Astrocyte-Neuron Interaction via the Glutamate-Glutamine Cycle and Its Dysfunction in Tau-Dependent Neurodegeneration.

Astroglia constitute the largest group of glial cells and are involved in numerous actions that are critical to neuronal development and functioning, such as maintaining the blood-brain barrier, forming synapses, supporting neurons with nutrients and trophic factors, and protecting them from injury. These properties are deeply affected in the course of many neurodegenerative diseases, including tauopathies, often before the onset of the disease. In this respect, the transfer of essential amino acids such as glutamate and glutamine between neurons and astrocytes in the glutamate-glutamine cycle (GGC) is one example. In this review, we focus on the GGC and the disruption of this cycle in tau-dependent neurodegeneration. A profound understanding of the complex functions of the GGC and, in the broader context, searching for dysfunctions in communication pathways between astrocytes and neurons via GGC in health and disease, is of critical significance for the development of novel mechanism-based therapies for neurodegenerative disorders.

Dual Implications of Nanosilver-Induced Autophagy: Nanotoxicity and Anti-Cancer Effects.

In recent years, efforts have been made to identify new anti-cancer therapies. Various types of nanomaterials, including silver nanoparticles (AgNPs), are being considered as an option. In addition to its well-known antibacterial activity, AgNPs exhibit cytotoxic potential in both physiological and cancer cells by inducing stress-mediated autophagy and apoptotic cell death. A rapidly growing collection of data suggests that the proper regulation of autophagic machinery may provide an efficient tool for suppressing the development of cancer. In this light, AgNPs have emerged as a potential anti-cancer agent to support therapy of the disease. This review summarizes current data indicating the dual role of AgNP-induced autophagy and highlights factors that may influence its protective vs. its toxic potential. It also stresses that our understanding of the cellular and molecular mechanisms of autophagy machinery in cancer cells, as well as AgNP-triggered autophagy in both normal and diseased cells, remains insufficient.

Memantine Improves the Disturbed Glutamine and γ-Amino Butyric Acid Homeostasis in the Brain of Rats Subjected to Experimental Autoimmune Encephalomyelitis.

Glutamine (Gln), glutamate (Glu), and γ-amino butyric acid (GABA) are essential amino acids for brain metabolism and function. Astrocyte-derived Gln is the precursor for the two most important neurotransmitters in the central nervous system (CNS), which are the excitatory neurotransmitter Glu and the inhibitory neurotransmitter GABA. In addition to their roles in neurotransmission, these amino acids can be used as alternative substrates in brain metabolism that enable metabolic coupling between astrocytes and neurons in the glutamate-glutamine cycle (GGC). The disturbed homeostasis of these amino acids within the tripartite synapse may be involved in the pathogenesis of various neurological diseases. Interactions between astrocytes and neurons in terms of Gln, Glu, and GABA homeostasis were studied in different phases of experimental allergic encephalomyelitis (EAE) in Lewis rats. The results of the study showed a decrease in the transport (uptake and release) of Gln and GABA in both neuronal and astrocyte-derived fractions. These effects were fully or partially reversed when the EAE rats were treated with memantine, a NMDA receptor antagonist. Changes in the expression and activity of selected glutamine/glutamate metabolizing enzymes, such as glutamine synthase (GS) and phosphate-activated glutaminase (PAG), which were affected by memantine, were observed in different phases of EAE. The results suggested perturbed homeostasis of Gln, Glu, and GABA during EAE, which may indicate alterations in neuron-astrocyte coupling and dysfunction of the tripartite synapse. Memantine appears to partially regulate the disturbed relationships between Gln, Glu, and GABA.

Mutant Tau protein-induced abnormalities in the Na+-dependent glutamine translocation and recycling and their impact on astrocyte-neuron integrity in vitro.

