3-MA administration. We found that lead exposure induced activation of astrocytes, as evidenced by increased GFAP levels and GFAP-positive staining cells in the rat hippocampus. Moreover, lead exposure induced autophagy in astrocytes, as evidenced by increased LC3II and Beclin 1 protein levels and decreased p62 expression in both the rat hippocampus and astrocytes, and it was confirmed that this autophagy was activated through blocking the downstream Akt/target of the rapamycin (mTOR) pathway in astrocytes. Furthermore, it was shown that treatment of lead acetate increased the release of tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß), and the accumulation of malondialdehyde (MDA) and myeloperoxidase (MPO) in astrocytes, which could be alleviated by further 3-MA induction. Therefore, we conclude that lead exposure can induce the autophagy of astrocytes via blocking the Akt/mTOR pathway, leading to accelerated release of inflammatory factors and oxidative stress indicators in astrocytes.


Assuntos
Astrócitos/metabolismo , Astrócitos/fisiologia , Autofagia/efeitos dos fármacos , Autofagia/genética , Compostos Organometálicos/toxicidade , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismo , Animais , Células Cultivadas , Proteína Glial Fibrilar Ácida/genética , Proteína Glial Fibrilar Ácida/metabolismo , Hipocampo/citologia , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Masculino , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Estresse Oxidativo/genética , Ratos Sprague-Dawley , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo
5.
Nat Commun ; 11(1): 4855, 2020 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-32978390

RESUMO

The atypical chemokine receptor 3 (ACKR3) plays a pivotal role in directing the migration of various cellular populations and its over-expression in tumors promotes cell proliferation and invasiveness. The intracellular signaling pathways transducing ACKR3-dependent effects remain poorly characterized, an issue we addressed by identifying the interactome of ACKR3. Here, we report that recombinant ACKR3 expressed in HEK293T cells recruits the gap junction protein Connexin 43 (Cx43). Cx43 and ACKR3 are co-expressed in mouse brain astrocytes and human glioblastoma cells and form a complex in embryonic mouse brain. Functional in vitro studies show enhanced ACKR3 interaction with Cx43 upon ACKR3 agonist stimulation. Furthermore, ACKR3 activation promotes ß-arrestin2- and dynamin-dependent Cx43 internalization to inhibit gap junctional intercellular communication in primary astrocytes. These results demonstrate a functional link between ACKR3 and gap junctions that might be of pathophysiological relevance.


Assuntos
Astrócitos/metabolismo , Comunicação Celular/fisiologia , Conexina 43/metabolismo , Junções Comunicantes/patologia , Receptores CXCR/metabolismo , Animais , Proliferação de Células , Conexina 43/efeitos dos fármacos , Conexinas/metabolismo , Técnicas de Introdução de Genes , Glioblastoma/metabolismo , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Domínios e Motivos de Interação entre Proteínas , Receptores CXCR/agonistas , Receptores CXCR/genética , Transdução de Sinais/fisiologia
6.
PLoS One ; 15(9): e0236611, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32941446

RESUMO

Treatment of diseases that affect the CNS by gene therapy requires delivery of oligonucleotides to target cells within the brain. As the blood brain barrier prevents movement of large biomolecules, current approaches involve direct injection of the oligonucleotides, which is invasive and may have only a localised effect. The aim of this study was to investigate the potential of 2 nm galactose-coated gold nanoparticles (NP-Gal) as a delivery system of oligonucleotides across brain endothelium. DNA oligonucleotides of different types were attached to NP-Gal by the place exchange reaction and were characterised by EMSA (electrophoretic mobility shift assay). Several nanoparticle formulations were created, with single- or double-stranded (20nt or 40nt) DNA oligonucleotides, or with different amounts of DNA attached to the carriers. These nanocarriers were applied to transwell cultures of human brain endothelium in vitro (hCMEC/D3 cell-line) or to a 3D-hydrogel model of the blood-brain barrier including astrocytes. Transfer rates were measured by quantitative electron microscopy for the nanoparticles and qPCR for DNA. Despite the increase in nanoparticle size caused by attachment of oligonucleotides to the NP-Gal carrier, the rates of endocytosis and transcytosis of nanoparticles were both considerably increased when they carried an oligonucleotide cargo. Carriers with 40nt dsDNA were most efficient, accumulating in vesicles, in the cytosol and beneath the basal membrane of the endothelium. The oligonucleotide cargo remained attached to the nanocarriers during transcytosis and the transport rate across the endothelial cells was increased at least 50fold compared with free DNA. The nanoparticles entered the extracellular matrix and were taken up by the astrocytes in biologically functional amounts. Attachment of DNA confers a strong negative charge to the nanoparticles which may explain the enhanced binding to the endothelium and transcytosis by both vesicular transport and the transmembrane/cytosol pathway. These gold nanoparticles have the potential to transport therapeutic amounts of nucleic acids into the CNS.


