Tau deposits in astrocytes are frequently found in Alzheimer's disease (AD) and other tauopathies. Since astrocytes do not express tau, the inclusions have been suggested to be of neuronal origin. However, the mechanisms behind their appearance and their relevance for disease progression remain unknown. Here we demonstrate, using a battery of experimental techniques that human astrocytes serve as an intermediator, promoting cell-to-cell spreading of pathological tau. Human astrocytes engulf and process, but fail to fully degrade dead neurons with tau pathology, as well as synthetic tau fibrils and tau aggregates isolated from AD brain tissue. Instead, the pathogenic tau is spread to nearby cells via secretion and tunneling nanotube mediated transfer. By performing co-culture experiments we could show that tau-containing astrocytes induce tau pathology in healthy human neurons directly. Furthermore, our results from a FRET based seeding assay, demonstrated that the tau proteoforms secreted by astrocytes have an exceptional seeding capacity, compared to the original tau species engulfed by the cells. Taken together, our study establishes a central role for astrocytes in mediating tau pathology, which could be of relevance for identifying novel treatment targets for AD and other tauopathies.
Alzheimer Disease , Tauopathies , Humans , tau Proteins/metabolism , Astrocytes/metabolism , Tauopathies/pathology , Alzheimer Disease/pathology , Neurons/metabolism , Brain/pathology
BACKGROUND: Astrocytes are crucial for maintaining brain homeostasis and synaptic function, but are also tightly connected to the pathogenesis of Alzheimer's disease (AD). Our previous data demonstrate that astrocytes ingest large amounts of aggregated amyloid-beta (Aß), but then store, rather than degrade the ingested material, which leads to severe cellular stress. However, the involvement of pathological astrocytes in AD-related synaptic dysfunction remains to be elucidated. METHODS: In this study, we aimed to investigate how intracellular deposits of Aß in astrocytes affect their interplay with neurons, focusing on neuronal function and viability. For this purpose, human induced pluripotent stem cell (hiPSC)-derived astrocytes were exposed to sonicated Αß42 fibrils. The direct and indirect effects of the Αß-exposed astrocytes on hiPSC-derived neurons were analyzed by performing astrocyte-neuron co-cultures as well as additions of conditioned media or extracellular vesicles to pure neuronal cultures. RESULTS: Electrophysiological recordings revealed significantly decreased frequency of excitatory post-synaptic currents in neurons co-cultured with Aß-exposed astrocytes, while conditioned media from Aß-exposed astrocytes had the opposite effect and resulted in hyperactivation of the synapses. Clearly, factors secreted from control, but not from Aß-exposed astrocytes, benefited the wellbeing of neuronal cultures. Moreover, reactive astrocytes with Aß deposits led to an elevated clearance of dead cells in the co-cultures. CONCLUSIONS: Taken together, our results demonstrate that inclusions of aggregated Aß affect the reactive state of the astrocytes, as well as their ability to support neuronal function.
Alzheimer Disease , Induced Pluripotent Stem Cells , Humans , Astrocytes/metabolism , Induced Pluripotent Stem Cells/metabolism , Culture Media, Conditioned/pharmacology , Cells, Cultured , Amyloid beta-Peptides/pharmacology , Amyloid beta-Peptides/metabolism , Neurons/metabolism , Alzheimer Disease/pathology
Studies suggest that astrocytic connexins (Cx) have an important role in the regulation of high brain functions through their ability to establish fine-tuned communication with neurons within the tripartite synapse. In light of these properties, growing evidence suggests a role of Cx in psychiatric disorders such as major depression but also in the therapeutic activity of antidepressant drugs. However, the real impact of Cx on treatment response and the underlying neurobiological mechanisms remain yet to be clarified. On this ground, the present study was designed to evaluate the functional activity of Cx in a mouse model of depression based on chronic corticosterone exposure and to determine to which extent their pharmacological inactivation influences the antidepressant-like activity of venlafaxine (VENLA). On the one hand, our results indicate that depressed mice have impaired Cx-based gap-junction and hemichannel activities. On the other hand, while VENLA exerts robust antidepressant-like activity in depressed mice; this effect is abolished by the pharmacological inhibition of Cx with carbenoxolone (CBX). Interestingly, the combination of VENLA and CBX is also associated with a higher rate of relapse after treatment withdrawal. To our knowledge, this study is one of the first to develop a model of relapse, and our results reveal that Cx-mediated dynamic neuroglial interactions play a critical role in the efficacy of monoaminergic antidepressant drugs, thus providing new targets for the treatment of depression.
