Glutamate Transporter 1-mediated Antidepressant-like Effect in a Rat Model of Chronic Unpredictable Stress

In recent years, more attention has been paid to the role of the glutamate transporter 1 （GLT-1, EAAT2） in major depressive disorder （MDD）. However, experimental data on brain GLT-1 levels are, to some extent, inconsistent in human postmortem and animal studies, These discrepancies imply that the role of GLT-1 in the pathophysiology of MDD and the action of antidepressants remain obscure. This work was designed to study the impact of chronic unpredictable stress （CUS） for 2 ses- sions per day for 35 days and four weeks of fluoxetine （FLX） on depressive-like behaviors in rats, as well as the concomitant expression of the GLT-1 protein in the hippocampus. Behavioral changes were assessed by the sucrose preference and open field tests. GLT-1 levels were detected by immunohisto- chemistry and Western blot analysis. Our study demonstrated that the animals exposed to CUS showed depressive-like behaviors and exhibited a significant decrease in GLT-1 expression in the hippocampus. Chronic FLX treatment reversed the behavioral deficits and the CUS-induced decrease in GLT-1 levels. Taken together, our results support the reduction of GLT-1 in human postmortem studies in MDD and suggest that GLT-1 may be involved in the antidepressant activity of FLX. Our studies further support the notion that GLT-1 is an attractive candidate molecule associated with the fundamental processes of MDD and may be a potential, and novel pharmacological target for the treatment of MDD.

Shuganheweitang Ameliorates Chronic Unpredictable Mild Stress-Induced Depression-Like Behaviors in Rats through the PI3K/AKT/mTOR Pathway: Involvement of Amino Acids, Glycerophospholipids, and Energy Metabolism

Background: Depression is a typical psychosomatic disease. Shuganheweitang (SGHWT) is a clinical formula that effectively treats depression. However, the potential mechanism used by SGHWT to ameliorate depression-like behaviors is still unclear. This study investigated the effects of SGHWT on metabolic change in the liver and hypothalamus with signaling pathways involved in chronic unpredictable mild stress (CUMS)-induced depression in rats to explore the mechanism of the anti-depressive effect. Methods: A total of 52 rats were used to create a model of depression by CUMS combined with solitary rearing for 6 weeks. Open field test (OFT), sucrose preference test (SPT), forced swim test (FST), and body weight (BW) were performed to analyze the pharmacodynamic effects of SGHWT. H&E staining, Nissl staining, immunofluorescence, immunohistochemistry, and western blot were used to evaluate the mechanism of action. Untargeted metabolomics techniques by ultra-performance liquid chromatography-quantitative time-of-flight tandem mass spectrometry (UPLC-Q-TOF-MS/MS) were used to analyze all the metabolic differences in the liver and hypothalamus. Results: SGHWT improved CUMS-induced depression-like behaviors in vivo. SGHWT reduced hepatic c-Fos protein expression and increased hypothalamic c-Fos protein expression. Moreover, p-PI3K, p-AKT473, p-AKT308, and p-mTOR protein expressions were significantly downregulated in the liver and hypothalamus of CUMS rats. Notably, these alterations were reversed by the SGHWT administration. Furthermore, the metabolomic analysis identified 15 and 5 key differential SPT-associated metabolites in the liver and hypothalamus, respectively. Conclusion: This study suggests that SGHWT ameliorates chronic unpredictable mild stress-induced depression-like behaviors, by the involvement of amino acids, glycerophospholipids, energy metabolism, and the PI3K/AKT/mTOR pathway. Highlights: 1) Shuganheweitang was derived from the TCM herbal formula Sinisan. 2) SGHWT treatment reverses depression-like behaviors in CUMS-induced rats. 3) The mechanism of SGHWT on depression by the liver and hypothalamus metabolomics. 4) SGHWT regulates amino acids, glycerophospholipids, and energy metabolism. 5) SGHWT exerts antidepressant effects through the PI3K/AKT/mTOR pathway.

