Peripheral Lipopolysaccharide Injection Research Articles

Acute neuroinflammation leads to disruption of neuronal chloride regulation and consequent hyperexcitability in the dentate gyrus

Neuroinflammation is a salient part of diverse neurological and psychiatric pathologies that associate with neuronal hyperexcitability, but the underlying molecular and cellular mechanisms remain to be identified. Here, we show that peripheral injection of lipopolysaccharide (LPS) renders the dentate gyrus (DG) hyperexcitable to perforant pathway stimulation invivo and increases the internal spiking propensity of dentate granule cells (DGCs) invitro 24h post-injection (hpi). In parallel, LPS leads to a prominent downregulation of chloride extrusion via KCC2 and to the emergence of NKCC1-mediated chloride uptake in DGCs under experimental conditions optimized to detect specific changes in transporter efficacy. These data show that acute neuroinflammation leads to disruption of neuronal chloride regulation, which unequivocally results in a loss of GABAergic inhibition in the DGCs, collapsing the gating function of the DG. The present work provides a mechanistic explanation for neuroinflammation-driven hyperexcitability and consequent cognitive disturbance.

Icilin, a cool/cold-inducing agent, alleviates lipopolysaccharide-induced septic sickness responses in mice

Sepsis is a significant global public health challenge, resulting in millions of human deaths annually. Transient receptor potential melastatin 8 (TRPM8), a non-selective ion channel, is the primary cold sensor in humans; however, its effects on endotoxin-induced inflammation remain unclear. We previously reported that TRPM8 knockout mice exhibited more severe physiological and behavioral endotoxemia responses upon a high-dose injection with lipopolysaccharide (LPS). In the present study, we investigated whether icilin, a TRPM8 agonist, was a target for the suppression of sickness responses using a mouse model of LPS-induced sepsis. A peripheral high-dose injection of LPS at 5 mg/kg showed a maximal body temperature decrease of 5.1 °C in mice subcutaneously pretreated with vehicle and 1.5 °C in icilin-pretreated animals. The decline in locomotor activity was attenuated in icilin-pretreated mice and its recovery was faster; however, the high-dose LPS injection rapidly decreased locomotor activity regardless of the icilin pretreatment. Furthermore, the icilin pretreatment attenuated LPS-induced decreases in body weight and food and water intakes and accelerated recovery from these sickness responses. Therefore, the present results demonstrated that the icilin pretreatment alleviated LPS-induced sickness responses or decreases in body temperature, locomotor activity, body weight loss, and food and water intakes, suggesting its potential as a therapeutic target for sepsis.

Menstrual blood-derived endometrial stem cells inhibit neuroinflammation by regulating microglia through the TLR4/MyD88/NLRP3/Casp1 pathway.

Neuroinflammation is a common response in various neurological disorders. Mesenchymal stem cell-based treatment has become a promising therapy for neuroinflammation-associated diseases. However, the effects of mesenchymal stem cells are controversial, and the underlying mechanism is incompletely understood. In the present study, menstrual blood-derived endometrial stem cells were intravenously transplanted into a mouse model of neuroinflammation established by peripheral injection of lipopolysaccharide. Microglial cells challenged with lipopolysaccharide were cultured with conditioned medium from endometrial stem cells. The levels of cytokines were detected by enzyme-linked immunosorbent assay. Cell proliferation and death were detected by Cell Counting Kit 8 and flow cytometry, respectively. The expression levels of Toll-like receptor 4 (TLR4), myeloid differentiation primary response gene 88 (MyD88), NLR family pyrin domain containing 3 (NLRP3) and caspase 1 (Casp1) were evaluated by western blotting. The results showed that intravenous transplantation of endometrial stem cells downregulated proinflammatory factors and upregulated anti-inflammatory factors in the brain of mice with neuroinflammation. Conditioned medium suppressed the inflammatory reaction and hyperactivation of microglial cells and protected microglial cells from cell death induced by lipopolysaccharide in vitro. The expression of TLR4, MyD88, NLRP3 and Casp1 in the brain of mice with neuroinflammation and in lipopolysaccharide-stimulated microglial cells was downregulated by endometrial stem cells and conditioned medium, respectively. These data suggested that menstrual blood-derived endometrial stem cells may suppress neuroinflammatory reactions partially by regulating microglia through the TLR4/MyD88/NLRP3/Casp1 signalling pathway. Our findings may be very useful for the development of an alternative stem cell-based therapy for neuroinflammation-associated disorders.

