Chronic Unpredictable Stress Group Research Articles

Chronic unpredictable stress during adolescence exerts sex-specific effects on depressive-like behavior and neural activation triggered by tail suspension test

During adolescence, acute stress can modify neuronal excitability in various brain regions, leading to negative behavioral outcomes. However, the impact of chronic stress during adolescence on neuronal responses to acute stimuli remains unclear. To address this, we subjected adolescent mice to 12 days of chronic unpredictable stress (CUS). Anxiety and depressive behaviors were evaluated, along with changes in c-Fos expression, which is one of the most widely used markers of neuronal activation. By comparing c-Fos immunoreactivity between the CUS and control groups both before and after the tail suspension test (TST), we found that adolescent CUS induced depressive behaviors in male mice, but not in female mice. Adolescent CUS primarily affected the excitability of neurons in the infralimbic cortex (IL), the dorsomedial and dorsolateral area of the bed nucleus of the stria terminalis (BNST), and the ventral hippocampus CA3. TST exerted a significant main effect on the density of c-Fos+ neurons in the prelimbic cortex (PL), infralimbic cortex (IL), cingulate areas 1 and 2 (Cg1, Cg2), the lateral septum (LS), Bed nucleus terminalis (BNST), and lateral habenular (LHb). Furthermore, the excitability of neurons in the paraventricular thalamic nucleus (PVT) was impacted by sex. These data suggest that adolescent CUS elicits region- and sex-specific modifications in TST-induced c-Fos expression, establishing a theoretical basis for understanding the pathophysiological alterations in mood disorders following adolescent stress.

Chronic unpredictable stress (CUS) reduced phoenixin expression, induced abnormal sperm and testis morphology in male rats

Chronic stress caused by prolonged emotional pressure can lead to various physiological issues, including reproductive dysfunction. Although reproductive problems can also induce chronic stress, the impact of chronic stress-induced reproductive dysfunction remains contentious. This study investigates the effects of chronic unpredictable stress (CUS) on reproductive neuropeptides, sperm quality, and testicular morphology. Sixteen twelve-week-old Sprague Dawley rats were divided into two groups: a non-stress control group and a CUS-induced group. The CUS regimen involved various stressors over 28 days, with both groups undergoing behavioural assessments through sucrose-preference and forced-swim tests. Hypothalamic gene expression levels of CRH, PNX, GPR173, kisspeptin, GnRH, GnIH, and spexin neuropeptides were measured via qPCR, while plasma cortisol, luteinizing hormone (LH), and testosterone concentrations were quantified using ELISA. Seminal fluid and testis samples were collected for sperm analysis and histopathological evaluation, respectively. Results showed altered behaviours in CUS-induced rats, reflecting stress impacts. Hypothalamic corticotropin-releasing hormone (CRH) expression and plasma cortisol levels were significantly higher in CUS-induced rats compared to controls (p < 0.05). Conversely, phoenixin (PNX) expression decreased in the CUS group (p < 0.05), while kisspeptin, spexin, and gonadotropin-inhibitory hormone (GnIH) levels showed no significant differences between groups. Despite a significant increase in GnRH expression (p < 0.05), plasma LH and testosterone concentrations were significantly lower (p < 0.05) in CUS-induced rats. Histopathological analysis revealed abnormal testis morphology in CUS-induced rats, including disrupted architecture, visible interstitial spaces between seminiferous tubules, and absence of spermatogenesis. In conclusion, CUS affects reproductive function by modulating PNX and GnRH expression, influencing cortisol levels, and subsequently reducing plasma LH and testosterone concentrations. This study highlights the complex interplay between chronic stress and reproductive health, emphasizing the significant impact of stress on reproductive functions.

Neuroprotective effect of bromelain on BDNF-TRKB signalling pathway in chronic unpredictable stress-induced depression model

BackgroundBromelain is a mixture of protease enzyme extract from the fruit or stem of the pineapple plant. It has a wide range of biological actions, and it is most commonly used as an anti-inflammatory agent. This study was designed to investigate the antidepressant effect of bromelain on chronic unpredictable stress (CUS)-induced depression in rat models by targeting various molecular mechanisms.ResultWe studied the in silico analysis of the antidepressant potential of bromelain by docking with various proteins involved in the pathophysiology of depression. As a result of in silico studies, bromelain showed good binding energy with IL1β, 5-HT, BDNF, CREB, and TrkB. The mRNA expression of BDNF, TrkB, AKT, ERK, and IL-1β was studied by qRT-PCR. Gene expression studies showed a significant decrease in BDNF, TrkB, AKT, and ERK in chronic unpredictable stress, whereas there was a significant increase in the case of the bromelain- and fluoxetine-treated group. Since neuroinflammation is also one of the major concerns in the pathophysiology of depression, pro-inflammatory cytokines were also studied along with apoptotic markers using ELISA. ELISA results showed a significant increase in inflammatory cytokines in CUS, and it was significantly decreased in the case of the bromelain- and fluoxetine-treated group. Similarly, there was an increased concentration of pro-apoptotic protein in the CUS group, whereas it was decreased in the bromelain and fluoxetine groups.ConclusionsFrom the results, it is clear that bromelain exerts an antidepressive effect by preventing neuroinflammation and neurodegeneration and by enhancing neurogenesis and neuroplasticity.Graphical abstract

