RESUMO
Human brain ontogeny is characterized by a considerably prolonged neotenic development of cortical neurons and circuits. Neoteny is thought to be essential for the acquisition of advanced cognitive functions, which are typically altered in intellectual disability (ID) and autism spectrum disorders (ASDs). Human neuronal neoteny could be disrupted in some forms of ID and/or ASDs, but this has never been tested. Here, we use xenotransplantation of human cortical neurons into the mouse brain to model SYNGAP1 haploinsufficiency, one of the most prevalent genetic causes of ID/ASDs. We find that SYNGAP1-deficient human neurons display strong acceleration of morphological and functional synaptic formation and maturation alongside disrupted synaptic plasticity. At the circuit level, SYNGAP1-haploinsufficient neurons display precocious acquisition of responsiveness to visual stimulation months ahead of time. Our findings indicate that SYNGAP1 is required cell autonomously for human neuronal neoteny, providing novel links between human-specific developmental mechanisms and ID/ASDs.
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The maintenance of a highly functional metabolic epithelium in vitro is challenging. Metabolic impairments in primary human hepatocytes (PHHs) over time is primarily due to epithelial-to-mesenchymal transitioning (EMT). The immature hepatoma cell line HepG2 was used as an in vitro model to explore strategies for enhancing the hepatic phenotype. The phenotypic characterization includes measuring the urea cycle, lipid storage, tricarboxylic acid-related metabolites, reactive oxygen species, endoplasmic reticulum calcium efflux, mitochondrial membrane potentials, oxygen consumptions rate, and CYP450 biotransformation capacity. Expression studies were performed with transcriptomics, co-immunoprecipitation and proteomics. CRISPR/Cas9 was also employed to genetically engineer HepG2 cells. After confirming that PHHs develop an EMT phenotype, expression of tankyrase1/2 was found to increase over time. EMT was reverted when blocking tankyrases1/2-dependent poly-ADP-ribosylation (PARylation) activity, by biochemical and genetic perturbation. Wnt/ß-catenin inhibitor XAV-939 blocks tankyrase1/2 and treatment elevated several oxygen-consuming reactions (electron-transport chain, OXHPOS, CYP450 mono-oxidase activity, phase I/II xenobiotic biotransformation, and prandial turnover), suggesting that cell metabolism was enhanced. Glutathione-dependent redox homeostasis was also significantly improved in the XAV-939 condition. Oxygen consumption rate and proteomics experiments in tankyrase1/2 double knockout HepG2 cells then uncovered PARylation as master regulator of aerobic-dependent cell respiration. Furthermore, novel tankyrase1/2-dependent PARylation targets, including mitochondrial DLST, and OGDH, were revealed. This work exposed a new mechanistic framework by linking PARylation to respiration and metabolism, thereby broadening the current understanding that underlies these vital processes. XAV-939 poses an immediate and straightforward strategy to improve aerobic activities, and metabolism, in (immature) cell cultures.
Assuntos
Transição Epitelial-Mesenquimal , Hepatócitos , Tanquirases , Humanos , Tanquirases/antagonistas & inibidores , Tanquirases/metabolismo , Células Hep G2 , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/fisiologia , Poli ADP Ribosilação/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Fenantrenos/farmacologiaRESUMO
Neuronal endosomal and lysosomal abnormalities are among the early changes observed in Alzheimer's disease (AD) before plaques appear. However, it is unclear whether distinct endolysosomal defects are temporally organized and how altered γ-secretase function or amyloid precursor protein (APP) metabolism contribute to these changes. Inhibiting γ-secretase chronically, in mouse embryonic fibroblast and hippocampal neurons, led to a gradual endolysosomal collapse initiated by decreased lysosomal calcium and increased cholesterol, causing downstream defects in endosomal recycling and maturation. This endolysosomal demise is γ-secretase dependent, requires membrane-tethered APP cytoplasmic domains, and is rescued by APP depletion. APP C-terminal fragments (CTFs) localized to late endosome/lysosome-endoplasmic reticulum contacts; an excess of APP-CTFs herein reduced lysosomal Ca2+ refilling from the endoplasmic reticulum, promoting cholesterol accretion. Tonic regulation by APP-CTFs provides a mechanistic explanation for their cellular toxicity: failure to timely degrade APP-CTFs sustains downstream signaling, instigating lysosomal dyshomeostasis, as observed in prodromal AD. This is the opposite of substrates such as Notch, which require intramembrane proteolysis to initiate signaling.
