CUMS-induced Mice Research Articles

Long Noncoding RNA LINC00473 Ameliorates Depression-Like Behaviors in Female Mice by Acting as a Molecular Sponge to Regulate miR-497-5p/BDNF Axis.

Background Depression was a common life-threatening psychiatric disorder and occurs more frequently in women than in men. Long noncoding RNAs (lncRNAs), such as LINC00473, had been reported to be involved in the progression of depression. Methods Chronic unpredictable moderate stress in mice (CUMS) was applied to construct a depression model. Subsequently, RT-qPCR was applied to check the level of LINC00473 and microRNA-497-5p (miR-497-5p) in the hippocampal region of the mice induced by CUMS. CUMS mice were injected with lentiviral vectors of LINC00473 (LV-LINC00473), miR-497-5p inhibitor, short hairpin- (sh-) brain-derived neurotrophic factor (sh-BDNF), or miR-497-5p mimic to evaluate depressive behaviors, including sucrose preference test, forced swim test, elevated plus maze, and tail suspension test. Moreover, the production of hypothalamic neurotransmitters was assessed with the usage of ELISA kits. Dual-luciferase reporter assay, RNA pull-down, and RIP analysis were performed to measure the relationship between miR-497-5p and LINC00473 or BDNF. Further, western blot was employed to determine the protein level of BDNF. Results We discovered that LINC00473 level was downregulated in the female mice with depression, but not in male mice. Besides, the depressive behaviors induced by CUMS in mice, including the decrease of sucrose preference and time in open arm, as well as the increase of immobility time and swimming resting time were all ameliorated by LINC00473 overexpression. Moreover, the concentration of neurotransmitters was decreased in CUMS-induced mouse hypothalamus, which was blocked by LV-LINC00473 lentiviral vector administration. Mechanistically, LINC00473 directly targeted miR-497-5p. Absence of miR-497-5p revealed the antidepression effects on CUMS-induced mice, and miR-497-5p upregulation could counter the antidepressive impacts of LINC00473 upregulation on CUMS-induced mice. Furthermore, LINC00473 could target miR-497-5p to modulate BDNF level. Knockdown of BDNF could abrogate the improving influences of miR-497-5p suppression on CUMS-induced depression. Conclusions LINC00473 ameliorated CUMS-caused depression by encouraging BDNF expression via binding to miR-497-5p, which might provide a potential therapeutic target for depression in females.

SIRT1/FOXO1 Axis-Mediated Hippocampal Angiogenesis is Involved in the Antidepressant Effect of Chaihu Shugan San.

ObjectiveChaihu Shugan San (CSS) has a long history for treating major depressive disorder (MDD), which has been verified effectively and safely in clinical studies. Deficient angiogenesis plays important roles in MDD. However, the underlying mechanisms of CSS on angiogenesis remain poorly understood.MethodsNetwork pharmacology analysis was applied to explore the potential angiogenic targets and pathways between CSS and MDD. These targets would be validated in chronic unpredictable mild stress (CUMS)-induced depressive-like mice by Western blots, immunofluorescence, and immunohistochemistry. Then, the underlying molecular mechanisms were further investigated in brain microvascular endothelial cells (BMVECs) with CSS-containing serum by Western blots and immunofluorescence.ResultsNetwork pharmacology analysis showed that the antidepressant role of CSS was closely associated with Silent information regulator protein 1 (SIRT1)/Forkhead box O1 (FOXO1) axis-mediated angiogenesis. This prediction was confirmed in the following experiments. CSS induced angiogenesis, increased SIRT1 expression, and decreased FOXO1 expression in the hippocampus of CUMS mice. Five percent CSS-containing serum produced a significant increase in BMVECs proliferation, migration, and tube formation, but these effects were reduced by SIRT1 silencing. CSS serum could also promote FOXO1 translocation to the cytoplasm through SIRT1 signaling, which triggered FOXO1 protein degradation. What is more, CSS upregulated VEGFA and BDNF expressions not only in the hippocampus of depressive mice but also in BMVECs supernatants. Of note, these trophic factors play important roles in promoting neurogenesis.ConclusionThe study showed that CSS could promote angiogenesis and neurogenesis in the hippocampus of CUMS-induced mice. The underlying molecular mechanism involves the SIRT1/FOXO1 axis and subsequent regulation of VEGFA and BDNF. These findings provide novel insight into CSS drug development, and targeting the SIRT1/FOXO1 axis might be a promising strategy to treat MDD.

Ginsenoside Rh2 administration produces crucial antidepressant-like effects in a CUMS-induced mice model of depression.