Tau-dependent neurodegeneration is accompanied by astrocytosis in a mouse trans-genic model, which replicates the neuropathological characteristic of tauopathy and other human neurodegenerative disorders where astrocyte activation precedes neuronal loss and is associated with disease progression. This indicates an important role of astrocytes in the development of the disease. Astrocytes derived from a transgenic mouse model expressing human Tau, exhibit changes in cellular markers of astrocyte neuroprotective function related to the glutamate-glutamine cycle (GGC), representing a key part of astrocyte-neuron integrity. Here, we focused on investigating the functional properties of key GGC components involved in the astrocyte-neuron network associated with Tau pathology in vitro. Mutant recombinant Tau (rTau) carrying the P301L mutation was added to the neuronal cultures, with or without control astrocyte-conditioned medium (ACM), to study glutamine translocation through the GGC. We demonstrated that mutant Tau in vitro induces neuronal degeneration, while control astrocytes response in neuroprotective way by preventing neurodegeneration. In parallel with this observation, we noticed the Tau-dependent decline of neuronal microtubule associated protein 2 (MAP2), followed by changes in glutamine (Gln) transport. Exposure to rTau decreases sodium-dependent Gln uptake in neurons and that effect was reversed when cells were co-incubated with control ACM after induction of rTau dependent pathology. Further, we found that neuronal Na+-dependent system A is the most specific system that is affected under rTau exposure. In addition, in rTau-treated astrocytes total Na+-dependent uptake of Gln, which is mediated by the N system, increases. Altogether, our study suggest mechanisms operating in Tau pathology may be related to the alterations in glutamine transport and recycling that affect neuronal-astrocytic integrity.

Endoplasmic Reticulum Stress Underlies Nanosilver-Induced Neurotoxicity in Immature Rat Brain.

The growing production of silver nanoparticles (AgNPs), and their widespread use in medical and consumer products, poses a potential threat to the environment and raises questions about biosafety. Immature organisms are particularly susceptible to various insults during development. The biological characteristics of immature organisms are different from those of adults, and dictate the consequences of exposure to various toxic substances, including AgNPs. Nanoparticles are highly reactive and can easily cross the blood-brain barrier (BBB) to accumulate in brain tissues. It is therefore important to investigate the molecular mechanisms of AgNP-induced neurotoxicity in the developing brain. Immature 2-week-old rats were exposed to a low dose of AgNPs (0.2 mg/kg b.w.) over a long period. Subsequently, brain tissues of the animals were subjected to ultrastructural and molecular analyses to determine endoplasmic reticulum (ER) stress. Ultrastructural markers of ER stress, such as pathological alterations in the ER and elongated forms of mitochondria accompanied by autophagy structures, were confirmed to be present in AgNP-exposed rat brain. Evidence for induction of ER stress in neurons was also provided by molecular markers. Upregulation of genes related to the ER-stress-induced unfolded protein response (UPR) pathway, such as GRP78, PERK, and CHOP ATF-6, was observed at the transcriptional and translational levels. The results show that prolonged exposure of immature rats to a low dose of AgNPs during the developmental period leads to induction of ER stress in the neurons of the developing brain. Simultaneously, in response to AgNP-induced ER stress, neurons promote protective mechanisms that partially compensate for ER stress by regulating the biodynamic processes of mitochondria and autophagy.

Developmental neurotoxicity of silver nanoparticles: the current state of knowledge and future directions.

The increasing production and use of silver nanoparticles (AgNPs) as an antimicrobial agent in an array of medical and commercial products, including those designed for infants and children, poses a substantial risk of exposure during the developmental period. This review summarizes current knowledge on developmental neurotoxicity of AgNPs in both pre- and post-natal stages with a focus on the biological specificity of immature organisms that predisposes them to neurotoxic insults as well as the molecular mechanisms underlying AgNP-induced neurotoxicity. The current review revealed that AgNPs increase the permeability of the blood-brain barrier (BBB) and selectively damage neurons in the brain of immature rats exposed pre and postnatally. Among the AgNP-induced molecular mechanisms underlying toxic insult is cellular stress, which can consequently lead to cell death. Glutamatergic neurons and NMDAR-mediated neurotransmission also appear to be a target for AgNPs during the postnatal period of exposure. Collected data indicate also that our current knowledge of the impact of AgNPs on the developing nervous system remains insufficient and further studies are required during different stages of development with investigation of environmentally-relevant doses of exposure.