Antidepressant-like effects of BCEF0083 in the chronic unpredictable stress models in mice

Background Up to now there have been no satisfactory drugs to treat psychiatric disorders, and now bioactive compound from entomagenous fungi (BCEF0083) is a new type of bioactive compound from entomopathogenic fungi. Our previous investigations have shown that BCEF has an inhibition effect on monoamine oxidase. So, BCEF may be a latent antidepressant. This study aimed at observing the antidepressant effects and its mechanism of BCEF in the chronic unpredictable stress models in mice. Methods The antidepressant effects of BCEF were examined on the chronic unpredictable stress models in mice. Sixty mice were randomly divided to six groups. Animals were housed and isolated except saline group. Mice were exposed to different stressors per day randomly from day 1 to day 21. Body weight were weighed on day 1,day 10 and on day 21 during the 21-day stress procedure. Awarding response was detected by using method of calculating the 24-hour consumption of saccharum water. Step through test was used to evaluate the behavioral response. AVP contents in plasma were also detected by using radioimmunoassays. Results Chronic unpredictable stress resulted in a significant decrease of the body weight and could apparently cause escape behavior disturbance and gradual reduction of sensitivity to reward in animal models. Drug treatment (BCEF 25, 50, 100 mg/kg) could significantly ameliorate the decreased body weight and effectively reverse the escape behavior disturbance. The gradual reduction of sensitivity to reward, the anhedonic state, was also effectively reversed by BCEF. BCEF (50, 100 mg/kg) could also effectively restore the AVP content in the plasma.Conclusions This evidence suggests that BCEF can effectively inhibit the depression behavior and show strong antidepressant effect. BCEF can effectively restore the plasma AVP release and this may be an important mechanism of its antidepressant effect.

Jobelyn^(■), a Sorghum-Based Nutritional Supplement Attenuates Unpredictable Chronic Mild Stress-Induced Memory Deficits in Mice

The ability of an organism to adapt to aversive stressful situations or life challenging circumstances is very crucial to its state of health and survival. However, breakdown in adaptation due to persistent uncontrollable stress, leads to impairment of bodily functions and onset of a variety of pathological disorders especially memory decline. This study was designed to evaluate the effect of Jobelyn&reg;(JB), a potent antioxidant sorghum-based food supplement on unpredictable chronic mild stress (UCMS)-induced memory impairment in mice. Male Swiss mice were given JB (5 - 50 mg/kg, p.o) 30 min prior to exposure to UCMS for 14 consecutive days before testing for memory. Thereafter, the serum corticosterone level was estimated by using ELISA kits. The levels of malondialdehyde (MDA) and glutathione (GSH) as well as acetylcholinesterase activity were estimated in the brain homogenate using spectrophotometer. Histology of the brain tissues and estimation of the populations of viable neurons in the hippocampal region were done after staining with hematoxyline and eosin. Our results showed that JB reversed memory impairment and suppressed corticosterone concentrations induced by UCMS. Moreover, JB reduced oxidative stress in the brain of UCMS-mice as shown by decreased MDA levels and elevated GSH concentrations. It also decreased brain acetylcholinesterase activity when compared with chronic stress group (p < 0.05). Furthermore, JB (5 - 10 mg/kg, p.o) offered significant protection against UCMS-induced degeneration and death of neuronal cells of the cornu ammonis 3 (CA3) of the hippocampal region of the brain indicating neuroprotection. Taken together, these findings suggest that JB attenuates memory deficits induced by UCMS in mice and may be useful therapeutically for stress-related cognitive dysfunctions. The reduction in the levels of serum corticosterone, antioxidation, neuroprotection and inhibition of cholinesterase enzyme might be contributing significantly to the positive effect of JB on memory in mice exposed to unpredictable chronic mild stress.

L-tyrosine improves neuroendocrine function in a mouse model of chronic stress

Adult BALB/c mice, individually housed, were stimulated with nine different stressors, arranged randomly, for 4 continuous weeks to generate an animal model of chronic stress. In chronically stressed mice, spontaneous locomotor activity was significantly decreased, escape latency in the Morris water maze test was prolonged, serum levels of total thyrotropin and total triiodothyronine were significantly decreased, and dopamine and norepinephrine content in the pallium, hippocampus and hypothalamus were significantly reduced. All of these changes were suppressed, to varying degrees, by L-tyrosine supplementation. These findings indicate that the neuroendocrine network plays an important role in chronic stress, and that L-tyrosine supplementation has therapeutic effects.