Melanocortin-4 receptors on neurons in the parabrachial nucleus mediate inflammation-induced suppression of food-seeking behavior

Anorexia is a common symptom during infectious and inflammatory disease. Here we examined the role of melanocortin-4 receptors (MC4Rs) in inflammation-induced anorexia. Mice with transcriptional blockage of the MC4Rs displayed the same reduction of food intake following peripheral injection of lipopolysaccharide as wild type mice but were protected against the anorexic effect of the immune challenge in a test in which fasted animals were to use olfactory cues to find a hidden cookie. By using selective virus-mediated receptor re-expression we demonstrate that the suppression of the food-seeking behavior is subserved by MC4Rs in the brain stem parabrachial nucleus, a central hub for interoceptive information involved in the regulation of food intake. Furthermore, the selective expression of MC4R in the parabrachial nucleus also attenuated the body weight increase that characterizes MC4R KO mice. These data extend on the functions of the MC4Rs and show that MC4Rs in the parabrachial nucleus are critically involved in the anorexic response to peripheral inflammation but also contribute to body weight homeostasis during normal conditions.

Peripheral Immune Activation in Mice Elicits Unfolded Protein Responses Independent on MyD88 Pathway in the Liver but not the Hypothalamus and Hippocampus

Neuroimmune interactions between the immune system and CNS as well as peripheral organs such as the liver play a key role in the pathophysiological state of diseases. Unfolded protein responses (UPRs), which are activated by cells in response to endoplasmic reticulum stress, have been linked to the occurrence of inflammation diseases, neurodegenerative diseases, and metabolic disorders such as type 2 diabetes. Peripheral injection of lipopolysaccharide (LPS) is known to induce a systemic inflammatory response, along with fever, anorexia, and depressive behaviors. LPS also elicits UPRs, although the underlying physiological mechanism remains unclear. In the present study, we investigated whether peripheral activation of the immune system can elicit UPRs in the CNS and liver. Peripheral injection of LPS is known to elevate pro-inflammatory cytokines in the liver, hypothalamus and hippocampus. We report that LPS-induced systemic inflammation elicits UPRs in the liver, but not the hypothalamus. Injection of LPS upregulated the expression levels of glucose-regulated protein 78 and pro-apoptotic transcription factor C/EBP homologous protein, along with increased splicing of X-box binding protein one mRNA in the liver, but not in the hypothalamus and hippocampus. Myeloid differentiation primary response 88 (MyD88), an adaptor protein, is known to play a key role in the signal transduction of LPS mediated by Toll-like receptor 4. Using MyD88 deficient mice, we found that LPS-induced UPRs occurred independently of MyD88 expression. In summary, peripheral activation of the immune system elicits UPRs in the liver, but not the hypothalamus and hippocampus, which may have implications for the pathophysiology of diseases.

Microglial Cannabinoid Type 1 Receptor Regulates Brain Inflammation in a Sex-Specific Manner.

Background: Neuroinflammation is a key feature shared by most, if not all, neuropathologies. It involves complex biological processes that act as a protective mechanism to fight against the injurious stimuli, but it can lead to tissue damage if self-perpetuating. In this context, microglia, the main cellular actor of neuroinflammation in the brain, are seen as a double-edged sword. By phagocyting neuronal debris, these cells can not only provide tissue repair but can also contribute to neuronal damage by releasing harmful substances, including inflammatory cytokines. The mechanisms guiding these apparent opposing actions are poorly known. The endocannabinoid system modulates the release of inflammatory factors such as cytokines and could represent a functional link between microglia and neuroinflammatory processes. According to transcriptomic databases and in vitro studies, microglia, the main source of cytokines in pathological conditions, express the cannabinoid type 1 receptor (CB1R). Methods: We thus developed a conditional mouse model of CB1R deletion specifically in microglia, which was subjected to an immune challenge (peripheral lipopolysaccharide injection). Results: Our results reveal that microglial CB1R differentially controls sickness behavior in males and females. Conclusion: These findings add to the comprehension of neuroinflammatory processes and might be of great interest for future studies aimed at developing therapeutic strategies for brain disorders with higher prevalence in men.