Assuntos
Barreira Hematoencefálica/metabolismo , Células Endoteliais/metabolismo , Nanopartículas Metálicas/química , Oligodesoxirribonucleotídeos/metabolismo , Astrócitos/metabolismo , Linhagem Celular , Células Cultivadas , Galactose/química , Ouro/química , Humanos , Oligodesoxirribonucleotídeos/administração & dosagem
7.
Nat Commun ; 11(1): 4458, 2020 09 07.
Artigo em Inglês | MEDLINE | ID: mdl-32895383

RESUMO

In rodent models of type 2 diabetes (T2D), sustained remission of hyperglycemia can be induced by a single intracerebroventricular (icv) injection of fibroblast growth factor 1 (FGF1), and the mediobasal hypothalamus (MBH) was recently implicated as the brain area responsible for this effect. To better understand the cellular response to FGF1 in the MBH, we sequenced >79,000 single-cell transcriptomes from the hypothalamus of diabetic Lepob/ob mice obtained on Days 1 and 5 after icv injection of either FGF1 or vehicle. A wide range of transcriptional responses to FGF1 was observed across diverse hypothalamic cell types, with glial cell types responding much more robustly than neurons at both time points. Tanycytes and ependymal cells were the most FGF1-responsive cell type at Day 1, but astrocytes and oligodendrocyte lineage cells subsequently became more responsive. Based on histochemical and ultrastructural evidence of enhanced cell-cell interactions between astrocytes and Agrp neurons (key components of the melanocortin system), we performed a series of studies showing that intact melanocortin signaling is required for the sustained antidiabetic action of FGF1. These data collectively suggest that hypothalamic glial cells are leading targets for the effects of FGF1 and that sustained diabetes remission is dependent on intact melanocortin signaling.


Assuntos
Diabetes Mellitus Experimental/dietoterapia , Diabetes Mellitus Tipo 2/tratamento farmacológico , Fator 1 de Crescimento de Fibroblastos/administração & dosagem , Hipoglicemiantes/administração & dosagem , Hipotálamo/efeitos dos fármacos , Proteínas Recombinantes/administração & dosagem , Proteína Relacionada com Agouti/metabolismo , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Glicemia/análise , Comunicação Celular , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/etiologia , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/etiologia , Diabetes Mellitus Tipo 2/patologia , Dieta Hiperlipídica/efeitos adversos , Sacarose na Dieta/administração & dosagem , Sacarose na Dieta/efeitos adversos , Humanos , Hipotálamo/citologia , Hipotálamo/patologia , Injeções Intraventriculares , Leptina/genética , Masculino , Melanocortinas/metabolismo , Hormônios Estimuladores de Melanócitos/administração & dosagem , Camundongos , Camundongos Knockout , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Oligodendroglia/efeitos dos fármacos , Oligodendroglia/metabolismo , RNA-Seq , Receptor Tipo 4 de Melanocortina/genética , Receptores de Melanocortina/antagonistas & inibidores , Receptores de Melanocortina/metabolismo , Indução de Remissão/métodos , Transdução de Sinais/efeitos dos fármacos , Análise de Célula Única , Técnicas Estereotáxicas , Transcriptoma/efeitos dos fármacos
8.
Nat Commun ; 11(1): 4388, 2020 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-32873805

RESUMO

Presynaptic spike timing-dependent long-term depression (t-LTD) at hippocampal CA3-CA1 synapses is evident until the 3rd postnatal week in mice, disappearing during the 4th week. At more mature stages, we found that the protocol that induced t-LTD induced t-LTP. We characterized this form of t-LTP and the mechanisms involved in its induction, as well as that driving this switch from t-LTD to t-LTP. We found that this t-LTP is expressed presynaptically at CA3-CA1 synapses, as witnessed by coefficient of variation, number of failures, paired-pulse ratio and miniature responses analysis. Additionally, this form of presynaptic t-LTP does not require NMDARs but the activation of mGluRs and the entry of Ca2+ into the postsynaptic neuron through L-type voltage-dependent Ca2+ channels and the release of Ca2+ from intracellular stores. Nitric oxide is also required as a messenger from the postsynaptic neuron. Crucially, the release of adenosine and glutamate by astrocytes is required for t-LTP induction and for the switch from t-LTD to t-LTP. Thus, we have discovered a developmental switch of synaptic transmission from t-LTD to t-LTP at hippocampal CA3-CA1 synapses in which astrocytes play a central role and revealed a form of presynaptic LTP and the rules for its induction.