Assuntos
Doença de Alzheimer , Secretases da Proteína Precursora do Amiloide , Precursor de Proteína beta-Amiloide , Retículo Endoplasmático , Endossomos , Lisossomos , Neurônios , Lisossomos/metabolismo , Animais , Endossomos/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Camundongos , Retículo Endoplasmático/metabolismo , Secretases da Proteína Precursora do Amiloide/metabolismo , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Neurônios/metabolismo , Colesterol/metabolismo , Hipocampo/metabolismo , Hipocampo/patologia , Cálcio/metabolismo , Humanos , Fibroblastos/metabolismo , Transdução de Sinais , ProteóliseRESUMO
Although human pluripotent stem cell (PSC)-derived brain organoids have enabled researchers to gain insight into human brain development and disease, these organoids contain solely ectodermal cells and are not vascularized as occurs during brain development. Here it is created less complex and more homogenous large neural constructs starting from PSC-derived neuroprogenitor cells (NPC), by fusing small NPC spheroids into so-called concentroids. Such concentroids consisted of a pro-angiogenic core, containing neuronal and outer radial glia cells, surrounded by an astroglia-dense outer layer. Incorporating PSC-derived endothelial cells (EC) around and/or in the concentroids promoted vascularization, accompanied by differential outgrowth and differentiation of neuronal and astroglia cells, as well as the development of ectodermal-derived pericyte-like mural cells co-localizing with EC networks. Single nucleus transcriptomic analysis revealed an enhanced neural cell subtype maturation and diversity in EC-containing concentroids, which better resemble the fetal human brain compared to classical organoids or NPC-only concentroids. This PSC-derived "vascularized" concentroid brain model will facilitate the study of neurovascular/blood-brain barrier development, neural cell migration, and the development of effective in vitro vascularization strategies of brain mimics.
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Células Endoteliais , Células-Tronco Pluripotentes , Humanos , Células Endoteliais/fisiologia , Neurogênese/fisiologia , Diferenciação Celular/fisiologia , EncéfaloRESUMO
Alzheimer's disease (AD) is a devastating neurodegenerative disorder characterized by a concerning rise in prevalence. It is projected that the number of affected individuals will reach a staggering 150 million by 2050. While recent advancements in monoclonal antibodies targeting Aß have shown some clinical effects, there is an urgent need for improved therapies to effectively address the impeding surge of AD patients worldwide. To achieve this, a deeper understanding of the intricate mechanisms underlying the disease is crucial. In recent years, mounting evidence has underscored the vital role of the innate immune system in AD pathology. However, limited findings persist regarding the involvement of the adaptive immune system. Here, we report on the impact of the adaptive immune system on various aspects of AD by using AppNL-G-F mice crossed into a Rag2-/- background lacking mature adaptive immune cells. In addition, to simulate the continuous exposure to various challenges such as infections that is commonly observed in humans, the innate immune system was activated through the repetitive induction of peripheral inflammation. We observed a remarkably improved performance on complex cognitive tasks when a mature adaptive immune system is absent. Notably, this observation is pathologically associated with lower Aß plaque accumulation, reduced glial activation, and better-preserved neuronal networks in the mice lacking a mature adaptive immune system. Collectively, these findings highlight the detrimental role of the adaptive immune system in AD and underscore the need for effective strategies to modulate it for therapeutic purposes.