Alterations in the transcriptional profile of genes related to glutamatergic signalling in animal models of Alzheimer's disease. The effect of fingolimod.

Alzheimer's disease (AD) is a multi-factorial illness that leads to progressive cognitive impairment. A glutamatergic system dysfunction has been reported to be implicated in the pathomechanism of AD. Therefore, in the current study we characterized the transcriptional profile of glutamate-related genes in transgenic AbPP V717I (TgAD) and sporadic (SAD, streptozotocin-induced) models of AD. Genes encoding glutamate membrane-bound (GLAST, GLT1, EAAC1) and vesicular (VGLUT1-3) transporters as well as ionotropic (AMPA, NMDA) and metabotropic (mGluR3, mGluR5) receptors were analysed. Based on qPCR analysis, we observed a discrepancy between TgAD and SAD mice in the profile of targeted genes. We noticed age-dependent upregulation of genes encoding VGLUT1, NMDAR1 and mGluR3 in 12-month-old TgAD mice. In the SAD model upregulation of genes encoding AMPAR1 and NMDAR1 as well as downregulation of GLAST, VGLUT3 and mGluR5 were found. Next, the effect of fingolimod (FTY720) was indicated. In the TgAD model, the drug reversed altered transcription of the mGluR3 glutamate receptor to the control level, whereas in the SAD model it downregulated the genes encoding VGLUT1, AMPAR2 and mGluR3. Interestingly, FTY720 influenced mGluR3 mRNA in both examined models. Observed alterations of gene transcription and the effects of FTY720 may potentially constitute an interesting target for further pharmacological studies.

Memantine Modulates Oxidative Stress in the Rat Brain following Experimental Autoimmune Encephalomyelitis.

Experimental autoimmune encephalomyelitis (EAE) is an animal model most commonly used in research on the pathomechanisms of multiple sclerosis (MS). The inflammatory processes, glutamate excitotoxicity, and oxidative stress have been proposed as determinants accompanying demyelination and neuronal degeneration during the course of MS/EAE. The aim of the current study was to characterize the role of NMDA receptors in the induction of oxidative stress during the course of EAE. The effect of memantine, the uncompetitive NMDA receptor antagonist, on modulation of neurological deficits and oxidative stress in EAE rats was analyzed using several experimental approaches. We demonstrated that the expression of antioxidative enzymes (superoxide dismutases SOD1 and SOD2) were elevated in EAE rat brains. Under the same experimental conditions, we observed alterations in oxidative stress markers such as increased levels of malondialdehyde (MDA) and decreased levels of sulfhydryl (-SH) groups, both protein and non-protein (indicating protein damage), and a decline in reduced glutathione. Importantly, pharmacological inhibition of ionotropic NMDA glutamate receptors by their antagonist memantine improved the physical activity of EAE rats, alleviated neurological deficits such as paralysis of tail and hind limbs, and modulated oxidative stress parameters (MDA, -SH groups, SOD's). Furthermore, the current therapy aiming to suppress NMDAR-induced oxidative stress was partially effective when NMDAR's antagonist was administered at an early (asymptomatic) stage of EAE.

Nanosystems and exosomes as future approaches in treating multiple sclerosis.