Assuntos
Astrócitos/metabolismo , Hipocampo/crescimento & desenvolvimento , Potenciação de Longa Duração/fisiologia , Transmissão Sináptica/fisiologia , Adenosina/metabolismo , Animais , Feminino , Ácido Glutâmico/metabolismo , Hipocampo/citologia , Masculino , Camundongos , Técnicas de Patch-Clamp , Receptores de Glutamato Metabotrópico/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo
9.
Nat Commun ; 11(1): 4411, 2020 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-32879313

RESUMO

The glymphatic system is a network of perivascular spaces that promotes movement of cerebrospinal fluid (CSF) into the brain and clearance of metabolic waste. This fluid transport system is supported by the water channel aquaporin-4 (AQP4) localized to vascular endfeet of astrocytes. The glymphatic system is more effective during sleep, but whether sleep timing promotes glymphatic function remains unknown. We here show glymphatic influx and clearance exhibit endogenous, circadian rhythms peaking during the mid-rest phase of mice. Drainage of CSF from the cisterna magna to the lymph nodes exhibits daily variation opposite to glymphatic influx, suggesting distribution of CSF throughout the animal depends on time-of-day. The perivascular polarization of AQP4 is highest during the rest phase and loss of AQP4 eliminates the day-night difference in both glymphatic influx and drainage to the lymph nodes. We conclude that CSF distribution is under circadian control and that AQP4 supports this rhythm.


Assuntos
Aquaporina 4/metabolismo , Líquido Cefalorraquidiano/metabolismo , Ritmo Circadiano/fisiologia , Sistema Glinfático/metabolismo , Animais , Astrócitos/metabolismo , Encéfalo/metabolismo , Cisterna Magna/metabolismo , Linfonodos/metabolismo , Camundongos
10.
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
11.
Life Sci ; 258: 118217, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32768575

RESUMO

AIMS: Astrocytes expressing the aquaporin-4 (AQP4) water channel are pathogenic, disease specific immunoglobulins (IgG) found in neuromyelitis optica spectrum disorder (NMOSD), referred to as NMO-IgG, which targets astrocytic AQP4. The interleukin-6 (IL-6) signaling when astrocytes were exposed to NMO-IgG present in the serum of NMOSD patients was evaluated. MAIN METHODS: Serum or human-IgG from NMOSD or healthy controls were exposed to astrocytes. The selectivity and immuno-pathological consequences of Ig binding to surface epitopes were measured by confocal microscopy. Astrocytes were exposed to medium, IL-6, soluble IL-6 receptor (sIL-6R), IL-6 + sIL-6R (IL-6/R), NMO-IgG or control-IgG, NMO-IgG + IL-6/R. The expression of key proteins in IL-6 signaling pathway, IL-6 cytokine and mRNA levels were evaluated by western blotting, enzyme-linked immunosorbent assay and quantitative polymerase chain reaction, respectively. KEY FINDINGS: Serum or NMO-IgG from NMOSD patients both induced the rapid downregulation of AQP4 expression on the surface of astrocytes. Stimulation of astrocytes with NMO-IgG, IL-6/R, and NMO-IgG + IL-6/R resulted in the enhancement of IL-6 mRNA expression. Meanwhile, the exogenous addition of NMO-IgG elicited an inflammatory transcriptional response that involved signaling through the Janus kinase/signal transducer and activator of transcription 3 (JAK/STAT3) pathway. Inhibition of the IL-6/JAK/STAT3 pathway with the JAK1/2 specific inhibitor, AZD1480, reversed the associated increase of IL-6. SIGNIFICANCE: Our findings suggest that NMO-IgG can stimulate the astrocytic JAK1/2/STAT3-dependent inflammatory response, which represents one of the important events in NMO pathogenesis. Inhibition of the JAK1/2 signaling pathway may be a novel promising therapy for NMOSD.