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Doença de Alzheimer , Humanos , Animais , Camundongos , Anticorpos Monoclonais , Sistema Imunitário , Inflamação , Placa AmiloideRESUMO
The enhanced cognitive abilities characterizing the human species result from specialized features of neurons and circuits. Here, we report that the hominid-specific gene LRRC37B encodes a receptor expressed in human cortical pyramidal neurons (CPNs) and selectively localized to the axon initial segment (AIS), the subcellular compartment triggering action potentials. Ectopic expression of LRRC37B in mouse CPNs in vivo leads to reduced intrinsic excitability, a distinctive feature of some classes of human CPNs. Molecularly, LRRC37B binds to the secreted ligand FGF13A and to the voltage-gated sodium channel (Nav) ß-subunit SCN1B. LRRC37B concentrates inhibitory effects of FGF13A on Nav channel function, thereby reducing excitability, specifically at the AIS level. Electrophysiological recordings in adult human cortical slices reveal lower neuronal excitability in human CPNs expressing LRRC37B. LRRC37B thus acts as a species-specific modifier of human neuron excitability, linking human genome and cell evolution, with important implications for human brain function and diseases.
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Neurônios , Células Piramidais , Canais de Sódio Disparados por Voltagem , Animais , Humanos , Camundongos , Potenciais de Ação/fisiologia , Axônios/metabolismo , Neurônios/metabolismo , Canais de Sódio Disparados por Voltagem/genética , Canais de Sódio Disparados por Voltagem/metabolismoRESUMO
Human cerebral organoids (COs) are self-organizing three-dimensional (3D) neural structures that provide a human-specific platform to study the cellular and molecular processes that underlie different neurological events. The first step of CO generation from human pluripotent stem cells (hPSCs) is neural induction, which is an in vitro simulation of neural ectoderm development. Several signaling pathways cooperate during neural ectoderm development and in vitro differentiation of hPSCs toward neural cell lineages is also affected by them. In this study, we considered some of the known sources of these variable signaling cues arising from cell culture media components and sought to modulate their effects by applying a comprehensive combination of small molecules and growth factors for CO generation. Histological analysis demonstrated that these COs recapitulate the neural progenitor zone and early cortical layer organization, containing different types of neuronal and glial cells which was in accordance with single-nucleus transcriptome profiling results. Moreover, patch clamp and intracellular Ca2+ dynamic studies demonstrated that the COs behave as a functional neural network. Thus, this method serves as a facile protocol for generating hPSC-derived COs that faithfully mimic the features of their in vivo counterparts in the developing human brain. See also Figure 1(Fig. 1).
RESUMO
Early Alzheimer's disease (AD) is associated with hippocampal hyperactivity and decreased sleep quality. Here we show that homeostatic mechanisms transiently counteract the increased excitatory drive to CA1 neurons in AppNL-G-F mice, but that this mechanism fails in older mice. Spatial transcriptomics analysis identifies Pmch as part of the adaptive response in AppNL-G-F mice. Pmch encodes melanin-concentrating hormone (MCH), which is produced in sleep-active lateral hypothalamic neurons that project to CA1 and modulate memory. We show that MCH downregulates synaptic transmission, modulates firing rate homeostasis in hippocampal neurons and reverses the increased excitatory drive to CA1 neurons in AppNL-G-F mice. AppNL-G-F mice spend less time in rapid eye movement (REM) sleep. AppNL-G-F mice and individuals with AD show progressive changes in morphology of CA1-projecting MCH axons. Our findings identify the MCH system as vulnerable in early AD and suggest that impaired MCH-system function contributes to aberrant excitatory drive and sleep defects, which can compromise hippocampus-dependent functions.