IntroductionThe most striking feature of depression is sadness and a loss of interest in activities, which represents a major cause of disability globally. Therefore, it is necessary to identify novel antidepressants for clinical practice. Ginsenoside Rh2 (Rh2) is one of the major bioactive ginsenosides that can be extracted from Panax ginseng and has been demonstrated to improve both memory and learning. The purpose of this study was to provide broad insight into the mechanisms underlying depression and gain greater insights into antidepressant therapy.MethodsIn this study, we first established an effective and feasible depression animal model of chronic unpredictable mild stress (CUMS) and behavioral testing was evaluated by the forced swim test (FST), the tail suspension test (TST) and the sucrose preference test. Following pretreatment with Rh2 (10 and 20 mg/kg), the immobility time of mice was reduced without affecting locomotor activity in both the FST and TST. Western blotting and immunofluorescence were used to investigate the activation of the hippocampal BDNF signaling pathway and hippocampal neurogenesis.ResultsDifferent concentrations of Rh2 significantly reduced depressive‐like symptoms in CUMS‐induced mice and downregulated the effects of the BDNF signaling cascade and neurogenesis in the hippocampus. Furthermore, the administration of K252a completely prevented the antidepressant‐like activity of Rh2 in mice.ConclusionThe results indicated that Rh2 possesses the antidepression action via the positive regulation of the BDNF‐TrkB pathway.

Antidepressant Mechanism of Traditional Chinese Medicine Formula Xiaoyaosan in CUMS-Induced Depressed Mouse Model via RIPK1-RIPK3-MLKL Mediated Necroptosis Based on Network Pharmacology Analysis.

Background: Depression is a stress-related disorder that seriously threatens people’s physical and mental health. Xiaoyaosan is a classical traditional Chinese medicine formula, which has been used to treat mental depression since ancient times. More and more notice has been given to the relationship between the occurrence of necroptosis and the pathogenesis of mental disorders. Objective: The purpose of present study is to explore the potential mechanism of Xiaoyaosan for the treatment of depression using network pharmacology and experimental research, and identify the potential targets of necroptosis underlying the antidepressant mechanism of Xiaoyaosan. Methods: The mice model of depression was induced by chronic unpredictable mild stress (CUMS) for 6 weeks. Adult C57BL/6 mice were randomly divided into five groups, including control group, chronic unpredictable mild stress group, Xiaoyaosan treatment group, necrostatin-1 (Nec-1) group and solvent group. Drug intervention performed from 4th to 6th week of modeling. The mice in Xiaoyaosan treatment group received Xiaoyaosan by intragastric administration (0.254 g/kg/d), and mice in CUMS group received 0.5 ml physiological saline. Meanwhile, the mice in Nec-1 group were injected intraperitoneally (i.p.) with Nec-1 (10 mg/kg/d), and the equivalent volume of DMSO/PBS (8.3%) was injected into solvent group mice. The behavior tests such as sucrose preference test, forced swimming test and novelty-suppressed feeding test were measured to evaluate depressive-like behaviors of model mice. Then, the active ingredients in Xiaoyaosan and the related targets of depression and necroptosis were compiled through appropriate databases, while the “botanical drugs-active ingredients-target genes” network was constructed by network pharmacology analysis. The expressions of RIPK1, RIPK3, MLKL, p-MLKL were detected as critical target genes of necroptosis and the potential therapeutic target compounds of Xiaoyaosan. Furthermore, the levels of neuroinflammation and microglial activation of hippocampus were measured by detecting the expressions of IL-1β, Lipocalin-2 and IBA1, and the hematoxylin and eosin (H&E) stained was used to observe the morphology in hippocampus sections. Results: After 6-weeks of modeling, the behavioral data showed that mice in CUMS group and solvent group had obvious depressive-like behaviors, and the medication of Xiaoyaosan or Nec-1 could improve these behavioral changes. A total of 96 active ingredients in Xiaoyaosan which could regulate the 23 key target genes were selected from databases. Xiaoyaosan could alleviate the core target genes in necroptosis and improve the hippocampal function and neuroinflammation in depressed mice. Conclusion: The activation of necroptosis existed in the hippocampus of CUMS-induced mice, which was closely related to the pathogenesis of depression. The antidepressant mechanism of Xiaoyaosan included the regulation of multiple targets in necroptosis. It also suggested that necroptosis could be a new potential target for the treatment of depression.

MicroRNA-135b-5p Downregulation Causes Antidepressant Effects by Regulating SIRT1 Expression.