Assuntos
Astrócitos/metabolismo , Imunoglobulina G/sangue , Interleucina-6/metabolismo , Janus Quinases/metabolismo , Neuromielite Óptica/sangue , Fator de Transcrição STAT3/metabolismo , Adulto , Idoso , Animais , Astrócitos/efeitos dos fármacos , Autoanticorpos/sangue , Autoanticorpos/farmacologia , Células Cultivadas , Feminino , Humanos , Imunoglobulina G/farmacologia , Interleucina-6/agonistas , Masculino , Pessoa de Meia-Idade , Ratos , Ratos Wistar , Fator de Transcrição STAT3/agonistas , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Adulto Jovem
12.
PLoS Biol ; 18(8): e3000826, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32776935

RESUMO

Ca2+/calmodulin-dependent kinase II (CaMKII) regulates synaptic plasticity in multiple ways, supposedly including the secretion of neuromodulators like brain-derived neurotrophic factor (BDNF). Here, we show that neuromodulator secretion is indeed reduced in mouse α- and ßCaMKII-deficient (αßCaMKII double-knockout [DKO]) hippocampal neurons. However, this was not due to reduced secretion efficiency or neuromodulator vesicle transport but to 40% reduced neuromodulator levels at synapses and 50% reduced delivery of new neuromodulator vesicles to axons. αßCaMKII depletion drastically reduced neuromodulator expression. Blocking BDNF secretion or BDNF scavenging in wild-type neurons produced a similar reduction. Reduced neuromodulator expression in αßCaMKII DKO neurons was restored by active ßCaMKII but not inactive ßCaMKII or αCaMKII, and by CaMKII downstream effectors that promote cAMP-response element binding protein (CREB) phosphorylation. These data indicate that CaMKII regulates neuromodulation in a feedback loop coupling neuromodulator secretion to ßCaMKII- and CREB-dependent neuromodulator expression and axonal targeting, but CaMKIIs are dispensable for the secretion process itself.


Assuntos
Astrócitos/metabolismo , Fator Neurotrófico Derivado do Encéfalo/genética , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/genética , Cálcio/metabolismo , Neurônios/metabolismo , Subunidades Proteicas/genética , Animais , Astrócitos/citologia , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/deficiência , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Retroalimentação Fisiológica , Regulação da Expressão Gênica , Hipocampo/citologia , Hipocampo/metabolismo , Camundongos , Camundongos Transgênicos , Neurônios/citologia , Fosforilação , Cultura Primária de Células , Subunidades Proteicas/deficiência , Sinapses/fisiologia , Transmissão Sináptica , Imagem com Lapso de Tempo
13.
Am J Physiol Cell Physiol ; 319(3): C605-C610, 2020 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-32783655

RESUMO

Epileptic seizures are the manifestation of hypersynchronous and excessive neuronal excitation. While the glutamatergic and GABAergic neurons play major roles in shaping fast neuronal excitation/inhibition homeostasis, it is well illustrated that astrocytes profoundly regulate neuronal excitation by controlling glutamate, GABA, cannabinoids, adenosine, and concentration of K+ around neurons. However, little is known about whether microglia take part in the regulation of acute neuronal excitation and ongoing epileptic behaviors. We proposed that if microglia are innately ready to respond to epileptic overexcitation, depletion of microglia might alter neuronal excitability and severity of acute epileptic seizures. We found that microglia depletion by plx3397, an inhibitor of CSF1R, exacerbates seizure severity and excitotoxicity-induced neuronal degeneration, indicating that microglia are rapidly responsive to the change of excitation/inhibition homeostasis and participate in the protection of neurons from overexcitation.


Assuntos
Astrócitos/metabolismo , Hipocampo/fisiologia , Microglia/fisiologia , Convulsões/fisiopatologia , Animais , Ácido Glutâmico/metabolismo , Camundongos , Neurônios/fisiologia , Convulsões/metabolismo
14.
Nature ; 583(7817): 603-608, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32641832