Vitamin D_(3) attenuates anxiety-like behavior in long-term ovariectomized rats with unpredictable mild stress

The impact of various vitamin D3(VD3)doses(1.0,2.5,or 5 mg/kg,s.c.)in mitigating the negative consequences of chronic unpredictable mild stress(CUMS)was investigated.Adult female rats with long-term estrogen deficiency were assessed using the sucrose preference test(SPT),the elevated plus-maze(EPM),the light/dark test(LDT),and the open-field test(OFT)to measure anhedonia-like and anxiety-like behavior.The corticosterone(CS)and adrenocorticotrophic hormone(ACTH)concentrations in blood serum and the brain-derived neurotrophic factor(BDNF)expression in the hippocampus of long-term ovariectomized(OVX)rats were measured by ELISA kits and/or western blotting.Treatment with VD3(5.0 mg/kg),similarly to fluoxetine(10.0 mg/kg),significantly reduced the anhedonia profile in the SPT and anxiety-like behavior in the EPM and LDT,and CS and ACTH levels in blood serum.It also elevated BDNF levels in the hippocampus of long-term OVX/CUMS compared to OVX/CUMS/solvent rats.Thus,these findings suggest that VD3(5.0 mg/kg)administration might attenuate the anxiety-like profile in long-term OVX adult rats subjected to the CUMS.This might occur via activation of the BDNF signaling pathway in the hippocampus and via restoration of CS and ACTH levels in blood serum.

Integration of animal behaviors under stresses with different time courses

We used animal models of ＂forced swim stress＂ and ＂chronic unpredictable stress＂, and tried to reveal whether a passive coping style of high flotation behavior in forced swim stress predicts an- hedonia behavior after chronic unpredictable stress, and whether the dopamine system regulates floating and anhedonia behaviors. Our results confirmed that depression-prone rats use ＂floating behavior＂ as a coping strategy in forced swim stress and more readily suffer from anhedonia during chronic unpredictable stress. Intraperitoneal injection or nucleus accumbens microin- jection of the dopamine 2/3 receptor subtype agonist ropinirole reduced floating behaviors in depression-prone animals, but increased sucrose preference in rats showing anhedonia. These data indicate that floating behavior is a defensive mode that is preferred by susceptible individ- uals under conditions of acute stress. Simultaneously, these animals more readily experienced anhedonia under long-term stress; that is, they were more readily affected by depression. Our results suggest that dopamine 2/3 receptor subtypes in the nucleus accumbens play an important role in floating behaviors and anhedonia.

Splenectomy does not affect mouse behaviors

The spleen is critical for immunity.It is the largest immune organ and immune center in the peripheral system.While the relationship between behavior and immunity has been demonstrated in physiology and diseases,the role of the spleen in behavior is not clear.To investigate the effects of the spleen on behaviors,we performed a refined splenectomy procedure on C57BL/6J mice and performed an open field test,circadian rhythm test,elevated plus maze,sucrose preference test,and Barnes maze test.Splenectomy did not induce changes in general locomotion,circadian rhythms,learning and memory,or depression/anxiety-related behaviors.To further investigate the effects of spleen on stress susceptibility,we established mouse models of depression through chronic unpredictable mild stress.The behavioral performances of mice subjected to splenectomy showed no differences from control animals.These findings suggest that splenectomy does not cause changes in baseline behavioral performance in mice.