Depression is a serious and potentially life-threatening mental illness. Recently, the role of sirtuin 1 (SIRT1) in chronic unpredictable mild stress (CUMS) management has been examined. The present study explored and clarified whether microRNA (miR)-135b-5p could play a role in depression by regulating the expression of SIRT1. SIRT1 was identified as the target gene of miR-135b-5p using TargetScan and the dual luciferase reporter assay. In addition, the expression levels of SIRT1 were significantly reduced in mouse peripheral blood and hippocampal tissue samples, while the expression of miR-135b-5p exhibited the opposite effects. Subsequently, the effects of miR-135b-5p inhibition were investigated in mice with depression. The results indicated that the miR-135b-5p inhibitor significantly increased the weight loss induced by CUMS compared with the model group, while reducing the expression levels of miR-135b-5p and further alleviating the depression-like behavior induced by CUMS. Concomitantly, the results indicated that the miR-135b-5p inhibitor inhibited CUMS-induced hippocampal cell apoptosis and significantly reduced the expression levels of cleaved caspase-3 and the ratio of cleaved caspase-3/caspase-3. Moreover, the miR-135b-5p inhibitor significantly reduced the CUMS-induced increase of the inflammatory factors IL-1β, IL-6 and TNF-α in the hippocampal mouse samples, while significantly increasing the expression levels of SIRT1. Finally, the results demonstrated that all the effects of the miR-135b-5p inhibitor on CUMS-induced mice were significantly reversed by SIRT1 silencing. In conclusion, the present study indicated that the miR-135b-5p/SIRT1 pathway was a key mediator of antidepressant effects induced in depressed mice. Therefore, it could be considered a potential therapeutic target for the treatment of CUMS-induced depression.

Detrimental Effect of Alprazolam on Sperm Viability ofMice (Mus musculus) Exposed to Chronic Unpredictable Mild Stress

Objective: This study aims to investigate the effect of alprazolam administration on sperm viability of mice (Mus musculus) strain Balb/C following CUMSfor 53 days. Method: This study used a true experimental design through randomization. Control group kept undisturbed without treatment and therapy throughout 60 days. K1 and K2 were exposed to a 53 day period of CUMS along with 7-day pretreatment adaptation. K2 was adapted for a week, stressed for 53 days along with cotreatment of alprazolam (4 mg/kg BW). Results: K1 and K2 showed insignificant improvement of sperm viability (p > 0.05).Meanwhile, K0 and K1 showed significant value compared to K2 (p<0.05). K2 showed little increase of sperm viability in CUMS group along with subsequent treatment of alprazolam. Conclusion: Overall, the results indicate an insignificant effect of alprazolam on sperm viability of CUMS-induced mice.

Baicalin promotes hippocampal neurogenesis via the Wnt/β-catenin pathway in a chronic unpredictable mild stress-induced mouse model of depression

Hippocampal neurogenesis is known to be related to depressive symptoms. Increasing evidence indicates that Wnt/β-catenin signaling regulates multiple aspects of adult hippocampal neurogenesis. Baicalin is a major flavonoid compound with multiple pharmacological effects such as anti-inflammatory, anti-apoptotic, and neuroprotective effects. The current study aimed to explore the antidepressant effects of baicalin and its possible molecular mechanisms affecting hippocampal neurogenesis via the regulation of the Wnt/β-catenin signaling pathway. A chronic mild unpredictable stress (CUMS) model of depression was used in the study. The CUMS-induced mice were treated with baicalin (50 and 100 mg/kg) for 21 days, orally, and the fluoxetine was used as positive control drug. The results indicated that baicalin alleviated CUMS-induced depression-like behaviour, and improved the nerve cells’ survival of the hippocampal dentate gyrus (DG) in CUMS-induced depression of model mice and increased Ki-67- and doublecortin (DCX)-positive cells to restore CUMS-induced suppression of hippocampal neurogenesis. The related proteins in the Wnt/β-catenin signaling pathway, which declined in the CUMS-induced depression model of mice, were upregulated after baicalin treatment, including Wingless3a (Wnt3a), dishevelled2 (DVL2), and β-catenin. Further study found that the phosphorylation rate of glycogen synthase kinase-3β (GSK3β) and β-catenin nuclear translocation increased, as the levels of the β-catenin target genes cyclinD1, c-myc, NeuroD1, and Ngn2 upregulated after baicalin treatment. In conclusion, these findings suggest that baicalin may promote hippocampal neurogenesis, thereby exerting the antidepressant effect via regulation of the Wnt/β-catenin signaling pathway.