Astrocytes , Connexins , Depressive Disorder , Animals , Mice , Antidepressive Agents/pharmacology , Antidepressive Agents/therapeutic use , Astrocytes/drug effects , Astrocytes/metabolism , Carbenoxolone/pharmacology , Connexins/drug effects , Connexins/metabolism , Phenotype , Recurrence , Depression/drug therapy , Depression/metabolism , Depressive Disorder/drug therapy , Depressive Disorder/metabolism
BACKGROUND: Alzheimer's disease (AD) research has relied on mouse models overexpressing human mutant A ßPP; however, newer generation knock-in models allow for physiological expression of amyloid-ß protein precursor (AßPP) containing familial AD mutations where murine AßPP is edited with a humanized amyloid-ß (Aß) sequence. The AppNL-F mouse model has shown substantial similarities to AD brains developing late onset cognitive impairment. OBJECTIVE: In this study, we aimed to characterize mature primary cortical neurons derived from homozygous AppNL-F embryos, especially to identify early mitochondrial alterations in this model. METHODS: Primary cultures of AppNL-F neurons kept in culture for 12-15 days were used to measure Aß levels, secretase activity, mitochondrial functions, mitochondrial-ER contacts, synaptic function, and cell death. RESULTS: We detected higher levels of Aß42 released from AppNL-F neurons as compared to wild-type neurons. AppNL-F neurons, also displayed an increased Aß42/Aß40 ratio, similar to adult AppNL-F mouse brain. Interestingly, we found an upregulation in mitochondrial oxygen consumption with concomitant downregulation in glycolytic reserve. Furthermore, AppNL-F neurons were more susceptible to cell death triggered by mitochondrial electron transport chain inhibition. Juxtaposition between ER and mitochondria was found to be substantially upregulated, which may account for upregulated mitochondrial-derived ATP production. However, anterograde mitochondrial movement was severely impaired in this model along with loss in synaptic vesicle protein and impairment in pre- and post-synaptic function. CONCLUSION: We show that widespread mitochondrial alterations can be detected in AppNL-F neurons in vitro, where amyloid plaque deposition does not occur, suggesting soluble and oligomeric Aß-species being responsible for these alterations.
Alzheimer Disease , Animals , Mice , Humans , Alzheimer Disease/metabolism , Mice, Transgenic , Amyloid beta-Peptides/metabolism , Amyloid beta-Protein Precursor/genetics , Amyloid beta-Protein Precursor/metabolism , Neurons/metabolism , Plaque, Amyloid/metabolism , Disease Models, Animal
The ion pump Na+,K+-ATPase is a critical determinant of neuronal excitability; however, its role in the etiology of diseases of the central nervous system (CNS) is largely unknown. We describe here the molecular phenotype of a Trp931Arg mutation of the Na+,K+-ATPase catalytic α1 subunit in an infant diagnosed with therapy-resistant lethal epilepsy. In addition to the pathological CNS phenotype, we also detected renal wasting of Mg2+. We found that membrane expression of the mutant α1 protein was low, and ion pumping activity was lost. Arginine insertion into membrane proteins can generate water-filled pores in the plasma membrane, and our molecular dynamic (MD) simulations of the principle states of Na+,K+-ATPase transport demonstrated massive water inflow into mutant α1 and destabilization of the ion-binding sites. MD simulations also indicated that a water pathway was created between the mutant arginine residue and the cytoplasm, and analysis of oocytes expressing mutant α1 detected a nonspecific cation current. Finally, neurons expressing mutant α1 were observed to be depolarized compared with neurons expressing wild-type protein, compatible with a lowered threshold for epileptic seizures. The results imply that Na+,K+-ATPase should be considered a neuronal locus minoris resistentia in diseases associated with epilepsy and with loss of plasma membrane integrity.
Epilepsy/genetics , Mutation, Missense , Sodium-Potassium-Exchanging ATPase/genetics , Animals , Anticonvulsants/pharmacology , Brain/drug effects , Brain/metabolism , Brain/pathology , Cells, Cultured , Drug Resistance , Epilepsy/drug therapy , Epilepsy/pathology , Humans , Infant , Molecular Dynamics Simulation , Mutation, Missense/drug effects , Protein Subunits/analysis , Protein Subunits/genetics , Sodium-Potassium-Exchanging ATPase/analysis , Xenopus
Major depression is a psychiatric disorder relying on different neurobiological mechanisms. In particular, a hypersensitivity of the hypothalamic-pituitary-adrenal axis leading to an excess of cortisol in blood and a deficit in monoaminergic neurotransmission have been associated with mood disorders. In keeping with these mechanisms, currently available antidepressant drugs act by increasing the extracellular levels of monoamines in the synaptic cleft. Since the discovery of the rapid and long-lasting antidepressant effects of ketamine, an NMDA receptor antagonist, a growing attention in psychiatry is paid to the pharmacological tools able to attenuate glutamatergic neurotransmission. Astrocytes play an important role in the excitatory/inhibitory balance of the central nervous system through the regulation of glutamate reuptake and secretion. Interestingly, the release of this excitatory amino acid is controlled, at least in part, by plasma membrane proteins (i.e. connexins) that cluster together to form gap junctions or hemichannels. Preclinical evidence suggests that these functional entities play a critical role in emotional behaviour. After a brief overview of the literature on mood disorders and related treatments, this review describes the role of astrocytes and connexins in glutamatergic neurotransmission and major depression. Moreover, we highlight the arguments supporting the therapeutic potential of connexins blockers but also the practical difficulties to target the hemichannels while maintaining gap junctions intact.