Vortioxetine Prevents Lipopolysaccharide-Induced Memory Impairment Without Inhibiting the Initial Inflammatory Cascade.

Vortioxetine is a novel multimodal antidepressant that modulates a wide range of neurotransmitters throughout the brain. Preclinical and clinical studies have shown that vortioxetine exerts positive effects on different cognitive domains and neuroprotective effects. Considering the key role of microglial cells in brain plasticity and cognition, we aimed at investigating the effects of pretreatment with vortioxetine in modulating behavioral and molecular effects induced by an immune challenge: peripheral injection of lipopolysaccharide (LPS). To this purpose, C57BL/6J male mice were first exposed to a 28-day standard diet or vortioxetine-enriched diet, which was followed by an acute immune challenge with LPS. Sickness symptoms and depressive-like behaviors (anhedonia and memory impairment) were tested 6 and 24 h after exposure to LPS, respectively. Moreover, the expressions of markers of immune activation and M1/M2 markers of microglia polarization were measured in the dorsal and ventral parts of the hippocampus. The pretreatment with vortioxetine did not affect both LPS-induced sickness behavior and anhedonia but prevented the deficit in the recognition memory induced by the immune challenge. At the transcriptional level, chronic exposure to vortioxetine did not prevent LPS-induced upregulation of proinflammatory cytokines 6 h after the immune challenge but rather seemed to potentiate the immune response to the challenge also by affecting the levels of expression of markers of microglia M1 phenotype, like cluster of differentiation (CD)14 and CD86, in an area-dependent manner. However, at the same time point, LPS injection significantly increased the expression of the M2 polarization inducer, interleukin 4, only in the hippocampus of animals chronically exposed to vortioxetine. These results demonstrate that a chronic administration of vortioxetine specifically prevents LPS-induced memory impairment, without affecting acute sickness behavior and anhedonia, and suggest that hippocampal microglia may represent a cellular target of this novel antidepressant medication. Moreover, we provide a useful model to further explore the molecular mechanisms specifically underlying cognitive impairments following an immune challenge.

Acute blockade of endogenous melatonin by Luzindole, with or without peripheral LPS injection, induces jejunal inflammation and morphological alterations in Swiss mice.

This study investigated the acute blockade of endogenous melatonin (MLT) using Luzindole with or without systemic lipopolysaccharide (LPS) challenge and evaluated changes in inflammatory and oxidative stress markers in the mouse jejunum. Luzindole is an MT1/MT2 MLT receptor antagonist. Both receptors occur in the small intestine. Swiss mice were treated with either saline (0.35 mg/kg, ip), Luzindole (0.35 mg/kg, ip), LPS (1.25 mg/kg, ip), or Luzindole+LPS (0.35 and 1.25 mg/kg, ip, respectively). Jejunum samples were evaluated regarding intestinal morphometry, histopathological crypt scoring, and PAS-positive villus goblet cell counting. Inflammatory Iba-1, interleukin (IL)-1β, tumor necrosis factor (TNF)-α, nuclear factor (NF)-kB, myeloperoxidase (MPO), and oxidative stress (NP-SHs, catalase, MDA, nitrate/nitrite) markers were assessed. Mice treated with Luzindole, LPS, and Luzindole+LPS showed villus height shortening. Crypt damage was worse in the LPS group. Luzindole, LPS, and Luzindole+LPS reduced the PAS-goblet cell labeling and increased Iba-1-immunolabelled cells compared to the saline group. Immunoblotting for IL-1β, TNF-α, and NF-kB was greater in the Luzindole group. The LPS-challenged group showed higher MPO activity than the saline and Luzindole groups. Catalase was reduced in the Luzindole and Luzindole+LPS groups compared to saline. The Luzindole group showed an increase in NP-SHs, an effect related to compensatory GSH activity. The acute blockade of endogenous MLT with Luzindole induced early changes in inflammatory markers with altered intestinal morphology. The other non-detectable deleterious effects of Luzindole may be balanced by the unopposed direct action of MLT in immune cells bypassing the MT1/MT2 receptors.