Traditional Chinese Formula Xiaoyaosan Alleviates Depressive-Like Behavior in CUMS Mice by Regulating PEBP1-GPX4-Mediated Ferroptosis in the Hippocampus

BackgroundAt present, the pathogenesis of depression is not fully understood, and nearly half of depression patients experience no obvious effects during treatment. This study aimed to establish a depression mouse model to explore the possible role of ferroptosis in the pathogenesis of depression, and observe the effects of Xiaoyaosan on PEBP1-GPX4-mediated ferroptosis in the hippocampus.MethodsForty-eight male C57BL/6 mice were randomly divided into a control group, CUMS group, Xiaoyaosan group and fluoxetine group, and the model was established by chronic unpredictable mild stress (CUMS) for a successive 6 weeks. The medication procedure was performed from the 4th to the 6th week of modeling. The behavioral evaluations were measured to evaluate depressive-like behaviors. The expressions of GPX4, FTH1, ACSL4 and COX2 were detected as ferroptosis-related indicators. Then, the total iron and ferrous content in the hippocampus were measured. The levels of PEBP1 and ERK1/2 were observed, and the expressions of GFAP and IBA1 were also detected to measure the functions of astrocytes and microglia in the hippocampus.ResultsEight herbs of Xiaoyaosan had 133 active ingredients which could regulate the 43 ferroptosis-related genes in depression. After 6 weeks of modeling, the data showed that mice in the CUMS group had obvious depressive-like behaviors, and medication with Xiaoyaosan or fluoxetine could significantly improve the behavioral changes. The expressions of GPX4, FTH1, ACSL4, COX2, PEBP1, ERK1/2, GFAP and IBA1 changed in the CUMS group mice, while the total iron and ferrous content also changed. Xiaoyaosan and fluoxetine had obvious curative effects that could significantly alleviate the above changes in the hippocampus.ConclusionOur results revealed that the activation of ferroptosis might exist in the hippocampi of CUMS-induced mice. The PEBP1-GPX4-mediated ferroptosis could be involved in the antidepressant mechanism of Xiaoyaosan. It also implied that ferroptosis could become a new target for research into the depression mechanism and antidepressant drugs.

Antidepressant-Like Effect of Geniposide in Mice Exposed to a Chronic Mild Stress Involves the microRNA-298-5p-Mediated Nox1.

Depression is a common mental disorder that presents a considerable challenge for public health. The natural product geniposide has neuroprotective effects on depression, but the underlying mechanism behind these effects had remained undefined. The present study was designed to investigate the role of microRNAs (miRs) in this mechanism. It studied mice with depression-like behavior established by exposure to chronic unpredictable mild stress (CUMS) for 2 months. The CUMS mice were intragastrically fed with geniposide at a dose of 10 ml/kg daily for two consecutive weeks. We monitored the depression-like behaviors of the CUMS mice by the forced swimming test (FST) and tail suspension test (TST). Then, we measured the cerebral expression of miR-298-5p and NADPH oxidase 1 (Nox1) mRNA in the CUMS mice by the RT-qPCR. The targeting relationship between miR-298-5p and Nox1 was evaluated by dual-luciferase reporter gene assay. The concentrations of adenosine triphosphate (ATP) and reactive oxygen species (ROS) were determined by the CellTiter-Glo® and flow cytometry, respectively. The mitochondrial membrane potential (MMP) was detected using JC-1 staining. Moreover, the expression of inflammatory cytokines (TNF-α, IL-1β, IL-6, and TGF-β) was determined by ELISA, RT-qPCR, and western blot analysis. We found that miR-298-5p was poorly-expressed while Nox1 was highly-expressed in the brain tissues of the CUMS-induced mice. Intriguingly, Geniposide treatment reversed the behavioral abnormalities of CUMS mice, including shortened immobility time. Geniposide inhibited the Nox1 expression by increasing miR-298-5p levels. There were increased ATP content and MMP and reduced contents of ROS and inflammatory cytokines in the CUMS mice receiving geniposide treatment. Hence, this study revealed an antidepressant effect of geniposide on CUMS-induced depression-like behavior in mice by down-regulating the miR-298-5p-targeted Nox1. This highlights a novel candidate target for the treatment of depression.

Fluoxetine ameliorates depressive symptoms by regulating lncRNA expression in the mouse hippocampus.