Antidepressant-Like Effects of Chaihu Shugan Powder(柴胡疏肝散)on Rats Exposed to Chronic Unpredictable Mild Stress through Inhibition of Endoplasmic Reticulum Stress-Induced Apoptosis

Objective:To investigate the antidepressant-like effects of Chaihu Shugan Powder(CSP,柴胡疏肝散)and to explore its underlying mechanisms.Methods:Thirty-two Sprague-Dawley rats were randomly divided into control(CON),chronic unpredictable mild stress(CUMS),fluoxetine(FLU),and CSP groups,8 rats in each group.All of the rats except for those in the control group were subjected to 3 consecutive weeks of CUMS to establish the depression model.The open field test(OFT),forced swimming test(FST),and sucrose preference test were used to assess the anti-anxiety and antidepressant effects of CSP.Terminal deoxynucleotidyl transferase(Td T)d UTP nick-end labeling was used to determine the apoptosis rate in the hippocampal tissues.The m RNA and protein levels of glucose-regulated protein(GRP)78,spliced X-box-binding protein(XBP)-1,CCAAT/enhancerbinding protein homologous protein(CHOP),caspase-12,and c-Jun N-terminal kinase(JNK)in the hippocampus of rats were evaluated by real-time PCR and Western blot analysis,respectively.Results:Administration of CSP alleviated anxiety and depression-like behavior in CUMS rats,as revealed by enhanced time and distance in the center of the OFT(P<0.05),an increased preference for sucrose,and longer swimming time and shorter immobility time during the FST(all P<0.05).In addition,CSP treatment significantly reduced the rate of apoptosis in rat hippocampal neurons(P<0.05).The m RNA and protein expression levels of GRP78,spliced XBP-1,and CHOP were down-regulated along with the expression of caspase-12 and cleaved caspase-12 proteins(all P<0.05),whereas total and phosphorylated JNK1 protein levels did not differ significantly between control and CSP-treated rats.Conclusion:CSP can improve depression-like behavior in rats exposed to CUMS,possibly by suppressing CHOP and caspase-12 mediated apoptosis in the rat hippocampus.

Fluoxetine Ameliorates the Aggravation of UC Symptoms in C57BL/6 Mice Induced by CUMS

Objective Patients with chronic ulcerative colitis(UC)often have mental symptoms such as depression and anxiety,and stress can lead to gastrointestinal diseases.However,the correlation between mental stress and UC is unclear.In this paper,chronic unpredictable mild stress(CUMS)was utilized to evaluate the involvement of mental factors in the pathogenesis of UC.Methods The CUMS model was used to evaluate the direct/indirect involvement of mental factors in the pathogenesis of UC.The behavior was evaluated by the open field,forced swimming,and tail suspension tests.Body weight,the disease activity index(DAI)score,colon length,and HE staining of colon tissue were used to evaluate the action of CUMS and fluoxetine.Results The results showed that weight loss and the DAI score increased in CUMS mice,but they had no meaningful effect on colon length and morphological structure of colon tissue.However,CUMS aggravated dextran sulfate sodium(DSS)-induced colon length shortening and colon morphological structure damage.Fluoxetine significantly improved the DAI score,shortened colon length,and damaged morphology and structure of the colons induced by CUMS combined with DSS in mice.Fluoxetine also decreased the level of IL-6 in the serum and the TNF-αand IFN-γlevels of colon tissue.Fluoxetine simultaneously improved behavioral abnormalities induced by CUMS combined with DSS in mice.Conclusion CUMS aggravated the UC symptoms induced by DSS,and fluoxetine could improve the UC symptoms due to its improvement in the inflammatory level and behavioral abnormalities.

Effects of CUMS combined with CRS on hippocampal glial cells and synaptic plasticity in depressed mice