TITLE: Rôle des connexines astrocytaires dans la régulation des taux extracellulaires de glutamate : implication pour le traitement des épisodes dépressifs majeurs. ABSTRACT: La dépression majeure est une pathologie psychiatrique reposant sur différents mécanismes neurobiologiques. Parmi ces mécanismes, on trouve une hypersensibilité de l'axe hypothalamo-hypophyso-surrénalien associée à un excès de cortisol dans le sang et un déficit de neurotransmission monoaminergique. Ainsi, l'efficacité thérapeutique des antidépresseurs actuels repose sur leur capacité à augmenter les taux extracellulaires de monoamines dans la fente synaptique. Depuis la découverte des effets antidépresseurs rapides et durables de la kétamine, un antagoniste des récepteurs NMDA, un intérêt croissant est porté sur les moyens pharmacologiques atténuant l'action du glutamate pour traiter la dépression majeure. Les astrocytes jouent un rôle prépondérant dans la balance excitation/inhibition du système nerveux central en régulant la recapture et la sécrétion du glutamate. De manière intéressante, la libération de cet acide aminé excitateur est contrôlée, du moins en partie, par des canaux membranaires regroupés au niveau de jonctions intercellulaires de type « gap ¼ ou d'hémicanaux formés par les connexines 30 et 43. Les données précliniques suggèrent que ces deux entités fonctionnelles ont des effets sur les comportements émotionnels dans différents modèles murins de dépression. Après un bref rappel sur les troubles de l'humeur et leurs traitements, cette revue de la littérature décrit le rôle des astrocytes et des connexines dans la neurotransmission glutamatergique et la dépression majeure. Les arguments avancés soulignent l'intérêt thérapeutique potentiel du blocage des connexines astrocytaires mais aussi les difficultés pratiques à cibler la fonction hémicanal sans impacter la fonction « gap ¼.
Astrocytes , Connexins , Depressive Disorder, Major , Hypothalamo-Hypophyseal System , Pituitary-Adrenal System , Antidepressive Agents/therapeutic use , Astrocytes/metabolism , Connexins/metabolism , Depressive Disorder, Major/drug therapy , Depressive Disorder, Major/metabolism , Glutamic Acid/metabolism , Humans , Hypothalamo-Hypophyseal System/metabolism , Pituitary-Adrenal System/metabolism , Synapses , Synaptic Transmission/drug effects
Donepezil (DPZ) is an acetylcholinesterase inhibitor used in Alzheimer's disease to restore cognitive functions but is endowed with limited efficacy. Recent studies pointed out the implication of astroglial networks in cognitive processes, notably via astrocyte connexins (Cxs), proteins involved in gap junction intercellular communications. Hence, we investigated the impact on cognition of pharmacological or genetic modulations of those astrocyte Cxs during DPZ challenge in two rodent models of Alzheimer's disease-like memory deficits. We demonstrated that the Cx modulator mefloquine (MEF) significantly enhanced the procognitive effect of DPZ in both models. In parallel, we determined that MEF potentiated DPZ-induced release of acetylcholine in hippocampus. Finally, local genetic silencing of astrocyte Cxs in the hippocampus was also found to enhance the procognitive effect of DPZ, pointing out the importance of Cx-dependent astrocyte networks in memory processes.