Depression is a prevalent mental disorder that is associated with aging and contributes to increased mortality and morbidity. The overall prevalence of geriatric depression with clinically significant symptoms is currently on the rise. Recent studies have demonstrated that altered expressions of long non-coding RNAs (lncRNAs) in the brain affect neurodevelopment and manifest modulating functions during the depression. However, most lncRNAs have not yet been studied. Herein, we analyzed the transcriptome of dysregulated lncRNAs to reveal their expressions in a mouse model exhibiting depressive-like behaviors, as well as their corresponding response following antidepressant fluoxetine treatment. A chronic unpredictable mild stress (CUMS) mouse model was applied. A six-week fluoxetine intervention in CUMS-induced mice attenuated depressive-like behaviors. In addition, differential expression analysis of lncRNAs was performed following RNA-sequencing. A total of 282 lncRNAs (134 up-regulated and 148 down-regulated) were differentially expressed in CUMS-induced mice relative to non-stressed counterparts (P<0.05). Moreover, 370 differentially expressed lncRNAs were identified in CUMS-induced mice after fluoxetine intervention. Gene Ontology (GO) analyses showed an association between significantly dysregulated lncRNAs and protein binding, oxygen binding, and transport activity, while the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that these dysregulated lncRNAs might be involved in inflammatory response pathways. Fluoxetine effectively ameliorated the symptoms of depression in CUMS-induced mice by regulating the expression of lncRNAs in the hippocampus. The findings herein provide valuable insights into the potential mechanism underlying depression in elderly people.

The effect of geniposide on chronic unpredictable mild stress-induced depressive mice through BTK/TLR4/NF-κB and BDNF/TrkB signaling pathways.

The purpose of this study was to estimate the pharmacological effect of geniposide (GEN) on depression, caused by chronic unpredictable mild stress (CUMS), and explore its potential mechanism. During the 6 week CUMS procedure, the mice were treated with GEN (10, 40 mg/kg) by gavage once daily for 3 weeks. As a result, the GEN treatment remarkably improved the behavioral manifestations and suppressed the generations of inflammatory cytokines both in vivo and in vitro. The MDA level was significantly increased, while the activities of SOD, GSH-PX were decreased in CUMS-challenged mice and corticosterone-stimulated PC12 cells. GEN administration significantly inhibited those changes. Moreover, GEN treatment could downregulate the expressions of p-BTK, TLR4, MyD88, p-NF-κB proteins, and upregulate BDNF, p-TrkB generations in CUMS-induced mice. Moreover, GEN administration inhibited the protein levels of p-BTK, TLR4, MyD88, p-NF-κB in corticosterone-induced PC12 cell. In summary, the results suggested that GEN exerted a therapeutic effect on CUMS-induced depressive mice possibly through the regulation of BTK/TLR4/NF-κB and BDNF/TrkB signaling pathways.

Protection of hUC-MSCs against neuronal complement C3a receptor-mediated NLRP3 activation in CUMS-induced mice

BackgroundHyperactivation of complement C3 and inflammation-related activation of NLR family pyrin domain containing 3 (NLRP3) inflammasome are implicated in the etiology of stress-related disorders. Studies have shown that human umbilical cord mesenchymal stromal cells (hUC-MSCs) have immunomodulatory and anti-inflammatory effects; however, the mechanism remains unclear. MethodshUC-MSCs were administered to chronic unpredictable mild stress (CUMS) model mice once a week for four weeks. After the administration of hUC-MSCs, several parameters were assessed, including behavioral performance, synapse-related proteins, complement C3 receptors (C3aR) in neurons, and the NLRP3 inflammasome. Then, CUMS mice were injected with SB290157, a complement C3aR antagonist, and the behavioral index and NLRP3 inflammasome activation were tested. ResultsThe open-field and forced swimming behavioral tests showed an improvement in depression-like behaviors in the CUMS-exposed mice after the administration of hUC-MSCs. Treatment with hUC-MSCs significantly decreased the neuronal C3aR levels and alleviated synaptic damage. Furthermore, the levels of the NLRP3 inflammasome and inflammatory factors were reduced after hUC-MSC administration. In particular, treatment with a C3aR antagonist also decreased NLRP3 inflammasome expression and inflammation, which was consistent with the observed improvements after hUC-MSC treatment. ConclusionhUC-MSCs can attenuate NLRP3 activation in CUMS-induced mice, which may be correlated with C3aR in neurons.

Geniposide Alleviates Oxidative Stress of Mice With Depression-Like Behaviors by Upregulating Six3os1.

Depression is a major cause of disease burden and severely impairs well-being of patients around the globe. Geniposide (GP) has been revealed to play a significant role in depression treatment. Of note, RNA sequencing of this study identified highly expressed long non-coding RNA Six3os1 in response to GP treatment. Thus, we aim to explore how GP affected chronic unpredictable mild stress (CUMS)-induced depression-like behaviors in mice in vivo and in vitro and the downstream molecular mechanism related to Six3os1. The relationship of Six3os1, miR-511-3p and Fezf1 was evaluated by dual-luciferase reporter gene assay, RIP assay, and RNA pulling down assay. Ectopic expression and knockdown experiments were developed in CUMS-induced mice and neurons with or without GP treatment. In vitro experiments and behavioral tests were conducted to examine alteration of CUMS-triggered oxidative stress following different interferences. The experimental data validated that GP treatment resulted in high expression of Six3os1 and Fezf1 and poor expression of miR-511-3p in CUMS-induced neurons. Six3os1 activated the AKT signaling pathway by upregulating miR-511-3p-targeted Fezf1. Either GP treatment or overexpression of Six3os1 or Fezf1 alleviated depression-like behaviors of CUMS-induced mice. GP treatment, miR-511-3p inhibition or overexpression of Six3os1 or Fezf1 not only reduced oxidative stress in CUMS-induced mice and neurons, but also reduced CUMS-induced neuronal apoptosis. Collectively, GP treatment-mediated Six3os1 upregulation ameliorated oxidative stress of mice with depression-like behaviors via the miR-511-3p/Fezf1/AKT axis.