Study the effects of empagliflozin on model of chronic depression and interleukin-6 in the brain of male rats

Abstract Background: Empagliflozin (EMP) is an oral anti-diabetic drug with pleiotropic effects such as anti-inflammatory properties. Objectives: To evaluate the antidepressant effect of EMP and describe the link between stress and interleukin-6 (IL-6) level in the brains of male rats. Materials and Methods: In this experiment, 50 rats were separated into five groups G1–G5. The sucrose preference test (SPT) was used to examine the antidepressant effects of fluoxetine and EMP. Rat interleukin enzyme-linked immunosorbent assay kit was used to measure the IL-6 level in rat brain tissue. SPT was performed on each rat on days 0, 10, and 25. Chronic unpredictable stress (CUS) was performed on each rat for 24 days. Results: By the end of day 10, all rats subjected to the CUS program had a substantial (P &lt; 0.05) reduction in sucrose intake index compared to day 0. EMP significantly increases sucrose intake compared to the stressed group. In comparison to the CUS group, fluoxetine significantly increases sucrose intake (P &lt; 0.05). In terms of IL-6, the mean IL-6 level in G2 was considerably greater than in G1. When compared to group 2, the mean IL-6 level was considerably lower in G3 and G5. Conclusions: EMP has antidepressant-like effects and can counteract the impact of stress-increased IL-6 levels in the brains of depressed rats.

Anti-depressant Potentials of Some Bioactive Components of Basella alba Leaves in Chronic Unpredictable Stress Rat Model

Bioactive components of Basella alba leaves are responsible for their antidepressant-like activity. However, the component with the greatest anti-depressant activity is unknown. This study investigated the antidepressant-like activities of the bioactive components (phenols, flavonoids, and glycosides) in Basella alba leaves. Forty-two male Wistar rats weighing 50–200 g were randomly divided into six groups (n=7). All the groups, except for the control, were subjected to chronic unpredictable stress (CUS) for five weeks. The rats in the CUS groups were treated with normal saline (1 mL/kg), escitalopram (5 mg/kg), and each of the phenol-, flavonoid-, and glycoside-rich Basella alba extracts (200 mg/kg) orally for twenty-one days. The tail suspension, sucrose preference, light-dark box, and hole-board tests were carried out before and after the induction of depression. In the CUS groups, reduced mobility time on tail suspension, increased percentage sucrose consumption, frequency of head dips on the hole board, and line-cross frequency in the light-dark box were observed. The latency on the hole board was reduced with Basella alba components, while there was a significant (p&lt;0.05) decrease in serum brain-derived neurotrophic factor and an increase in serum IL-6 in the CUS animals not treated with extracts. The phenol-rich Basella alba extract showed the most potent antidepressant-like activity, followed by the flavonoid-rich extract. The bioactive components of Basella alba, particularly phenols, were effective in ameliorating the depressive features of CUS and should be further studied for use as an adjunct or stand-alone antidepressant.

Antidepressant evaluation of Andrographis paniculata Nees extract and andrographolide in chronic unpredictable stress zebrafish model

Andrographis paniculata (A. paniculata) showed an anti-depressive effect in rodent models. Zebrafish has recently emerged as a worthy complementary translational model for antidepressant drug discovery study. This study investigates the anti-depressive effect of A. paniculata extract and andrographolide in the chronic unpredictable stress (CUS)- zebrafish model. Four groups of zebrafish (n = 10/group), i.e. control, CUS (stressed, untreated), CUS + A. paniculata (100 mg/L) and CUS + fluoxetine (0.01 mg/L) were assessed in open-field and social interaction tests, 24 h after treatment. After extract screening, behavioural and cortisol analysis of andrographolide (5, 25 and 50 mg/kg, i.p.) and fluoxetine (10 mg/kg, i.p.) were evaluated. Before the behavioural study, acute toxicity and characterization of A. paniculata extract using UHPLC-ESI-MS/MS were performed. A significant reduction in freezing duration was found in A. paniculata- (t-test, p = 0.0234) and fluoxetine-treated groups (t-test, p < 0.0001) compared to the CUS group. A significant increase in total distance travelled, and contact duration was observed only in the fluoxetine-treated group (t-test, p = 0.0007) and (t-test, p = 0.0207), respectively. A significant increase in highly mobile duration was observed in both treatment groups. Andrographolide (50 mg/kg, i.p.) acute treatment showed a significant reduction in freezing duration (p = 0.0042), duration in a dark area (p = 0.0338) and cortisol level (p = 0.0156) and increased total distance travelled (p = 0.0144). Twenty-six compounds were tentatively characterized by LC-MS/MS method, and andrographolide content is 0.042 μg/g. According to cortisol analysis, A. paniculata's LC50 is 627.99 mg/L, while andrographolide's EC50 was determined as 26.915 mg/kg. Further assessment of the cellular and molecular underpinnings of the anti-depressive effect of andrographolide is strongly recommended to evaluate the potential as an antidepressant.