Objective:To explore the effects of CUMS combined with CRS on mouse hippocampal glial cells and synaptic plasticity-related proteins. Methods: Forty mice were randomly divided into normal group (n=20) and model group (n=20). The model group used CUMS combined with CRS to prepare a mouse model of depression for 7 weeks. The behavioral evaluation of the mice at 3 weeks and 7 weeks after modeling was performed by sugar water preference test, open field test and tail suspension test. After the experiment, the samples were collected, and the content of TNF-a in the hippocampus of mice was detected by enzyme-linked immunosorbent assay. Immunohistochemical method was used to detect the Iba-1 and GFAP MOD values of mouse hippocampal CA1 area, CA3 area and DG area. Western blot was used to detect the protein expression of Iba-1, GFAP, SYN1 and PSD-95 in the hippocampus. fluorescence quantitative PCR method was used to detect the expression of SYN1, PSD-95 mRNA in hippocampus. Results: At the 3rd week after modeling, the body weight, sugar water preference rate, total distance moved, number of standing uprights, and stay time in the central area of the mice in the model group were all lower than those in the normal group (P<0.05), and the tail suspension immobility time was longer than that in the normal group (P<0.01). After 7 weeks of modeling, the body weight, sugar water preference rate, total distance moved, number of erection times, central area residence time, and average movement speed of the mice in the model group were lower than those in the normal group (P< 0.05), the tail suspension immobility time was longer than that in the normal group (P<0.01). The contents of TNF-a in the hippocampus were higher than those in the normal group (P<0.05). The GFAP MOD value and the relative expression of GFAP protein in hippocampal CA1, CA3 and DG regions were significantly lower than those in the normal group (P<0.05). The Iba-1 MOD value and the relative expression of Iba-1 protein in hippocampal CA1, CA3 and DG regions were significantly higher than those in the normal group (P<0.05). The relative expression of SYN1 and PSD-95 protein and the relative expression of SYN1 and PSD-95 mRNA in the hippocampus were significantly lower than those in the normal group (P<0.05). Conclusion: After 3 weeks of CUMS and CRS modeling, the depression-like behavior of mice appeared, and the depression of mice was more obvious after 7 weeks of modeling. The depression mouse model made by CUMS combined with CRS method may be related to increased hippocampal inflammation, excessive activation of microglia, decreased number of astrocytes and decreased synaptic plasticity.

Momordica charantia fruit extract with antioxidant capacity improves the expression of tyrosine-phosphorylated proteins in epididymal fluid of chronic stress rats

Objective:Although the protective effects of Momordica charantia L.(MC)extract on chemical-induced testicular damage have been studied,the preventive effects of MC extract on functional proteins in the epididymis under chronic stress have never been reported.This study investigated the protective effects of MC fruit extract on protein secretion,especially tyrosine-phosphorylated proteins,in the epididymis of rats exposed to chronic unpredictable stress(CUS).Methods:Total phenolic compounds(TPC),total flavonoid compounds(TFC)and antioxidant capacities of MC extract were measured.Adult male rats were divided into 4 groups:control group,CUS group,and 2groups of CUS that received different doses of MC extract(40 or 80 mg/kg).In treated groups,rats were given MC daily,followed by induction of CUS(1 stressor was randomly applied from a battery of 9 potential stressors)for 60 consecutive days.Plasma corticosterone and testosterone levels were analyzed after the end of experiment.Expressions of heat-shock protein 70(HSP-70)and tyrosine-phosphorylated proteins present in the fluid of the head and tail of the epididymis were quantified using Western blot.Results:MC extract contained TPC of(19.005±0.270)mg gallic acid equivalents and TFC of(0.306±0.012)mg catechin equivalents per gram,and had 2,2-diphenyl-1-picrylhydrazyl antioxidant capacity of(4.985±0.086)mg trolox equivalents per gram,radical 50%inhibitory concentration of(2.011±0.008)mg/mL and ferric reducing antioxidant power of(23.697±0.819)μmol Fe(Ⅱ)per gram.Testosterone level in the epididymis was significantly increased,while the corticosterone level was significantly improved in groups treated with MC extract,compared to the CUS animals.Particularly,an 80 mg/kg dose of MC extract prevented the impairments of HSP-70 and tyrosine-phosphorylated protein expressions in the luminal fluid of the epididymis of CUS rats.Conclusion:MC fruit extract had antioxidant activities and improved the functional proteins secreted from the head and tail of the epididymis.It is possible to develop the MC fruit extract as a male fertility supplement for enhancing functional sperm maturation in stressed men.