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Doença de Alzheimer , Hormônios Hipotalâmicos , Camundongos , Animais , Doença de Alzheimer/genética , Neurônios/fisiologia , Hormônios Hipofisários , Sono , Camundongos TransgênicosRESUMO
The gut microbiota represents a diverse and dynamic population of microorganisms that can influence the health of the host. Increasing evidence supports the role of the gut microbiota as a key player in the pathogenesis of neurodegenerative diseases, including Alzheimer's disease (AD). Unfortunately, the mechanisms behind the interplay between gut pathogens and AD are still elusive. It is known that bacteria-derived outer membrane vesicles (OMVs) act as natural carriers of virulence factors that are central players in the pathogenesis of the bacteria. Helicobacter pylori (H. pylori) is a common gastric pathogen and H. pylori infection has been associated with an increased risk to develop AD. Here, we are the first to shed light on the role of OMVs derived from H. pylori on the brain in healthy conditions and on disease pathology in the case of AD. Our results reveal that H. pylori OMVs can cross the biological barriers, eventually reaching the brain. Once in the brain, these OMVs are taken up by astrocytes, which induce activation of glial cells and neuronal dysfunction, ultimately leading to exacerbated amyloid-ß pathology and cognitive decline. Mechanistically, we identified a critical role for the complement component 3 (C3)-C3a receptor (C3aR) signalling in mediating the interaction between astrocytes, microglia and neurons upon the presence of gut H. pylori OMVs. Taken together, our study reveals that H. pylori has a detrimental effect on brain functionality and accelerates AD development via OMVs and C3-C3aR signalling.
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Doença de Alzheimer , Vesículas Extracelulares , Helicobacter pylori , Humanos , Doença de Alzheimer/etiologia , Doença de Alzheimer/patologia , Encéfalo , Vesículas Extracelulares/patologiaRESUMO
Neuronal development in the human cerebral cortex is considerably prolonged compared with that of other mammals. We explored whether mitochondria influence the species-specific timing of cortical neuron maturation. By comparing human and mouse cortical neuronal maturation at high temporal and cell resolution, we found a slower mitochondria development in human cortical neurons compared with that in the mouse, together with lower mitochondria metabolic activity, particularly that of oxidative phosphorylation. Stimulation of mitochondria metabolism in human neurons resulted in accelerated development in vitro and in vivo, leading to maturation of cells weeks ahead of time, whereas its inhibition in mouse neurons led to decreased rates of maturation. Mitochondria are thus important regulators of the pace of neuronal development underlying human-specific brain neoteny.
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Mitocôndrias , Neurogênese , Neurônios , Animais , Humanos , Camundongos , Córtex Cerebral/citologia , Córtex Cerebral/crescimento & desenvolvimento , Metabolismo Energético , Mitocôndrias/metabolismo , Neurônios/metabolismoRESUMO
KIF2A is an atypical kinesin that has the capacity to depolymerize microtubules. Patients carrying mutations in KIF2A suffer from progressive microcephaly and mental disabilities. While the role of this protein is well documented in neuronal migration, the relationship between its dysfunction and the pathobiology of brain disorders is unclear. Here, we report that KIF2A is dispensable for embryogenic neurogenesis but critical in postnatal stages for maturation, connectivity, and maintenance of neurons. We used a conditional approach to inactivate KIF2A in cortical progenitors, nascent postmitotic neurons, and mature neurons in mice. We show that the lack of KIF2A alters microtubule dynamics and disrupts several microtubule-dependent processes, including neuronal polarity, neuritogenesis, synaptogenesis, and axonal transport. KIF2A-deficient neurons exhibit aberrant electrophysiological characteristics, neuronal connectivity, and function, leading to their loss. The role of KIF2A is not limited to development, as fully mature neurons require KIF2A for survival. Our results emphasize an additional function of KIF2A and help explain how its mutations lead to brain disorders.