RESUMO

Astrocytes take up glucose from the bloodstream to provide energy to the brain, thereby allowing neuronal activity and behavioural responses1-5. By contrast, astrocytes are under neuronal control through specific neurotransmitter receptors5-7. However, whether the activation of astroglial receptors can directly regulate cellular glucose metabolism to eventually modulate behavioural responses is unclear. Here we show that activation of mouse astroglial type-1 cannabinoid receptors associated with mitochondrial membranes (mtCB1) hampers the metabolism of glucose and the production of lactate in the brain, resulting in altered neuronal functions and, in turn, impaired behavioural responses in social interaction assays. Specifically, activation of astroglial mtCB1 receptors reduces the phosphorylation of the mitochondrial complex I subunit NDUFS4, which decreases the stability and activity of complex I. This leads to a reduction in the generation of reactive oxygen species by astrocytes and affects the glycolytic production of lactate through the hypoxia-inducible factor 1 pathway, eventually resulting in neuronal redox stress and impairment of behavioural responses in social interaction assays. Genetic and pharmacological correction of each of these effects abolishes the effect of cannabinoid treatment on the observed behaviour. These findings suggest that mtCB1 receptor signalling can directly regulate astroglial glucose metabolism to fine-tune neuronal activity and behaviour in mice.


Assuntos
Astrócitos/metabolismo , Metabolismo Energético , Glucose/metabolismo , Mitocôndrias/metabolismo , Receptor CB1 de Canabinoide/metabolismo , Animais , Astrócitos/citologia , Astrócitos/efeitos dos fármacos , Agonistas de Receptores de Canabinoides/farmacologia , Células Cultivadas , Dronabinol/farmacologia , Complexo I de Transporte de Elétrons/química , Complexo I de Transporte de Elétrons/metabolismo , Metabolismo Energético/efeitos dos fármacos , Glicólise/efeitos dos fármacos , Humanos , Fator 1 Induzível por Hipóxia/metabolismo , Ácido Láctico/metabolismo , Masculino , Camundongos , Mitocôndrias/efeitos dos fármacos , Membranas Mitocondriais/metabolismo , Oxirredução , Fosforilação , Espécies Reativas de Oxigênio/metabolismo , Receptor CB1 de Canabinoide/agonistas , Comportamento Social
15.
PLoS One ; 15(7): e0235852, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32628717

RESUMO

The medulloblastoma (MB) microenvironment is diverse, and cell-cell interactions within this milieu is of prime importance. Astrocytes, a major component of the microenvironment, have been shown to impact primary tumor cell phenotypes and metastasis. Based on proximity of MB cells and astrocytes in the brain microenvironment, we investigated whether astrocytes may influence MB cell phenotypes directly. Astrocyte conditioned media (ACM) increased Daoy MB cell invasion, adhesion, and in vivo cellular protrusion formation. ACM conditioning of MB cells also increased CD133 surface expression, a key cancer stem cell marker of MB. Additional neural stem cell markers, Nestin and Oct-4A, were also increased by ACM conditioning, as well as neurosphere formation. By knocking down CD133 using short interfering RNA (siRNA), we showed that ACM upregulated CD133 expression in MB plays an important role in invasion, adhesion and neurosphere formation. Collectively, our data suggests that astrocytes influence MB cell phenotypes by regulating CD133 expression, a key protein with defined roles in MB tumorgenicity and survival.


Assuntos
Antígeno AC133/genética , Astrócitos/metabolismo , Meduloblastoma/metabolismo , Fenótipo , Antígeno AC133/metabolismo , Animais , Adesão Celular , Linhagem Celular Tumoral , Movimento Celular , Células Cultivadas , Meios de Cultivo Condicionados , Humanos , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/fisiologia , Nestina/genética , Nestina/metabolismo , Fator 3 de Transcrição de Octâmero/genética , Fator 3 de Transcrição de Octâmero/metabolismo , Microambiente Tumoral , Peixe-Zebra
16.
J Pharmacol Sci ; 144(2): 83-88, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32709559

RESUMO

Astrocytes generate robust intracellular Ca2+ signals that are assumed to be key regulators of astrocytic function. Among various Ca2+ mobilization mechanisms, Ca2+ release from the endoplasmic reticulum (ER) via the inositol 1,4,5-trisphosphate receptor (IP3R) has attracted attention as a major component of astrocytic Ca2+ signaling. Manipulation of astrocytic IP3-Ca2+ signaling, such as genetic deletion of the type 2 IP3R, has revealed multifaceted roles of astrocytic ER Ca2+ release in health and disease. Recent developments in Ca2+ imaging techniques including ER intraluminal Ca2+ imaging have been indispensable in determining the physiological and pathophysiological significance of astrocytic ER Ca2+ release via IP3Rs. Beneficial and detrimental roles of IP3R-dependent Ca2+ release in astrocytes have been revealed in wide variety of disorders in the brain, strongly suggesting astrocytic IP3-Ca2+ signaling as a novel and promising therapeutic target.