Multiple sclerosis (MS) is an immune-mediated demyelinating disease of the central nervous system which leads to neurological dysfunctions and severe disabilities. MS pathology is characterised by damage of the blood-brain barrier and infiltration of autoreactive T cells that overactivate glial cells, thereby initiating neuroinflammation accompanied by the formation of demyelinating plaques and neurodegeneration. Clinical deficits in this multifactorial disease depend on the progression of myelin loss, the stage of inflammation, the status of axons and the activity of oligodendrocyte precursor cells (OPCs). Despite significant progress in the treatment of MS, current therapies remain limited and new approaches are highly desirable. Nanosystems based on liposomes and nanoparticles are among some of the more noteworthy therapeutic strategies being investigated. Applications of nanosystems alone or as drug carriers in animal models of MS have been found to successfully alleviate the symptoms of the disease and exert anti-inflammatory potential. Exosomes are a specific type of nanosystem based on nanometre-sized extracellular vesicles released by different cells which exhibit important healing features. Exosomes contain an array of anti-inflammatory and neuroprotective agents which may contribute to modulation of the immune system as well as promoting remyelination and tissue repair. In this review, opportunities to use nanosystems against progression of MS will be discussed in context of cell-specific pathologies associated with MS.

Astroglial and Microglial Purinergic P2X7 Receptor as a Major Contributor to Neuroinflammation during the Course of Multiple Sclerosis.

Multiple sclerosis (MS) is an autoimmune inflammatory disease of the central nervous system that leads to the progressive disability of patients. A characteristic feature of the disease is the presence of focal demyelinating lesions accompanied by an inflammatory reaction. Interactions between autoreactive immune cells and glia cells are considered as a central mechanism underlying the pathology of MS. A glia-mediated inflammatory reaction followed by overproduction of free radicals and generation of glutamate-induced excitotoxicity promotes oligodendrocyte injury, contributing to demyelination and subsequent neurodegeneration. Activation of purinergic signaling, in particular P2X7 receptor-mediated signaling, in astrocytes and microglia is an important causative factor in these pathological processes. This review discusses the role of astroglial and microglial cells, and in particular glial P2X7 receptors, in inducing MS-related neuroinflammatory events, highlighting the importance of P2X7R-mediated molecular pathways in MS pathology and identifying these receptors as a potential therapeutic target.

Early and Delayed Impact of Nanosilver on the Glutamatergic NMDA Receptor Complex in Immature Rat Brain.

Silver nanoparticles (AgNPs) are the one of the most extensively used nanomaterials. The strong antimicrobial properties of AgNPs have led to their use in a wide range of medical and consumer products. Although the neurotoxicity of AgNPs has been confirmed, the molecular mechanisms have not been extensively studied, particularly in immature organisms. Based on information gained from previous in vitro studies, in the present work, we examine whether ionotropic NMDA glutamate receptors contribute to AgNP-induced neurotoxicity in an animal model of exposure. In brains of immature rats subjected to a low dose of AgNPs, we identified ultrastructural and molecular alterations in the postsynaptic region of synapses where NMDA receptors are localized as a multiprotein complex. We revealed decreased expression of several NMDA receptor complex-related proteins, such as GluN1 and GluN2B subunits, scaffolding proteins PSD95 and SynGAP, as well as neuronal nitric oxide synthase (nNOS). Elucidating the changes in NMDA receptor-mediated molecular mechanisms induced by AgNPs, we also identified downregulation of the GluN2B-PSD95-nNOS-cGMP signaling pathway which maintains LTP/LTD processes underlying learning and memory formation during development. This observation is accompanied by decreased density of NMDA receptors, as assessed by a radioligand binding assay. The observed effects are reversible over the post-exposure time. This investigation reveals that NMDA receptors in immature rats are a target of AgNPs, thereby indicating the potential health hazard for children and infants resulting from the extensive use of products containing AgNPs.

Response of immature rats to a low dose of nanoparticulate silver: Alterations in behavior, cerebral vasculature-related transcriptome and permeability.