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Encefalopatias , Proteínas Repressoras , Animais , Camundongos , Proteínas Repressoras/metabolismo , Cinesinas/genética , Microtúbulos/metabolismo , Neurônios/metabolismo , Encefalopatias/metabolismoRESUMO
Loss-of-function mutations in the PRKN, PINK1 and PARK7 genes (encoding parkin, PINK1 and DJ-1, respectively) cause autosomal recessive forms of Parkinson's disease. PINK1 and parkin jointly mediate selective autophagy of damaged mitochondria (mitophagy), but the mechanisms by which loss of DJ-1 induces Parkinson's disease are not well understood. Here, we investigated PINK1/parkin-mediated mitophagy in cultured human fibroblasts and induced pluripotent stem cell-derived neurons with homozygous PARK7 mutations. We found that DJ-1 is essential for PINK1/parkin-mediated mitophagy. Loss of DJ-1 did not interfere with PINK1 or parkin activation after mitochondrial depolarization but blocked mitophagy further downstream by inhibiting recruitment of the selective autophagy receptor optineurin to depolarized mitochondria. By contrast, starvation-induced, non-selective autophagy was not affected by loss of DJ-1. In wild-type fibroblasts and induced pluripotent stem cell-derived dopaminergic neurons, endogenous DJ-1 translocated to depolarized mitochondria in close proximity to optineurin. DJ-1 translocation to depolarized mitochondria was dependent on PINK1 and parkin and did not require oxidation of cysteine residue 106 of DJ-1. Overexpression of DJ-1 did not rescue the mitophagy defect of PINK1- or parkin-deficient cells. These findings position DJ-1 downstream of PINK1 and parkin in the same pathway and suggest that disruption of PINK1/parkin/DJ-1-mediated mitophagy is a common pathogenic mechanism in autosomal recessive Parkinson's disease.
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Mitofagia , Doença de Parkinson , Proteínas Quinases , Humanos , Mitocôndrias/metabolismo , Mitofagia/genética , Mitofagia/fisiologia , Doença de Parkinson/metabolismo , Proteínas Quinases/genética , Ubiquitina-Proteína Ligases/genéticaRESUMO
The ability of immune-modulating biologics to prevent and reverse pathology has transformed recent clinical practice. Full utility in the neuroinflammation space, however, requires identification of both effective targets for local immune modulation and a delivery system capable of crossing the blood-brain barrier. The recent identification and characterization of a small population of regulatory T (Treg) cells resident in the brain presents one such potential therapeutic target. Here, we identified brain interleukin 2 (IL-2) levels as a limiting factor for brain-resident Treg cells. We developed a gene-delivery approach for astrocytes, with a small-molecule on-switch to allow temporal control, and enhanced production in reactive astrocytes to spatially direct delivery to inflammatory sites. Mice with brain-specific IL-2 delivery were protected in traumatic brain injury, stroke and multiple sclerosis models, without impacting the peripheral immune system. These results validate brain-specific IL-2 gene delivery as effective protection against neuroinflammation, and provide a versatile platform for delivery of diverse biologics to neuroinflammatory patients.
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Astrócitos , Produtos Biológicos , Animais , Encéfalo , Humanos , Interleucina-2/genética , Interleucinas , Camundongos , Doenças Neuroinflamatórias , Linfócitos T ReguladoresRESUMO
Single domain antibodies (VHHs) are potentially disruptive therapeutics, with important biological value for treatment of several diseases, including neurological disorders. However, VHHs have not been widely used in the central nervous system (CNS), largely because of their restricted blood-brain barrier (BBB) penetration. Here, we propose a gene transfer strategy based on BBB-crossing adeno-associated virus (AAV)-based vectors to deliver VHH directly into the CNS. As a proof-of-concept, we explored the potential of AAV-delivered VHH to inhibit BACE1, a well-characterized target in Alzheimer's disease. First, we generated a panel of VHHs targeting BACE1, one of which, VHH-B9, shows high selectivity for BACE1 and efficacy in lowering BACE1 activity in vitro. We further demonstrate that a single systemic dose of AAV-VHH-B9 produces positive long-term (12 months plus) effects on amyloid load, neuroinflammation, synaptic function, and cognitive performance, in the AppNL-G-F Alzheimer's mouse model. These results constitute a novel therapeutic approach for neurodegenerative diseases, which is applicable to a range of CNS disease targets.