Assuntos
Astrócitos/metabolismo , Sinalização do Cálcio/fisiologia , Cálcio/metabolismo , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/fisiologia , Animais , Encefalopatias/etiologia , Encefalopatias/genética , Encefalopatias/terapia , Deleção de Genes , Humanos , Inositol 1,4,5-Trifosfato/metabolismo , Receptores de Inositol 1,4,5-Trifosfato/genética , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Imagem Molecular , Terapia de Alvo Molecular
17.
Life Sci ; 258: 118140, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32730838

RESUMO

AIMS: This study was performed to investigate the expression profile of cytochrome P450 (CYP) isoforms and effects of polycyclic aromatic hydrocarbons (PAHs) and antiepileptic drugs on CYP1 expression in human astrocytoma MOG-G-CCM cells. MAIN METHODS: CYP1A1 and CYP1B1 expression were determined by quantitative real-time polymerase chain reaction, Western blotting, and immunocytochemistry. KEY FINDINGS: MOG-G-CCM cells expressed various CYP isoforms. Among the CYP isoforms analyzed, CYP1B1 showed the highest expression level, followed by CYP1A1. Furthermore, CYP1B1 was localized in both the endoplasmic reticulum and mitochondria. 3-Methylcholanthrene (3-MC), benz[a]anthracene (B[a]A), benzo[a]pyrene (B[a]P), and valproic acid (VPA) increased the expression of CYP1B1 and CYP1A1. The potent aryl hydrocarbon receptor antagonist GNF351 significantly suppressed the 3-MC- and VPA-mediated upregulation of CYP1B1 and CYP1A1. In addition, VPA potentiated the induction of CYP1B1 and CYP1A1 by 3-MC, B[a]A, and B[a]P, although the augmentation of CYP1A1 was more remarkable than that of CYP1B1. In contrast, other antiepileptic drugs (carbamazepine, lamotrigine, levetiracetam, phenytoin) did not affect the 3-MC-mediated upregulation of CYP1B1 and CYP1A1. VPA is known to act as a histone deacetylase (HDAC) inhibitor. Therefore, the effects of trichostatin A, a representative HDAC inhibitor, on CYP1 induction by 3-MC were examined. Trichostatin A enhanced the 3-MC-mediated upregulation of CYP1A1 but not CYP1B1. SIGNIFICANCE: These results partially indicated that VPA may augment the PAH-mediated induction of CYP1B1 and CYP1A1 through the activation of transcription by HDAC inhibition.


Assuntos
Anticonvulsivantes/farmacologia , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1B1/genética , Hidrocarbonetos Policíclicos Aromáticos/farmacologia , Regulação para Cima/efeitos dos fármacos , Ácido Valproico/farmacologia , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Linhagem Celular Tumoral , Humanos , Transcriptoma/efeitos dos fármacos
18.
PLoS One ; 15(7): e0234614, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32649728

RESUMO

Exosomes appear to be effective inter-cellular communicators delivering several types of molecules, such as proteins and RNAs, suggesting that they could influence neural stem cell (NSC) differentiation. Our RNA sequencing studies demonstrated that the RNAs related to cell proliferation and astrocyte differentiation were upregulated in human mesenchymal stem cells (hMSC) when co-cultured with exosomes obtained from the culture medium of human glioma cells (U87). Metallothionein 3 and elastin genes, which are related to cell proliferation, increased 10 and 7.2 fold, respectively. Expression of genes for astrocyte differentiation, such as tumor growth factor alpha, induced protein 3 of the NOTCH1 family, colony stimulating factor and interleukin 6 of the STAT3 family and Hes family bHLH transcription factor 1 also increased by 2.3, 10, 4.7 and 2.9 fold, respectively. We further examined the effects of these exosomes on rat fetal neural stem cell (rNSC) differentiation using the secreted exosomes from U87 glioma cells or exosomes from U87 cells that were stimulated with interleukin 1ß (IL-1ß). The rNSCs, extracted from rat brains at embryonic day 14 (E14), underwent a culture protocol that normally leads to predominant (~90%) differentiation to ODCs. However, in the presence of the exosomes from untreated or IL-1ß-treated U87 cells, significantly more cells differentiated into astrocytes, especially in the presence of exosomes obtained from the IL-1ß-challenged glioma cells. Moreover, glioma-derived exosomes appeared to inhibit rNSC differentiation into ODCs or astrocytes as indicated by a significantly increased population of unlabeled cells. A portion of the resulting astrocytes co-expressed both CD133 and glial fibrillary acidic protein (GFAP) suggesting that exosomes from U87 cells could promote astrocytic differentiation of NSCs with features expected from a transformed cell. Our data clearly demonstrated that exosomes secreted by human glioma cells provide a strong driving force for rat neural stem cells to differentiate into astrocytes, uncovering potential pathways and therapeutic targets that might control this aggressive tumor type.