The increasing production and use of silver nanoparticles (AgNPs) as antimicrobial agents in medicinal and commercial products creates a substantial risk of exposure, especially for infants and children. Our current knowledge concerning the impact of AgNPs on developing brain is insufficient. Therefore we investigated the temporal profile of transcriptional changes in cellular components of the neurovascular unit in immature rats exposed to a low dose of AgNPs. The behavior of animals under these conditions was also monitored. Significant deposition of AgNPs in brain of exposed rats was identified and found to persist over the post-exposure time. Substantial changes were noted in the transcriptional profile of tight junction proteins such as occludin and claudin-5, and pericyte-related molecules such as angiopoietin-1. Moreover, downregulation of platelet-derived growth factor (PDGFß) and its receptor (PDGFßR) which constitute the main signaling pathway between endothelial cells and pericytes was observed. These were long-lasting effects, accompanied by overexpression of astroglial-specific GFAP mRNA and endothelial cell adhesion molecule, ICAM-1, involved in the pathomechanism of neuroinflammation. The profile of changes indicates that even low doses of AgNPs administered during the early stage of life induce dysregulation of neurovascular unit constituents which may lead to disintegration of the blood-brain barrier. This was confirmed by ultrastructural analysis that revealed enhanced permeability of cerebral microvessels resulting in perivascular edema. Changes in the behavior of exposed rats indicating pro-depressive and anti-anxiety impacts were also identified. The results show a high risk of using AgNPs in medical and consumer products dedicated for infants and children.

Dysfunctional glia: contributors to neurodegenerative disorders.

Astrocytes are integral components of the central nervous system, where they are involved in numerous functions critical for neuronal development and functioning, including maintenance of blood-brain barrier, formation of synapses, supporting neurons with nutrients and trophic factors, and protecting them from injury. These roles are markedly affected in the course of chronic neurodegenerative disorders, often before the onset of the disease. In this review, we summarize the recent findings supporting the hypothesis that astrocytes play a fundamental role in the processes contributing to neurodegeneration. We focus on α-synucleinopathies and tauopathies as the most common neurodegenerative diseases. The mechanisms implicated in the development and progression of these disorders appear not to be exclusively neuronal, but are often related to the astrocytic-neuronal integrity and the response of astrocytes to the altered microglial function. A profound understanding of the multifaceted functions of astrocytes and identification of their communication pathways with neurons and microglia in health and in the disease is of critical significance for the development of novel mechanism-based therapies against neurodegenerative disorders.

Early Postnatal Exposure to a Low Dose of Nanoparticulate Silver Induces Alterations in Glutamate Transporters in Brain of Immature Rats.

Due to strong antimicrobial properties, silver nanoparticles (AgNPs) are used in a wide range of medical and consumer products, including those dedicated for infants and children. While AgNPs are known to exert neurotoxic effects, current knowledge concerning their impact on the developing brain is scarce. During investigations of mechanisms of neurotoxicity in immature rats, we studied the influence of AgNPs on glutamate transporter systems which are involved in regulation of extracellular concentration of glutamate, an excitotoxic amino acid, and compared it with positive control-Ag citrate. We identified significant deposition of AgNPs in brain tissue of exposed rats over the post-exposure time. Ultrastructural alterations in endoplasmic reticulum (ER) and Golgi complexes were observed in neurons of AgNP-exposed rats, which are characteristics of ER stress. These changes presumably underlie substantial long-lasting downregulation of neuronal glutamate transporter EAAC1, which was noted in AgNP-exposed rats. Conversely, the expression of astroglial glutamate transporters GLT-1 and GLAST was not affected by exposure to AgNPs, but the activity of the transporters was diminished. These results indicate that even low doses of AgNPs administered during an early stage of life create a substantial risk for health of immature organisms. Hence, the safety of AgNP-containing products for infants and children should be carefully considered.

A Low Dose of Nanoparticulate Silver Induces Mitochondrial Dysfunction and Autophagy in Adult Rat Brain.