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Doença de Alzheimer , Secretases da Proteína Precursora do Amiloide , Ácido Aspártico Endopeptidases , Anticorpos de Domínio Único , Doença de Alzheimer/patologia , Secretases da Proteína Precursora do Amiloide/imunologia , Secretases da Proteína Precursora do Amiloide/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Ácido Aspártico Endopeptidases/imunologia , Ácido Aspártico Endopeptidases/metabolismo , Barreira Hematoencefálica , Dependovirus/genética , Modelos Animais de Doenças , Vetores Genéticos/uso terapêutico , Camundongos , Camundongos TransgênicosRESUMO
Biological responses to stress are complex and highly conserved. Corticotropin-releasing factor (CRF) plays a central role in regulating these lifesaving physiological responses to stress. We show that, in mice, CRF rapidly changes Schaffer Collateral (SC) input into hippocampal CA1 pyramidal cells (PC) by modulating both functional and structural aspects of these synapses. Host exposure to acute stress, in vivo CRF injection, and ex vivo CRF application all result in fast de novo formation and remodeling of existing dendritic spines. Functionally, CRF leads to a rapid increase in synaptic strength of SC input into CA1 neurons, e.g., increase in spontaneous neurotransmitter release, paired-pulse facilitation, and repetitive excitability and improves synaptic plasticity: long-term potentiation (LTP) and long-term depression (LTD). In line with the changes in synaptic function, CRF increases the number of presynaptic vesicles, induces redistribution of vesicles towards the active zone, increases active zone size, and improves the alignment of the pre- and postsynaptic compartments. Therefore, CRF rapidly enhances synaptic communication in the hippocampus, potentially playing a crucial role in the enhanced memory consolidation in acute stress.
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Hormônio Liberador da Corticotropina , Células Piramidais , Animais , Hipocampo , Potenciação de Longa Duração , Camundongos , Sinapses , Transmissão SinápticaRESUMO
Astrocytes, the main supportive cell type of the brain, show functional impairments upon ageing and in a broad spectrum of neurological disorders. Limited access to human astroglia for pre-clinical studies has been a major bottleneck delaying our understanding of their role in brain health and disease. We demonstrate here that functionally mature human astrocytes can be generated by SOX9 overexpression for 6 days in pluripotent stem cell (PSC)-derived neural progenitor cells. Inducible (i)SOX9-astrocytes display functional properties comparable to primary human astrocytes comprising glutamate uptake, induced calcium responses and cytokine/growth factor secretion. Importantly, electrophysiological properties of iNGN2-neurons co-cultured with iSOX9-astrocytes are indistinguishable from gold-standard murine primary cultures. The high yield, fast timing and the possibility to cryopreserve iSOX9-astrocytes without losing functional properties makes them suitable for scaled-up production for high-throughput analyses. Our findings represent a step forward to an all-human iPSC-derived neural model for drug development in neuroscience and towards the reduction of animal use in biomedical research.
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Astrócitos , Células-Tronco Neurais , Animais , Astrócitos/citologia , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Camundongos , Células-Tronco Neurais/fisiologia , Neurogênese/fisiologia , Neurônios/citologia , Fatores de Transcrição SOX9/metabolismoRESUMO
Tau is a major driver of neurodegeneration and is implicated in over 20 diseases. Tauopathies are characterized by synaptic loss and neuroinflammation, but it is unclear if these pathological events are causally linked. Tau binds to Synaptogyrin-3 on synaptic vesicles. Here, we interfered with this function to determine the role of pathogenic Tau at pre-synaptic terminals. We show that heterozygous knockout of synaptogyrin-3 is benign in mice but strongly rescues mutant Tau-induced defects in long-term synaptic plasticity and working memory. It also significantly rescues the pre- and post-synaptic loss caused by mutant Tau. However, Tau-induced neuroinflammation remains clearly upregulated when we remove the expression of one allele of synaptogyrin-3. Hence neuroinflammation is not sufficient to cause synaptic loss, and these processes are separately induced in response to mutant Tau. In addition, the pre-synaptic defects caused by mutant Tau are enough to drive defects in cognitive tasks.