Antidepressant effects of a polysaccharide from okra (Abelmoschus esculentus (L) Moench) by anti-inflammation and rebalancing the gut microbiota

The present study aimed to evaluate the antidepressant-like effect of a polysaccharide (OP), which is isolated from okra (Abelmoschus esculentus (L) Moench), in CUMS-induced mice and its possible mechanisms. OPT, FST and TST were employed to examine the anxiety and depressive behavior in CUMS-induced mice and fecal microbiota transplantation (FMT) CUMS-induced mice, while proinflammatory cytokines, TLR4/NF-κB pathway and MAPKs signaling were detected in both CUMS-induced mice and LPS-induced BV2 cells. The results showed that anxiety- and depressive-like behaviors, gut microbiota dysbiosis and changes of SCFAs, and activation of inflammatory reactions in the colon, serum, and hippocampus of CUMS-induced mice, as well as activation of inflammatory reactions in BV2 cells, could be alleviated by the treatment of OP. The mice that were colonized by OP microbiota showed improved anxiety and depressive behaviors and lower inflammatory response. Furthermore, OP inhibited the expression of TLR4, the nuclear translocation of NF-κB and high levels of proinflammatory cytokines, and enhanced the MAPKs signaling, these effects of OP also observed in LPS-induced BV2 cells. Above all, suggested that the potential mechanism of the antidepressant-like effects of OP was closely correlated with the bidirectional communication of microbiota-gut-brain axis via regulation of inflammation response.

Xiaoyaosan exerts antidepressant-like effects by regulating the functions of astrocytes and EAATs in the prefrontal cortex of mice

BackgroundMounting evidence indicates that the cerebral cortex is an important physiological system of emotional activity, and its dysfunction may be the main cause of stress. Glutamate is the primary excitatory neurotransmitter in the central nervous system (CNS), which initiates rapid signal transmission in the synapse before its reuptake into the surrounding glia, specifically astrocytes (ASTs). The astrocytic excitatory amino acid transporters 1 (EAAT1) and 2 (EAAT2) are the major transporters that take up synaptic glutamate to maintain optimal extracellular glutamic levels, thus preventing accumulation in the synaptic cleft and ensuing excitotoxicity. Growing evidence has shown that excitotoxicity is associated with depression. Therefore, we hypothesized that the underlying antidepressant-like mechanism of Xiaoyaosan (XYS), a Chinese herbal formula, may be related to the regulation of astrocytic EAATs. Therefore, we studied the antidepressant mechanism of XYS on the basis of EAAT dysfunction in ASTs.MethodsEighty adult C57BL/6 J mice were randomly divided into 4 groups: a control group, a chronic unpredictable mild stress (CUMS) group, a Xiaoyaosan (XYS) treatment group and a fluoxetine hydrochloride (Flu) treatment group. Except for the control group, mice in the other groups all received chronic unpredictable mild stress for 21 days. Mice in the control and CUMS groups received gavage administration with 0.5 mL of normal saline (NS) for 21 days, and mice in the XYS and Flu treatment groups were administered dosages of 0.25 g/kg/d and 2.6 mg/kg/d by gavage. The effects of XYS on the depressive-like behavioral tests, including the open field test (OFT), forced swimming test (FST) and sucrose preference test (SPT), were examined. The glutamate (Glu) concentrations of the prefrontal cortex (PFC) were detected with colorimetry. The morphology of neurons in the PFC was observed by Nissl staining. The expression of glial fibrillary acidic protein (GFAP), NeuN, EAAT1 and EAAT2 proteins in the PFC of mice was detected by using Western blotting and immunohistochemistry. Quantitative real-time PCR (qPCR) was used to detect the expression of the GFAP, NeuN, EAAT1 and EAAT2 genes in the PFC of mice.ResultsThe results of behavioral tests showed that CUMS-induced mice exhibited depressive-like behavior, which could be improved in some tests with XYS and Flu treatment. Immunohistochemistry and Western blot analysis showed that the protein levels of GFAP, NeuN, EAAT1 and EAAT2 in the PFC of CUMS mice were significantly lower than those in the control group, and these changes could be reversed by XYS and Flu. The results of qPCR analysis showed that the expression of GFAP, NeuN, EAAT1 and EAAT2 mRNAs in the PFC of CUMS mice was not significantly changed, with the exception of EAAT2, compared with that of the control group, while the expression of the above mRNAs was significantly higher in the XYS and Flu groups than that in the CUMS group.ConclusionXYS may exert antidepressant-like effects by improving the functions of AST and EAATs and attenuating glutamate-induced neuronal damage in the frontal cortex.