Lipopolysaccharide-Induced Exosomal miR-146a Is Involved in Altered Expression of Alzheimer's Risk Genes Via Suppression of TLR4 Signaling.

Repeated exposure to toll-like receptor 4 (TLR4) ligands, such as lipopolysaccharide (LPS), reduces responses of monocytes/macrophages to LPS (LPS/endotoxin tolerance). Microglial exposure to Aβ deposits, a TLR4 ligand, may cause "Aβ/LPS tolerance," leading to decreased Aβ clearance. We demonstrated that microglial activation by LPS is diminished in Aβ deposit-bearing 12-month-old model mice of Alzheimer's disease (AD), compared with non-AD mice and Aβ deposit-free 2-month-old AD mice. Because miR-146a plays a predominant role in inducing TLR tolerance in macrophages and because miR-146a in extracellular vesicles (EVs) shed by inflammatory macrophages increases in circulation, we investigated potential roles of miR-146a and inflammatory EVs in inducing TLR tolerance in microglia and in altering expression of inflammatory AD risk genes. We found that miR-146a upregulation induces TLR tolerance and alters expression of inflammatory AD risk genes in response to LPS treatment in BV2 microglia. LPS brain injection altered expression of the AD risk genes in 12-month-old AD mice but not in non-AD littermates. EVs from inflammatory macrophages polarize BV2 microglia to M1 phenotype and induce TLR tolerance. Microglia exposed to Aβ in the brain show reduced cytokine responses to systemic inflammation due to peripheral LPS injection, indicating TLR/Aβ tolerance in microglia. Our results suggest that increased miR-146a induces microglial Aβ/LPS tolerance and that circulating EVs shed by inflammatory macrophages contribute to microglial Aβ/LPS tolerance, leading to reduced Aβ clearance. Our study also suggests that altered expression of inflammatory AD risk genes may contribute to AD development via the same molecular mechanism underlying LPS tolerance.

Effect of Pentoxifylline on Apoptotic-Related Gene Expression Profile, Learning and Memory Impairment Induced by Systemic Lipopolysaccharide Administration in the Rat Hippocampus.

Background:Inflammation is one of the effective factors, in the development of functional disorders of the nervous system. Pentoxifylline (PTX) has an inhibitory effect on inflammatory factors. Therefore the aim of this study was to evaluate the effect of PTX on learning, memory and expression of genes, involved in neuronal survival in the rat hippocampus, following systemic lipopolysaccharide (LPS) injection.Methods:Male rats were randomly divided into 5 groups of control, LPS and LPS + PTX, receiving doses of 10, 25 and 50 mg/kg of PTX, respectively. In LPS groups, LPS was injected (5 mg/kg; intraperitoneal), and after one week, rats received intraperitoneal PTX for 14 days, in the treatment groups. Learning and memory were evaluated by object location task (OLT) and novel object recognition (NOR). Then, the hippocampus was dissected in order to measure the expression of the associated genes.Results:The results showed that peripheral LPS injection caused significant damage (P < 0.01) to learning and memory with respect to controls, but PTX with doses of 10 and 50 mg/kg prevented these impairments. Results from reverse transcription polymerase chain reaction (RT-PCR) showed that LPS significantly increased the expression of Bax and TNF- α with respect to controls. PTX in the LPS + PTX group significantly increased the expression of Bcl-2, BAD and Caspase-3.Conclusions:Other than the increased Bcl-2 expression, PTX had no significant effect on the expression of other genes, therefore further studies are needed to find out how PTX improves the learning and memory impairments, following the peripheral inflammation.