Assuntos
Astrócitos/metabolismo , Diferenciação Celular/fisiologia , Exossomos/fisiologia , Células-Tronco Neurais/metabolismo , Animais , Astrócitos/fisiologia , Proliferação de Células , Células Cultivadas , Técnicas de Cocultura , Elastina/metabolismo , Exossomos/metabolismo , Regulação da Expressão Gênica/genética , Glioma/metabolismo , Humanos , Interleucina-6/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Células-Tronco Neurais/fisiologia , Neurônios/metabolismo , Cultura Primária de Células , Ratos , Fator de Transcrição STAT3/metabolismo
19.
Proc Natl Acad Sci U S A ; 117(31): 18810-18821, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32690710

RESUMO

In the central nervous system, glycogen-derived bioenergetic resources in astrocytes help promote tissue survival in response to focal neuronal stress. However, our understanding of the extent to which these resources are mobilized and utilized during neurodegeneration, especially in nearby regions that are not actively degenerating, remains incomplete. Here we modeled neurodegeneration in glaucoma, the world's leading cause of irreversible blindness, and measured how metabolites mobilize through astrocyte gap junctions composed of connexin 43 (Cx43). We elevated intraocular pressure in one eye and determined how astrocyte-derived metabolites in the contralateral optic projection responded. Remarkably, astrocyte networks expand and redistribute metabolites along distances even 10 mm in length, donating resources from the unstressed to the stressed projection in response to intraocular pressure elevation. While resource donation improves axon function and visual acuity in the directly stressed region, it renders the donating tissue susceptible to bioenergetic, structural, and physiological degradation. Intriguingly, when both projections are stressed in a WT animal, axon function and visual acuity equilibrate between the two projections even when each projection is stressed for a different length of time. This equilibration does not occur when Cx43 is not present. Thus, Cx43-mediated astrocyte metabolic networks serve as an endogenous mechanism used to mitigate bioenergetic stress and distribute the impact of neurodegenerative disease processes. Redistribution ultimately renders the donating optic nerve vulnerable to further metabolic stress, which could explain why local neurodegeneration does not remain confined, but eventually impacts healthy regions of the brain more broadly.


Assuntos
Astrócitos , Glaucoma/metabolismo , Doenças Neurodegenerativas/metabolismo , Animais , Astrócitos/metabolismo , Astrócitos/fisiologia , Conexina 43/genética , Conexina 43/metabolismo , Feminino , Junções Comunicantes/metabolismo , Pressão Intraocular/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos
20.
Nat Commun ; 11(1): 3240, 2020 07 06.
Artigo em Inglês | MEDLINE | ID: mdl-32632168

RESUMO

Astrocytic Ca2+ signaling has been intensively studied in health and disease but has not been quantified during natural sleep. Here, we employ an activity-based algorithm to assess astrocytic Ca2+ signals in the neocortex of awake and naturally sleeping mice while monitoring neuronal Ca2+ activity, brain rhythms and behavior. We show that astrocytic Ca2+ signals exhibit distinct features across the sleep-wake cycle and are reduced during sleep compared to wakefulness. Moreover, an increase in astrocytic Ca2+ signaling precedes transitions from slow wave sleep to wakefulness, with a peak upon awakening exceeding the levels during whisking and locomotion. Finally, genetic ablation of an important astrocytic Ca2+ signaling pathway impairs slow wave sleep and results in an increased number of microarousals, abnormal brain rhythms, and an increased frequency of slow wave sleep state transitions and sleep spindles. Our findings demonstrate an essential role for astrocytic Ca2+ signaling in regulating slow wave sleep.


Assuntos
Astrócitos/metabolismo , Sinalização do Cálcio , Sono de Ondas Lentas/fisiologia , Animais , Camundongos , Vigília/fisiologia
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