Extensive incorporation of silver nanoparticles (AgNPs) into many medical and consumer products has raised concerns about biosafety. Since nanosilver accumulates persistently in the central nervous system, it is important to assess its neurotoxic impacts. We investigated a model of prolonged exposure of adult rats to a low environmentally relevant dose of AgNPs (0.2 mg/kg b.w.). Ultrastructural analysis revealed pathological alterations in mitochondria such as swelling and cristolysis. Besides, elongated forms of mitochondria were present. Level of adenosine triphosphate was not altered after exposure, although a partial drop of mitochondrial membrane potential was noted. Induction of autophagy with only early autophagic forms was observed in AgNP-exposed rat brains as evidenced by ultrastructural markers. Increased expression of two protein markers of autophagy, beclin 1 and microtubule-associated proteins 1A/1B light chain 3B (MAP LC3-II), was observed, indicating induction of autophagy. Expression of lysosome-related Rab 7 protein and cathepsin B did not change, suggesting inhibition of physiological flux of autophagy. Our results show that exposure to a low, environmentally relevant dose of AgNPs leads to induction of autophagy in adult rat brain in response to partial mitochondrial dysfunction and to simultaneous interfering with an autophagic pathway. The cell compensates for the defective autophagy mechanism via development of enhanced mitochondrial biodynamic.

Prolonged inflammation leads to ongoing damage after spinal cord injury.

The pathogenesis of spinal cord injury (SCI) remains poorly understood and treatment remains limited. Emerging evidence indicates that post-SCI inflammation is severe but the role of reactive astrogliosis not well understood given its implication in ongoing inflammation as damaging or neuroprotective. We have completed an extensive systematic study with MRI, histopathology, proteomics and ELISA analyses designed to further define the severe protracted and damaging inflammation after SCI in a rat model. We have identified 3 distinct phases of SCI: acute (first 2 days), inflammatory (starting day 3) and resolution (>3 months) in 16 weeks follow up. Actively phagocytizing, CD68+/CD163- macrophages infiltrate myelin-rich necrotic areas converting them into cavities of injury (COI) when deep in the spinal cord. Alternatively, superficial SCI areas are infiltrated by granulomatous tissue, or arachnoiditis where glial cells are obliterated. In the COI, CD68+/CD163- macrophage numbers reach a maximum in the first 4 weeks and then decline. Myelin phagocytosis is present at 16 weeks indicating ongoing inflammatory damage. The COI and arachnoiditis are defined by a wall of progressively hypertrophied astrocytes. MR imaging indicates persistent spinal cord edema that is linked to the severity of inflammation. Microhemorrhages in the spinal cord around the lesion are eliminated, presumably by reactive astrocytes within the first week post-injury. Acutely increased levels of TNF-alpha, IL-1beta, IFN-gamma and other pro-inflammatory cytokines, chemokines and proteases decrease and anti-inflammatory cytokines increase in later phases. In this study we elucidated a number of fundamental mechanisms in pathogenesis of SCI and have demonstrated a close association between progressive astrogliosis and reduction in the severity of inflammation.

Astroglial contribution to tau-dependent neurodegeneration.

Astrocytes, by maintaining an optimal environment for neuronal function, play a critical role in proper function of mammalian nervous system. They regulate synaptic transmission and plasticity and protect neurons against toxic insults. Astrocytes and neurons interact actively via glutamine-glutamate cycle (GGC) that supports neuronal metabolic demands and neurotransmission. GGC deficiency may be involved in different diseases of the brain, where impaired astrocytic control of glutamate homeostasis contributes to neuronal dysfunction. This includes tau-dependent neurodegeneration, where astrocytes lose key molecules involved in regulation of glutamate/glutamine homeostasis, neuronal survival and synaptogenesis. Astrocytic dysfunction in tauopathy appears to precede neurodegeneration and overt tau neuropathology such as phosphorylation, aggregation and formation of neurofibrillary tangles. In this review, we summarize recent studies demonstrating that activation of astrocytes is strictly associated with neurodegenerative processes including those involved in tau related pathology. We propose that astrocytic dysfunction, by disrupting the proper neuron-glia signalling early in the disease, significantly contributes to tauopathy pathogenesis.

Early P2X7R-dependent activation of microglia during the asymptomatic phase of autoimmune encephalomyelitis.