ABCA7 haplodeficiency disturbs microglial immune responses in the mouse brain

Carrying premature termination codons in 1 allele of the ABCA7 gene is associated with an increased risk for Alzheimer's disease (AD). While the primary function of ABCA7 is to regulate the transport of phospholipids and cholesterol, ABCA7 is also involved in maintaining homeostasis of the immune system. Since inflammatory pathways causatively or consequently participate in AD pathogenesis, we studied the effects of Abca7 haplodeficiency in mice on brain immune responses under acute and chronic conditions. When acute inflammation was induced through peripheral lipopolysaccharide injection in control or heterozygous Abca7 knockout mice, partial ABCA7 deficiency diminished proinflammatory responses by impairing CD14 expression in the brain. On breeding to AppNL-G-F knockin mice, we observed increased amyloid-β (Aβ) accumulation and abnormal endosomal morphology in microglia. Taken together, our results demonstrate that ABCA7 loss of function may contribute to AD pathogenesis by altering proper microglial responses to acute inflammatory challenges and during the development of amyloid pathology, providing insight into disease mechanisms and possible treatment strategies.

Circadian clock protein Rev-erbα regulates neuroinflammation

Circadian dysfunction is a common attribute of many neurodegenerative diseases, most of which are associated with neuroinflammation. Circadian rhythm dysfunction has been associated with inflammation in the periphery, but the role of the core clock in neuroinflammation remains poorly understood. Here we demonstrate that Rev-erbα, a nuclear receptor and circadian clock component, is a mediator of microglial activation and neuroinflammation. We observed time-of-day oscillation in microglial immunoreactivity in the hippocampus, which was disrupted in Rev-erbα-/- mice. Rev-erbα deletion caused spontaneous microglial activation in the hippocampus and increased expression of proinflammatory transcripts, as well as secondary astrogliosis. Transcriptomic analysis of hippocampus from Rev-erbα-/- mice revealed a predominant inflammatory phenotype and suggested dysregulated NF-κB signaling. Primary Rev-erbα-/- microglia exhibited proinflammatory phenotypes and increased basal NF-κB activation. Chromatin immunoprecipitation revealed that Rev-erbα physically interacts with the promoter regions of several NF-κB-related genes in primary microglia. Loss of Rev-erbα in primary astrocytes had no effect on basal activation but did potentiate the inflammatory response to lipopolysaccharide (LPS). In vivo, Rev-erbα-/- mice exhibited enhanced hippocampal neuroinflammatory responses to peripheral LPS injection, while pharmacologic activation of Rev-erbs with the small molecule agonist SR9009 suppressed LPS-induced hippocampal neuroinflammation. Rev-erbα deletion influenced neuronal health, as conditioned media from Rev-erbα-deficient primary glial cultures exacerbated oxidative damage in cultured neurons. Rev-erbα-/- mice also exhibited significantly altered cortical resting-state functional connectivity, similar to that observed in neurodegenerative models. Our results reveal Rev-erbα as a pharmacologically accessible link between the circadian clock and neuroinflammation.

The Effect of Pentoxifylline on Passive Avoidance Learning and Expression of Tumor Necrosis Factor-Alpha and Caspase-3 in the Rat Hippocampus Following Lipopolysaccharide-Induced Inflammation.

Background:Peripheral inflammation is effective in the development of neurodegenerative diseases. Pentoxifylline (PTX) has an inhibitory effect on inflammatory cytokines; therefore, we aimed to evaluate the effect of PTX on passive avoidance learning and the expression of tumor necrosis factor-alpha (TNF-α) and caspase-3 in the rat hippocampus, following systemic lipopolysaccharide (LPS) injection.Materials and Methods:Male Wistar rats were randomly divided into five groups: control, LPS, and LPS + PTX, receiving doses of 10, 25, and 50 mg/kg of PTX, respectively. The animals received daily injections of PTX (i.p.) 1 week before and 2 weeks after the LPS injection (5 mg/kg; i.p.). Learning and memory were evaluated by passive avoidance learning. Then, the expression of the associated genes was measured in the hippocampus.Results:The results showed that the peripheral LPS injection had no significant effect on learning and memory. PTX only with a dose of 10 mg/kg shows an improvement (P < 0.05). Results from reverse transcription polymerase chain reaction showed that LPS had no significant effect on the expression of caspase-3 and TNF-α. PTX with a dose of 10 mg/kg decreased the caspase-3 expression in the LPS + PTX group (P < 0.001), but the expression of both genes increased, using other concentrations.Conclusions:Findings showed that systemic application of LPS after 2 weeks had no effect on learning and memory and the expression of inflammatory genes in the hippocampus, but PTX led to an increase in the expression of these genes, which could be due to its direct effects or possible exacerbation of LPS effects.