Overexpression of brain-derived neurotrophic factor in the hippocampus protects against post-stroke depression

Post-stroke depression is associated with reduced expression of brain-derived neurotrophic factor （BDNF）. In this study, we evaluated whether BDNF overexpression affects depression-like behavior in a rat model of post-stroke depression. The middle cerebral artery was occluded to produce a model of focal cerebral ischemia. These rats were then subjected to isolation-housing combined with chronic unpredictable mild stress to generate a model of post-stroke depression. A BDNF gene lentiviral vector was injected into the hippocampus. At 7 days after injection, western blot assay and real-time quantitative PCR revealed that BDNF expression in the hippo- campus was increased in depressive rats injected with BDNF lentivirus compared with depressive rats injected with control vector. Furthermore, sucrose solution consumption was higher, and horizontal and vertical movement scores were increased in the open field test in these rats as well. These findings suggest that BDNF overexpression in the hippocampus of post-stroke depressive rats alleviates depression-like behaviors.

Shuganjieyu capsule increases neurotrophic factor expression in a rat model of depression

Shuganjieyu capsule has been approved for clinical treatment by the State Food and Drug Ad-ministration of China since 2008. In the clinic, Shuganjieyu capsule is often used to treat mild to moderate depression. In the rat model of depression established in this study, Shuganjieyu capsule was administered intragastrically daily before stress. Behavioral results conifrmed that depressive symptoms lessened after treatment with high-dose （150 mg/kg） Shuganjieyu capsule. Immunohistochemistry results showed that high-dose Shuganjieyu capsule signiifcantly increased phosphorylation levels of phosphorylation cyclic adenosine monophosphate response element binding protein and brain-derived neurotrophic factor expression in the medial prefrontal cortex and hippocampal CA3 area. Overall, our results suggest that in rats, Shuganjieyu capsule effec-tively reverses depressive-like behaviors by increasing expression levels of neurotrophic factors in the brain.

Exploration of the mechanism by which icariin modulates hippocampal neurogenesis in a rat model of depression

Icariin(ICA) has a significant capacity to protect against depression and hippocampal injury,but it cannot effectively cross the bloodbrain barrier and accumulate in the brain.Therefore,the mechanism by which ICA protects against hippocampal injury in depression remains unclear.In this study,we performed proteomics analysis of cerebrospinal fluid to investigate the mechanism by which ICA prevents dysfunctional hippocampal neurogenesis in depression.A rat model of depression was established through exposure to chronic unpredictable mild stress for 6 weeks,after which 120 mg/kg ICA was administered subcutaneously every day.The results showed that ICA alleviated depressive symptoms,learning and memory dysfunction,dysfunctional neurogenesis,and neuronal loss in the dentate gyrus of rats with depression.Neural stem cells from rat embryonic hippocampi were cultured in media containing 20% cerebrospinal fluid from each group of rats and then treated with 100 μM corticosterone.The addition of cerebrospinal fluid from rats treated with ICA largely prevented the corticosterone-mediated inhibition of neuronal proliferation and differentiation.Fifty-two differentially expressed proteins regulated by chronic unpredictable mild stress and ICA were identified through proteomics analysis of cerebrospinal fluid.These proteins were mainly involved in the ribosome,PI3 K-Akt signaling,and interleukin-17 signaling pathways.Parallel reaction monitoring mass spectrometry showed that Rps4 x,Rps12,Rps14,Rps19,Hsp90 b1,and Hsp90 aa1 were up-regulated by chronic unpredictable mild stress and down-regulated by ICA.In contrast,Htr A1 was down-regulated by chronic unpredictable mild stress and up-regulated by ICA.These findings suggest that ICA can prevent depression and dysfunctional hippocampal neurogenesis through regulating the expression of certain proteins found in the cerebrospinal fluid.The study was approved by the Experimental Animal Ethics Committee of Guangzhou University of Chinese Medicine of China in March 2017.

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 sixweek 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.