AIM: Astroglial connexins (Cxs) 30 and 43 are engaged in gap junction and hemichannel activities. Evidence suggests that these functional entities contribute to regulating neurotransmission, thereby influencing brain functions. In particular, preclinical and clinical findings highlight a role of Cx43 in animal models of depression. However, the role of these proteins in response to currently available psychotropic drugs is still unknown. METHODS: To investigate this, we evaluated the behavioural effects of the genetic and pharmacological inactivation of Cx43 on the antidepressant- and anxiolytic-like activities of the selective serotonin reuptake inhibitor fluoxetine and the benzodiazepine diazepam, respectively. RESULTS: A single administration of fluoxetine (18 mg/kg; i.p.) produced a higher increase in hippocampal extracellular serotonin levels, and a greater antidepressant-like effect in the tail suspension test in Cx43 knock-down (KD) mice bred on a C57BL/6 background compared to their wild-type littermates. Similarly, in outbred Swiss wild-type mice, the intra-hippocampal injection of a shRNA-Cx43 or the acute systemic injection of the Cxs inhibitor carbenoxolone (CBX: 10 mg/kg; i.p.) potentiated the antidepressant-like effects of fluoxetine. Evaluating the effects of such strategies on diazepam (0.5 mg/kg; i.p.), the results indicate that Cx43 KD mice or wild-types injected with a shRNA-Cx43 in the amygdala, but not in the hippocampus, attenuated the anxiolytic-like effects of this benzodiazepine in the elevated plus maze. The chronic systemic administration of CBX mimicked the latter observations. CONCLUSION: Collectively, these data pave the way to the development of potentiating strategies in the field of psychiatry based on the modulation of astroglial Cx43.
Anti-Anxiety Agents/pharmacology , Antidepressive Agents/pharmacology , Astrocytes/drug effects , Connexin 43/antagonists & inhibitors , Connexin 43/genetics , Animals , Astrocytes/metabolism , Benzodiazepines/pharmacology , Diazepam/pharmacology , Fluoxetine/pharmacology , Male , Mice , Mice, Inbred C57BL , Selective Serotonin Reuptake Inhibitors/pharmacology
Astrocytes are glial cells organized in dynamic and structured networks in the brain. These plastic networks, involving key proteins such as connexin 43 (Cx43), are engaged in fine neuronal tuning and have recently been considered as emerging therapeutic targets in central nervous system disorders. We developed and validated a new application of the manganese-enhanced magnetic resonance imaging (MEMRI) technique allowing in vivo investigations of astrocyte-neuron interactions through quantification of brain Cx43 functional activity. The proof of concept has been achieved by quantification of MEMRI signals in brain after either local astrocyte-specific Cx43 knockdown with shRNA or systemic administration of Cx43 blockers. Unilateral hippocampal Cx43 genetical silencing was associated with an ipsilateral local increase of MEMRI signal. Furthermore, Cx43 blockers also enhanced MEMRI signal responses in hippocampus. Altogether, these data reveal the MEMRI technique as a tool for quantitative imaging of in vivo Cx43-dependent function in astrocytes under physiological and pathological conditions.
Astrocytes , Brain , Connexin 43/metabolism , Magnetic Resonance Imaging/methods , Manganese/metabolism , Neurons , Animals , Astrocytes/cytology , Astrocytes/metabolism , Brain/cytology , Brain/metabolism , Connexin 43/antagonists & inhibitors , Gene Knockdown Techniques , Male , Mice , Mice, Inbred C57BL , Neurons/cytology , Neurons/metabolism
Clinical and preclinical studies have implicated glial anomalies in major depression. Conversely, evidence suggests that the activity of antidepressant drugs is based, at least in part, on their ability to stimulate density and/or activity of astrocytes, a major glial cell population. Despite this recent evidence, little is known about the mechanism(s) by which astrocytes regulate emotionality. Glial cells communicate with each other through gap junction channels (GJCs), while they can also directly interact with neurons by releasing gliotransmitters in the extracellular compartment via an hemichannels (HCs)-dependent process. Both GJCs and HCs are formed by two main protein subunits: connexins (Cx) 30 and 43 (Cx30 and Cx43). Here we investigate the role of hippocampal Cx43 in the regulation of depression-like symptoms using genetic and pharmacological approaches. The first aim of this study was to evaluate the impact of the constitutive knock-down of Cx43 on a set of behaviors known to be affected in depression. Conversely, the expression of Cx43 was assessed in the hippocampus of mice subjected to prolonged corticosterone (CORT) exposure, given either alone or in combination with an antidepressant drug, the selective serotonin reuptake inhibitor fluoxetine. Our results indicate that the constitutive deficiency of Cx43 resulted in the expression of some characteristic hallmarks of antidepressant-/anxiolytic-like behavioral activities along with an improvement of cognitive performances. Moreover, in a new cohort of wild-type mice, we showed that CORT exposure elicited anxiety and depression-like abnormalities that were reversed by chronic administration of fluoxetine. Remarkably, CORT also increased hippocampal amounts of phosphorylated form of Cx43 whereas fluoxetine treatment normalized this parameter. From these results, we envision that antidepressant drugs may exert their therapeutic activity by decreasing the expression and/or activity of Cx43 resulting from a lower level of phosphorylation in the hippocampus.