Recovery from an acute systemic and central LPS-inflammation challenge is affected by mouse sex and genetic background

Genetic and sexual factors influence the prevalence and the pathogenesis of many inflammatory disorders. In this study their relevance on the peripheral and central inflammatory status induced by a peripheral injection of lipopolysaccharide (LPS) was evaluated. BALB/c and CD-1 male and female mice were intraperitoneally injected with LPS. Spleens and brains were collected 2 and 72 hours later to study the levels of IL-6, TNF-α and IL-1β. Percentage of microglia and astrocytes was determined in the cortex and hippocampus. Locomotor activity was registered before and during the 72 hours after LPS-treatment. Two hours after LPS-injection, a peripheral increase of the three cytokines was found. In brains, LPS increased TNF-α only in males with higher levels in CD-1 than BALB/c. IL-1β increased only in CD-1 males. IL-6 increased in both strains with lower levels in BALB/c females. Peripheral and central levels of cytokines decline 72 hrs after LPS-treatment whilst a significantly increase of Iba-1 expression was detected. A dramatic drop of the locomotor activity was observed immediately after LPS injection. Our results show that acute systemic administration of LPS leads to peripheral and central increase of pro-inflammatory cytokines and microglia activation, in a strain and sex dependent manner.

Monocytes, microglia, and CD200-CD200R1 signaling are essential in the transmission of inflammation from the periphery to the central nervous system.

Peripheral inflammation is known to trigger neuroinflammation and neurodegenerative disease. However, the key components during the propagation of inflammation from the periphery to the central nervous system (CNS) remain unclear. Lipopolysaccharide (LPS) was administered to Sprague-Dawley rats to induce peripheral inflammation. An intravenous injection and an intranigral injection of clodronate liposomes were given to deplete monocytes and microglia, respectively. Recombinant CD200 fusion protein (CD200Fc) or an anti-CD200R1 antibody was injected into the substantia nigra to manipulate the involvement of CD200 and CD200R1. Immunohistochemistry and immunofluorescence staining were used to measure microglial activation and dopaminergic neuronal loss. The expression of brain pro-inflammatory cytokines (i.e., tumor necrosis factor alpha, IL-1β) and CD200-CD200R1 signaling were measured by quantitative RT-PCR. Our data showed that the peripheral LPS injection activated the microglia and induced an increase in the levels of pro-inflammatory cytokines (i.e., tumor necrosis factor alpha, IL-1β). The depletion of either monocytes or microglia suppressed these inflammatory effects that were induced by peripheral LPS administration. The peripheral LPS injection increased the expression of CD200 and CD200R1 in the substantia nigra. Dopaminergic neuronal loss induced by the peripheral LPS injection was accelerated by the blockade of CD200-CD200R1 signaling with an anti-CD200R1 antibody and attenuated by intensifying the signaling with CD200Fc. These results highlight the importance of monocytes, microglia, and CD200-CD200R1 signaling in the transmission of inflammation from the periphery to the CNS.

Central CCL2 signaling onto MCH neurons mediates metabolic and behavioral adaptation to inflammation.

Sickness behavior defines the endocrine, autonomic, behavioral, and metabolic responses associated with infection. While inflammatory responses were suggested to be instrumental in the loss of appetite and body weight, the molecular underpinning remains unknown. Here, we show that systemic or central lipopolysaccharide (LPS) injection results in specific hypothalamic changes characterized by a precocious increase in the chemokine ligand 2 (CCL2) followed by an increase in pro-inflammatory cytokines and a decrease in the orexigenic neuropeptide melanin-concentrating hormone (MCH). We therefore hypothesized that CCL2 could be the central relay for the loss in body weight induced by the inflammatory signal LPS. We find that central delivery of CCL2 promotes neuroinflammation and the decrease in MCH and body weight. MCH neurons express CCL2 receptor and respond to CCL2 by decreasing both electrical activity and MCH release. Pharmacological or genetic inhibition of CCL2 signaling opposes the response to LPS at both molecular and physiologic levels. We conclude that CCL2 signaling onto MCH neurons represents a core mechanism that relays peripheral inflammation to sickness behavior.