Andrographolide triggers autophagy-mediated inflammation inhibition and attenuates chronic unpredictable mild stress (CUMS)-induced depressive-like behavior in mice

Depression is one of the most common psychiatric disorders in the world. Andrographolide is a natural product that displays evident anti-inflammatory activities. The purpose of the present study was to explore the antidepressant potential of andrographolide in chronic unpredictable mild stress (CUMS)-induced depressive-like behavior in mice. Performance in behavioral tests such as the forced swim test, sucrose preference test, tail suspension test and Y-maze was improved following andrographolide administration. The pro-inflammatory mediator NO and cytokines IL-1β, IL-6 as well as TNF-α were measured in the prefrontal cortex using a reagent kit, ELISA and real-time PCR. NF-κB signaling, NLRP3 inflammasome assembly and autophagy process were examined in the prefrontal cortex using western blotting. It was observed that 5 mg/kg andrographolide treatment obviously improved depressive-like behavior. In addition, 5 mg/kg andrographolide treatment also decreased the expression of pro-inflammatory mediators and cytokines (NO, COX-2, iNOS, IL-1β, IL-6 and TNF-α), NF-κB signaling (p-p65, p-IκBα) and NLRP3 inflammasome assembly (NLRP3, ASC and caspase-1) in the prefrontal cortex. Moreover, autophagy levels increased after andrographolide treatment. Finally, the antidepressant and anti-inflammatory effects of andrographolide were compromised by the application of chloroquine (CQ), which suggested that andrographolide-induced autophagy was mainly affected by the initiation rather than the blocking of autophagic flux. In conclusion, these results suggest that andrographolide produces antidepressant-like and anti-inflammatory effects in CUMS-induced mice which maybe mediated by the upregulation of autophagy.

Identification of key genes, pathways, and miRNA/mRNA regulatory networks of CUMS-induced depression in nucleus accumbens by integrated bioinformatics analysis.

IntroductionMajor depressive disorder (MDD) is a recurrent, devastating mental disorder, which affects >350 million people worldwide, and exerts substantial public health and financial costs to society. Thus, there is a significant need to discover innovative therapeutics to treat depression efficiently. Stress-induced dysfunction in the subtype of neuronal cells and the change of synaptic plasticity and structural plasticity of nucleus accumbens (NAc) are implicated in depression symptomology. However, the molecular and epigenetic mechanisms and stresses to the NAc pathological changes in depression remain elusive.Materials and methodsIn this study, treatment group mice were treated continually with the chronic unpredictable mild stress (CUMS) until expression of depression-like behaviors were found. Depression was confirmed with sucrose preference, novelty-suppressed feeding, forced swimming, and tail suspension tests. We applied high-throughput RNA sequencing to assess microRNA expression and transcriptional profiles in the NAc tissue from depression-like behaviors mice and control mice. The regulatory network of miRNAs/mRNAs was constructed based on the high-throughput RNA sequence and bioinformatics software predictions.ResultsA total of 17 miRNAs and 10 mRNAs were significantly upregulated in the NAc of CUMS-induced mice with depression-like behaviors, and 12 miRNAs and 29 mRNAs were downregulated. A series of bioinformatics analyses showed that these altered miRNAs predicted target mRNA and differentially expressed mRNAs were significantly enriched in the MAPK signaling pathway, GABAergic synapse, dopaminergic synapse, cytokine–cytokine receptor interaction, axon guidance, regulation of autophagy, and so on. Furthermore, dual luciferase report assay and qRT-PCR results validated the miRNA/mRNA regulatory network.ConclusionThe deteriorations of GABAergic synapses, dopaminergic synapses, neurotransmitter synthesis, as well as autophagy-associated apoptotic pathway are associated with the molecular pathological mechanism of CUMS-induced depression.

Evaluating the Anti-depression Effect of Xiaoyaosan on Chronically-stressed Mice.