Establishment and evaluation of a mouse model of affective disorder combined with atherosclerosis

Objective:To establish a mouse model of affective disorder complicated with atherosclerosis(AS)by high fat feeding and chronic mild unpredictable stimulation(CUMS),and to provide an animal model for the later study of the prevention and treatment of affective disorder complicated with atherosclerosis by traditional Chinese medicine.Methods:10 C57BL/6J mice were used as blank group,and 20 ApoE-/-mice were randomly divided into AS group and AS+CUMS group.After one week of adaptive feeding,except for the blank group,the other two groups were fed with high fat diet.Meanwhile,the AS+CUMS group was given chronic unpredictable mild stress.The model was evaluated after 16 weeks of modeling.During the experiment,the body weight,food intake,excitability,hair color and other general morphology of mice in each group were observed and recorded.Behavioral indexes(Sucrose preference tests and Open Field test)were detected in each group.The levels of 5-hydroxytryptamine(5-HT)and hypothalamic-pituitary-adrenal(HPA)axis hormones,including adrenocorticotropin(ACTH)and corticosterone(CORT),were detected by ELISA.Serum lipid levels,including total cholesterol(TC),triglyceride(TG),low density lipoprotein cholesterol(LDL-C)and high density lipoprotein cholesterol(HDL-C),were detected by ELISA.HE staining was used to observe the pathological condition of aorta.Results:Compared with blank group,the excitability and food intake of AS+CUMS group were significantly decreased.There were no significant differences in sugar water consumption and activity capacity of mice in AS group,while sugar water consumption and activity capacity of mice in AS+CUMS group were significantly decreased(P<0.01).The serum 5-HT levels in AS group and AS+CUMS group were decreased,and the levels of ACTH and CORT in AS+CUMS group were significantly decreased(P<0.01),and the levels of ACTH and CORT in AS+CUMS group were significantly increased(P<0.01).The serum TC,TG and LDL-C levels of mice in AS group and AS+CUMS group were significantly increased(P<0.01),while the HDL-C level of mice in both groups was significantly decreased(P<0.01).HE staining results showed that the area and degree of plaques in the active vascular lumen of AS group and AS+CUMS group were larger and heavier.Conclusion:High fat feeding combined with CUMS was successful in establishing a mouse model of emotional disorder combined with atherosclerosis.

Depression and stress levels increase risk of liver cancer through epigenetic downregulation of hypocretin

Recent studies suggest that Hypocretin (HCRT, Orexin) are involved in stress regulation of depression through the hypothalamic-pituitary-adrenal (HPA) axis. However, the molecular mechanism by which Hypocretin regulate neurobiological responses is unknown. Herein, the effects of chronic stress on the epigenetic modification of HCRT and its association with depression were explored with regard to a potential role in cancer progression. In the study, Sprague Dawley (SD) rats were used to establish an animal model of cancer with depression by administrating n-nitrosodiethylamine (DEN) and chronic unpredictable mild stress (CUMS). RNA-sequencing was used to detect differentially expressed genes in the hippocampus of rats and quantitative real-time polymerase chain reaction (qRT-PCR) was used to validate the results of RNA-sequencing. The status of HCRT promoter methylation was assessed by methylation specific polymerase chain reaction. Behavioral tests showed that rats exposed to CUMS had significant depressive-like behaviors. The number of liver tumors and tumor load in depressed rats exposed to CUMS was higher than in SD rats without CUMS. RNA-sequencing revealed that HCRT was one of the most siginificantly downregulated gene in the hippocampus of SD rats with CUMS compared to non-stressed group, which was validated by qRT-PCR. HCRT mRNA expression was downregulated and the promoter for HCRT was hyper-methylated in those with depression. These results identified a critical role for chronic psychological stressors in tumorigenesis and cancer progression, via epigenetic HCRT downregulation. Such epigenetic downregulation may be the molecular basis for the association of cancer with depression.