Elevation of cortical serotonin transporter activity upon peripheral immune challenge is regulated independently of p38 mitogen-activated protein kinase activation and transporter phosphorylation.

The serotonin transporter (SERT) is responsible for high-affinity serotonin (5-HT) uptake from extracellular fluid and is a prominent pharmacological target in the treatment of depression. In recent years, depression has also been linked to immune system activation. Inflammatory conditions can cause sickness behaviour and depression-like symptoms in both animals and humans. Since SERT has been proposed as one of the molecular targets in inflammation-induced depression, we applied the widely used lipopolysaccharides (LPS) model to study the effects of peripheral inflammation on SERT activity in the brain. We show that 24h after intraperitoneal LPS administration, SERT-mediated 5-HT uptake is significantly enhanced in the frontal cortex. Analysis of uptake kinetics revealed that the transport capacity (Vmax ) of cortical SERT was increased in LPS-injected animals, while the Km value remained unchanged. The increase in Vmax was neither due to increased SERT protein expression nor increased synaptic surface exposure. The suppression of SERT activity upon inhibition of p38 MAPK was not selective for LPS-induced enhancement of SERT function. In addition, SERT activity changes in LPS-treated rats are unaffected by nitric oxide synthase and protein kinase G inhibitors. Using the Phos-Tag method, we identified five SERT-specific protein bands representing distinct phosphorylation states of SERT. However, the enhancement of SERT activity in LPS-treated rats was not correlated with altered transporter phosphorylation. Together with previous studies by others, our results suggest that SERT is regulated by multiple mechanisms in response to peripheral immune system activation. Peripheral injection of lipopolysaccharide (LPS) induces characteristic sickness and depression-like behaviour in rats over a period of at least 24h. We show here that the activity of the serotonin transporter (SERT), a prominent antidepressant target, is up-regulated 24h following LPS administration. In contrast to previous studies focusing on earlier responses to LPS treatment, we found that SERT function is selectively enhanced in the frontal cortex, independently of the p38 MAPK pathway. Our study provides further insight into molecular mechanisms underlying inflammation-induced depression.

Molecular and cellular neuroinflammatory status of mouse brain after systemic lipopolysaccharide challenge: importance of CCR2/CCL2 signaling.

BackgroundGenetic and environmental factors are critical elements influencing the etiology of major depression. It is now accepted that neuroinflammatory processes play a major role in neuropsychological disorders. Neuroinflammation results from the dysregulation of the synthesis and/or release of pro- and anti-inflammatory cytokines with central or peripheral origin after various insults. Systemic bacterial lipopolysaccharide (LPS) challenge is commonly used to study inflammation-induced depressive-like behaviors in rodents. In the present study, we investigated immune-to-brain communication in mice by examining the effects of peripheral LPS injection on neuroinflammation encompassing cytokine and chemokine production, microglia and central nervous system (CNS)-associated phagocyte activation, immune cell infiltration and serotonergic neuronal function.MethodsLPS was administered to C57BL/6 J mice by intraperitoneal injection; brains were collected and pro-inflammatory cytokine mRNA and proteins were measured. To examine the relative contribution of the different populations of brain immune cells to the occurrence of neuroinflammation after acute systemic inflammation, we precisely characterized them by flow cytometry, studied changes in their proportions and level of activation, and measured the amount of cytokines they released by Cytometric Bead Array™ after cell sorting and ex vivo culture. Because of the central role that the chemokine CCL2 seems to play in our paradigm, we studied the effect of CCL2 on the activity of serotonergic neurons of the raphe nucleus using electrophysiological recordings.ResultsWe report that systemic LPS administration in mice caused a marked increase in pro-inflammatory IL-1β, IL-6, TNFα and CCL2 (monocyte chemoattractant protein-1) mRNA and protein levels in the brain. Moreover, we found that LPS caused microglia and CNS-associated phagocyte activation characterized by upregulation of CCR2, TLR4/CD14, CD80 and IL-4Rα, associated with overproduction of pro-inflammatory cytokines and chemokines, especially CCL2. LPS also induced a marked and selective increase of CCR2+ inflammatory monocytes within the brain. Finally, we showed that CCL2 hyperpolarized serotonergic raphe neurons in mouse midbrain slices, thus probably reducing the serotonin tone in projection areas.ConclusionTogether, we provide a detailed characterization of the molecular and cellular players involved in the establishment of neuroinflammation after systemic injection of LPS. This highlights the importance of the CCL2/CCR2 signaling and suggests a possible link with depressive disorders.