In addition to the standardized use of antidepressant medications and psychotherapy, the usage of traditional Chinese medicine has lead to an overall improvement of patients with major depressive disorder (MDD). Therefore, the purpose of this study was to establish the mouse depressive model, observe the behavior changes associated with chronic unpredictable mild stress (CUMS), and then evaluate the anti-depression effect of Xiaoyaosan. Mice were randomly divided into four groups: a control group, a model group, a treatment group with Xiaoyaosan, and a treatment group with fluoxetine. All mice were individually kept in cages, and depression was induced in the mice by exposing them to several designed manipulations of CUMS for 21 days, as described in the protocol. Mice in the control group and model group received 0.5 mL of distilled water, while mice in the treatment groups received either Xiaoyaosan (0.25 g/kg/day) or fluoxetine (2.6 mg/kg/day). The drugs used in the study were given intragastrically daily during the entire three weeks. To estimate the depressive-like behaviors, a series of parameters including the coat state, body weight, open field test score, and sucrose preference test score were recorded. Data analysis showed that behaviors of model mice were significantly changed compared to behaviors of mice in the control group, which were improved by the treatment of Xiaoyaosan and fluoxetine. The current findings demonstrated the anti-depression effects of Xiaoyaosan on the behaviors of CUMS-induced mice and revealed that compounds from the Xiaoyaosan prescription may be worthwhile for treating depression, considering their beneficial effects on depressive-like behaviors.

(+)-Sesamin attenuates chronic unpredictable mild stress-induced depressive-like behaviors and memory deficits via suppression of neuroinflammation

Depression is a mood disorder that is related to neuroinflammation and cognition loss. This study is aimed to determine the potential antidepressant effects of (+)-sesamin, a lignan component of sesame, in a mild stress-induced depression mouse model. CD-1 mice were treated with chronic unpredictable mild stress (CUMS) process and orally administrated with sesamin (50 mg/kg/d) for 6 weeks. Behavioral tests including forced swimming test, tail suspension test, open field test, and elevated plus maze test demonstrated that sesamin treatment inhibited CUMS-induced mice depressant-like behaviors and anxiety, without changing immobility. It was found that sesamin prevented stress-induced decease levels of 5-HT and NE in striatum and serum. Cognitive deficits were assessed using Y-maze and Morris water maze test. Sesamin treatment also prevented stressed-induced memory impairments and neuronal damages. Consistently, sesamin also enhanced synapse ultrastructure and improved expressions of PSD-95 in stressed mice hippocampus with improving neurotrophic factors expression including BDNF and NT3. Moreover, sesamin treatment significantly prevented CUMS-induced neuroinflammation by inhibiting over-activation of microglia and expressions of inflammatory mediators including iNOS, COX-2, TNF-α and IL-1β in stressed mice hippocampus and cortex. These results illustrated that sesamin markedly improved CUMS-induced depression and memory loss via inhibiting neuroinflammation, which indicate that as food component, sesamin might be also a novel potential therapeutic for depression.

Xiaoyaosan Improves Depressive-Like Behaviors in Mice through Regulating Apelin-APJ System in Hypothalamus.

Background: The apelin-APJ system has been considered to play a crucial role in HPA axis function, and how the traditional Chinese compound prescription Xiaoyaosan regulates the apelin-APJ system as a supplement to treat depressive disorders. Objective: To investigate the depression-like behaviors and expression of apelin and APJ in hypothalamus of chronic unpredictable mild stress (CUMS) mice and study whether these changes related to the regulation of Xiaoyaosan. Methods: 60 adult C57BL/6J mice were randomly divided into four groups, including control group, CUMS group, Xiaoyaosan treatment group and fluoxetine treatment group. Mice in the control group and CUMS group received 0.5 mL physiological saline once a day by intragastric administration. Mice in two treatment groups received Xiaoyaosan (0.25 g/kg/d) and fluoxetine (2.6 mg/kg/d), respectively. After 21 days of modeling with CUMS, the expression of apelin and APJ in hypothalamus were measured by real-time fluorescence quantitative PCR, western blot and immunohistochemical staining. The physical condition, body weight, food intake and behavior tests such as open field test, sucrose preference test and force swimming test were measured to evaluate depressive-like behaviors. Results: In this study, significant behavioral changes were found in CUMS-induced mice, meanwhile the expressions of apelin and APJ in the hypothalamus were changed after modeling. The body weight, food-intake and depressive-like behaviors in CUMS-induced mice could be improved by Xiaoyaosan treatment which is similar with the efficacy of fluoxetine, while the expressions of apelin and APJ in hypothalamus were modified by Xiaoyaosan. Conclusions: The data suggest that apelin-APJ system changes in the hypothalamus may be a target of depressive disorders, and the beneficial effects of Chinese compound prescription Xiaoyaosan on depressive-like behaviors may be mediated by the apelin-APJ system.