Microglia-mediated neuroinflammation accompanies many central nervous system (CNS) diseases, including multiple sclerosis (MS), and is strongly dependent on the purinergic P2X7 receptor. The nature of the inflammatory response in MS is studied for decades indicating, that proinflammatory microgliosis is involved in advanced stages of MS and is associated with active tissue damage and neurological dysfunctions. Evidence on the role of microgliosis in initial stages of the disease is scarce. Thus, in the present study, we investigated the time course of microglial activation in rat brain subjected to experimental autoimmune encephalomyelitis (EAE) which is the animal model of MS. We show that activation of microglia occurs in brains of immunized rats at a very early stage of EAE, well before the development of neurological symptoms of the disease. Enhanced immunoreactivity of microglia/macrophage-specific protein Iba-1, together with morphological features of microgliosis, was identified beginning at day 4 post immunization. Concomitantly, microglial expression of P2X7R was also examined. Moreover, our results reveal that administration of Brilliant Blue G, an antagonist of P2X7R, delays the onset of the disease and partially inhibits development of neurological symptoms in EAE rats. Blockage of P2X7R significantly reduces activation of microglia as confirmed by decreased Iba-1 immunoreactivity and suppresses neuroinflammation in EAE rat brains, as indicated by decreased protein levels of investigated proinflammatory cytokines: IL-1ß, IL-6 and TNF-α. Our results indicate that microglia are involved in inducing neuroinflammation at a very early stage of MS/EAE via a P2X7R-dependent mechanism.

Prolonged Exposure to Silver Nanoparticles Results in Oxidative Stress in Cerebral Myelin.

Currently, silver nanoparticles (AgNPs) are frequently used in a wide range of medical and consumer products. Substantial usage of AgNPs is considered to create substantive risks to both the environment and the human health. Since there is increasing evidence that the main mechanism of toxicity of AgNPs relates to oxidative stress, in the current study we investigate oxidative stress-related biochemical parameters in myelin isolated from adult rat brain subjected to a low dose of AgNPs. Animals were exposed for 2 weeks to 0.2 mg/kg b.w. of small (10 nm) AgNPs stabilized in citrate buffer or silver citrate established as a control to compare the effects of particulate and ionic forms of silver. We observe enhanced peroxidation of lipids and decreased concentrations of protein and non-protein -SH groups in myelin membranes. Simultaneously, expression of superoxide dismutase, a free radical scavenger, is increased whereas the process of protein glutathionylation, being a cellular protective mechanism against irreversible oxidation, is found to be inefficient. Results indicate that oxidative stress-induced alterations in myelin membranes may be the cause of ultrastructural disturbances in myelin sheaths.

Administration of an antagonist of P2X7 receptor to EAE rats prevents a decrease of expression of claudin-5 in cerebral capillaries.

Purinergic P2X receptors, when activated under pathological conditions, participate in induction of the inflammatory response and/or cell death. Both neuroinflammation and neurodegeneration represent hallmarks of multiple sclerosis (MS), an autoimmune disease of the central nervous system. In the current study, we examined whether P2X7R is expressed in brain microvasculature of rats subjected to experimental autoimmune encephalomyelitis (EAE) and explore possible relationships with blood-brain barrier (BBB) protein-claudin-5 after administration of P2X7R antagonist-Brilliant Blue G (BBG). Capillary fraction isolated from control and EAE rat brains was subjected to immunohistochemical and Western blot analyses. We document the presence of P2X7R in brain capillaries isolated from brain tissue of EAE rats. P2X7R is found to be localized on the abluminal surface of the microvessels and is co-expressed with PDGFßR, a marker of pericytes. We also show over-expression of this receptor in isolated capillaries during the course of EAE, which is temporally correlated with a lower protein level of PDGFßR, as well as claudin-5, a tight junction-building protein. Administration of a P2X7R antagonist to the immunized rats significantly reduced clinical signs of EAE and enhances protein expression of both claudin-5 and PDGFßR. These results indicate that P2X7 receptor located on pericytes may contribute to pathological mechanisms operated during EAE in cerebral microvessels influencing the BBB integrity.