A New Animal Model of Depression Induced by Repeated Central Lipopolysaccharide Administration

The administration of lipopolysaccharide(LPS), a component of the cell wall of gram negative bacteria, is commonly-used method to cause immune activation and the release of cytokines both in the periphery and in the brain in rodents. The cytokine theory of depression indicated that pro-inflammatory cytokines play an important role in the pathological mechanism of depression. Many studies used the peripheral administration of LPS to induce depressive-like behaviors. However, in these studies, the altered behaviors usually last only a few hours, seldom longer than 24h. Such short-term depressive behavior can hardly be regarded as a model of depression.. Thus, the aim of the present study was therefore to develop a new animal model of depression with apparent depressive-like behavior at- and after- 24h post-LPS injection. In this study, single and triple central LPS administration were used to induce depressive-like behavior respectively. Sprague-Dawley rats were randomly divided into LPS group and control group. LPS(100ng/rat, one injection; or once every second day, total three times) or isotonic saline was administered by intracerebroventricular microinjection. The depressive-like behavior was measured by preference to saccharin, locomotor activity and immobility time of tail suspension. The result indicated that single central LPS injection induced partial depressive-like behaviors. There was significant difference in locomotor activity, but not in the preference of saccharin and immobility time of tail suspension. However, repeated central LPS administration induced significant depressive-like behaviors after 24h of the last LPS injection. The animals with triple central LPS administration consumed less saccharin solution, exhibited less locomotor activity in the open field, and maintained immobility time in tail suspension. The changes in locomotor activity and immobility time of tail suspension were even apparent until 72h after the last LPS injection. Our results demonstrate that a new effective model of depression can be established by means of repeated lateral ventricle LPS injections, and the induced depressive-like behavior has longer time duration than by the peripheral injection of LPS.

Brain protein expression changes in WAG/Rij rats, a genetic rat model of absence epilepsy after peripheral lipopolysaccharide treatment

Peripheral injection of bacterial lipopolysaccharide (LPS) facilitates 8-10Hz spike-wave discharges (SWD) characterizing absence epilepsy in WAG/Rij rats. It is unknown however, whether peripherally administered LPS is able to alter the generator areas of epileptic activity at the molecular level. We injected 1mg/kg dose of LPS intraperitoneally into WAG/Rij rats, recorded the body temperature and EEG, and examined the protein expression changes of the proteome 12h after injection in the fronto-parietal cortex and thalamus. We used fluorescent two-dimensional differential gel electrophoresis to investigate the expression profile. We found 16 differentially expressed proteins in the fronto-parietal cortex and 35 proteins in the thalamus. It is known that SWD genesis correlates with the transitional state of sleep-wake cycle thus we performed meta-analysis of the altered proteins in relation to inflammation, epilepsy as well as sleep. The analysis revealed that all categories are highly represented by the altered proteins and these protein-sets have considerable overlap. Protein network modeling suggested that the alterations in the proteome were largely induced by the immune response, which invokes the NFkB signaling pathway. The proteomics and computational analysis verified the known functional interplay between inflammation, epilepsy and sleep and highlighted proteins that are involved in their common synaptic mechanisms. Our physiological findings support the phenomenon that high dose of peripheral LPS injection increases SWD-number, modifies its duration as well as the sleep-wake stages and decreases body temperature.