Quercetin regulates depression-like behavior in CUMS rat models via TLR4/NF-κB signaling

Background:Depression is becoming increasingly prevalent around the world,imposing a substantial burden on individuals,families,as well as society.Quercetin is known to be highly effective in treating depression.However,additional research is needed to dissect the mechanisms of its anti-depressive effects.Methods:For this study,Sprague-Dawley(SD)rats were randomized into the control,model,quercetin,or fluoxetine group.The latter three groups were exposed to chronic unpredictable mild stress(CUMS)for 42 d.The first two groups received saline solution daily via oral gavage.Meanwhile,the quercetin group was orally administered a quercetin suspension(52.08 mg/kg)every day,while the fluoxetine group was orally administered a fluoxetine solution(2.08 mg/kg).Here,fluoxetine served as the positive control drug to compare the therapeutic effects of quercetin.The experimental period was 6 weeks.Depressive behaviors in rats were assessed through various physiological and behavioral measures.Additionally,pathological changes in hippocampal tissues were examined using Nissl staining.Serum cytokines were detected using an enzymelinked immunosorbent assay(ELISA),and immunohistochemistry was employed to quantify the levels and integral optical density(IOD)values of ionized calcium binding adaptor molecule-1(Iba-1)expression in the brain.Real-time fluorescence quantitative PCR(RT-qPCR)was utilized to evaluate the mRNA levels of inflammatory indicators as well as toll-like receptor 4(TLR4),and nuclear factor-κappa B P65(NF-κB P65)in hippocampus.Western blot(WB)technique was employed to observe the protein levels of TLR4,NF-κB P65,and phospho-NF-κB P65(p-NF-κB P65).Results:After 42 d of exposure to CUMS,rats exhibited a slow increase in body weight,a reduction in food intake,an abnormal preference for sugar water,and aberrant open-field behaviors.Pathological analysis revealed the disintegration,rupture,interruption,and disorganization of hippocampal neuronal cells after CUMS exposure,along with a decrease in Nissl bodies in the CA1 region.This was accompanied by the elevated expression of interleukin-1β(IL-1β),tumor necrosis factor-α(TNF-α),and interleukin-6(IL-6)in the serum and the upregulation of IL-1β,IL-6,and TNF-αmRNA expression in the hippocampus.Increases in Iba-1-positive cells and the IOD values of Iba-1 were detected in hippocampal microglia.Furthermore,TLR4 and NF-κB P65 mRNA and protein levels were upregulated in hippocampal tissues.Quercetin,an antidepressant,could alleviate depression-like symptoms in rats and downregulate inflammatory factors associated with the TLR4/NF-κB signaling pathway in hippocampal microglia,and its therapeutic effect was comparable to fluoxetine.Conclusion:In rat models of CUMS,quercetin may act as an antidepressant by inhibiting inflammation in hippocampal microglia via TLR4/NF-κB signaling pathway.These results offer experimental and theoretical support for applying quercetin in the clinical management of depression.

Pinching Spine:A Potential Treatment for Depression

Objective： To investigate whether pinching spine （PS, i.e.捏脊, a traditional Chinese manipulativetherapy） is beneficial to ameliorating the depressive state （including behavioral deficit, retardative weight gainand decreased sucrose consumption） in a rat model of depression induced by chronic unpredictable stress （CUS）and to explore the candidate mechanism of action. Methods： PS was performed on rats＇ spine once daily for 1week after exposure to CUS. The open-field test, body weight measuring, and sucrose intake test were appliedon different dates： before stress （dO）, at the end of stress （d21） and after PS treatment （d28）, respectively. Thenthe rats＇ hippocampuses were performed genome-wide microarray analysis, and the expression levels of severalgenes were evaluated by real-time polymerase chain reaction （PCR）. Results： Exposure to CUS resulted indecreases of behavioral activity and sucrose consumption, which were reversed significantly after PS treatment.The expression of several genes relevant to energy metabolism, anti-oxidation, and olfactory receptor, etc., weredown-regulated, while the expression of those relevant to hemostasis, immunity-inflammation, and restrictionof activities and ingestion, etc., were up-regulated in hippocampuses of rats exposed to CUS. PS treatmentsignificantly inverted these changes. Furthermore, increase or decrease in gone expression evaluated by real-time PCR was concordant with up-regulated or down-regulated expression evaluated by microarray analysis.Conclusion： PS showed a potential antidepressant-like effect, of which the action mechanism might be due togone expression regulation in hippocampus.