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Transtornos da Memória/fisiopatologia , Microglia/fisiologia , Terminações Pré-Sinápticas/fisiologia , Sinaptogirinas/fisiologia , Proteínas tau/fisiologia , Animais , Encefalite/fisiopatologia , Feminino , Hipocampo/fisiopatologia , Hipocampo/ultraestrutura , Masculino , Camundongos Knockout , Plasticidade Neuronal , Terminações Pré-Sinápticas/ultraestrutura , Sinaptogirinas/genéticaRESUMO
Excitatory and inhibitory neurons are connected into microcircuits that generate circuit output. Central in the hippocampal CA3 microcircuit is the mossy fiber (MF) synapse, which provides powerful direct excitatory input and indirect feedforward inhibition to CA3 pyramidal neurons. Here, we dissect its cell-surface protein (CSP) composition to discover novel regulators of MF synaptic connectivity. Proteomic profiling of isolated MF synaptosomes uncovers a rich CSP composition, including many CSPs without synaptic function and several that are uncharacterized. Cell-surface interactome screening identifies IgSF8 as a neuronal receptor enriched in the MF pathway. Presynaptic Igsf8 deletion impairs MF synaptic architecture and robustly decreases the density of bouton filopodia that provide feedforward inhibition. Consequently, IgSF8 loss impairs excitation/inhibition balance and increases excitability of CA3 pyramidal neurons. Our results provide insight into the CSP landscape and interactome of a specific excitatory synapse and reveal IgSF8 as a critical regulator of CA3 microcircuit connectivity and function.
Assuntos
Região CA3 Hipocampal/fisiologia , Proteínas de Transporte/metabolismo , Potenciais Pós-Sinápticos Excitadores/fisiologia , Proteínas de Membrana/metabolismo , Fibras Musgosas Hipocampais/metabolismo , Células Piramidais/fisiologia , Animais , Proteínas de Transporte/genética , Células Cultivadas , Células HEK293 , Humanos , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout , Técnicas de Patch-Clamp , Cultura Primária de Células , Proteômica , Ratos , Sinaptossomos/metabolismoRESUMO
Classically, neurexins are thought to mediate synaptic connections through trans interactions with a number of different postsynaptic partners. Neurexins are cleaved by metalloproteases in an activity-dependent manner, releasing the soluble extracellular domain. Here, we report that in both immature (before synaptogenesis) and mature (after synaptogenesis) hippocampal neurons, the soluble neurexin-1ß ectodomain triggers acute Ca2+-influx at the dendritic/postsynaptic side. In both cases, neuroligin-1 expression was required. In immature neurons, calcium influx required N-type calcium channels and stimulated dendritic outgrowth and neuronal survival. In mature glutamatergic neurons the neurexin-1ß ectodomain stimulated calcium influx through NMDA-receptors, which increased presynaptic release probability. In contrast, prolonged exposure to the ectodomain led to inhibition of synaptic transmission. This secondary inhibition was activity- and neuroligin-1 dependent and caused by a reduction in the readily-releasable pool of vesicles. A synthetic peptide modeled after the neurexin-1ß:neuroligin-1 interaction site reproduced the cellular effects of the neurexin-1ß ectodomain. Collectively, our findings demonstrate that the soluble neurexin ectodomain stimulates growth of neurons and exerts acute and chronic effects on trans-synaptic signaling involved in setting synaptic strength.