Chronic unpredictable stress changes left ventricular geometry and activation of the inflammasome

Background: Psychosocial stress, particularly chronic unpredictable stress (CUS) has been identified as a risk factor for cardiovascular disease, but the effects of CUS on left ventricle (LV) function, geometry and systemic hemodynamics remains unclear. Moreover, whether and to what degree CUS activates intracellular formation of the inflammasome, including the NOD-LRR pyrin containing protein 3 (NLRP3) remains unknown. Accordingly, the central hypothesis of this project was that in a murine model of CUS, shifts in LV geometry and systemic hemodynamics would occur and be associated with the induction of NLRP3. Methods and Results: Mice (C57BL/6, 10wks of age, equal sex, n=16) were subjected to CUS for 4 weeks which entailed daily random stressors including tilted cage, wet bedding, and overnight light. Referent age and sex matched non CUS mice served as controls (n=16). LV function was assessed by high fidelity ultrasound and blood pressure measurements by tail cuff for both referent control and CUS mice. The changes in ambient heart rate (HR), LV end-systolic volume (LVEDV), and systolic blood pressure (SBP) were expressed as a function of referent control. All results are reported as mean ± SEM. While LV ejection fraction was equivalent between CUS and control groups (70 ± 3 vs. 67 ± 3%, respectively, p=0.48), HR trended higher (4.5 ± 3.1%, p=0.08), LVEDV was lower (-11 ± 5%, p=0.03) and SBP was elevated (12.1 ± 4.9%, p=0.04) - all indicative of heightened sympathetic tone. LV myocardial NLRP3 (rtPCR) was increased (1.47 ± 0.22 fold change, p=0.04) with CUS as was IL-1β (1.43 ± 0.20 fold change, p=0.05), a downstream cytokine activated by NLRP3. Conclusion: CUS, in and of itself, shifts LV geometry, causes mild hypertension and induction of the myocardial inflammasome- all of which would exacerbate/contribute to cardiovascular disease. This work was supported in part by National Institutes of Health grant R01HL130972-01A1 (F.G.S.), R01HL5949 (F.G.S.), RO1DK132948 (C.W. &amp; F.P.), R01 MH129798 (SKW) and P20GM109091 (F.H.) as well as a Merit Award from the Veterans Health Administration, BX000168-10A1 (F.G.S.), BX005320 (F.G.S.), BX002604 (M.J.R.), and VISN7 RDA (F.H.). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Effect of Bromelain on Chronic Unpredictable Stress-induced Behavioral, Biochemical, and Monoamine Changes in Wistar Albino Rat Model of Depression.

Bromelain is a complex mixture of protease enzyme extract from the fruit or stem of the pineapple plant and it has a history of folk medicine use. It is known to have a wide range of biological actions and it is most commonly used as an anti-inflammatory agent, though scientists have also discovered its potential as an anticancer and antimicrobial agent, it has been reported to have positive effects on the respiratory, digestive, circulatory systems and potentially on the immune system. This study was designed to investigate the antidepressant potential of Bromelain in the chronic unpredictable stress (CUS) model of depression. We studied the antioxidant activity, and neuroprotective effect of Bromelain by analyzing the fear and anxiety behavior, antioxidants, and neurotransmitter levels, and also by analyzing the histopathological changes. Adult male Wistar albino rats were divided into 5 groups, Control; Bromelain; CUS; CUS + Bromelain, CUS + fluoxetine. Animals of the CUS group, CUS + Bromelain group, and CUS + Fluoxetine group were exposed to CUS for 30 days. Animals of the Bromelain group and CUS + Bromelain group were treated orally with 40mg/kg Bromelain throughout the period of CUS whereas, the positive control group was treated with fluoxetine. Results showed a significant decrease in oxidative stress marker (lipid peroxidation), and the stress hormone cortisol, in Bromelain-treated CUS-induced depression. Bromelain treatment in CUS has also resulted in a significant increase in neurotransmitter levels, which indicates the efficacy of Bromelain to counteract the monamine neurotransmitter changes in depression by increasing their synthesis and reducing their metabolism. In addition, the antioxidant activity of Bromelain prevented oxidative stress in depressed rats. Also, hematoxylin and eosin staining of hippocampus sections has revealed that Bromelain treatment has protected the degeneration of nerve cells by chronic unpredictable stress exposure. This data provides evidence for the antidepressant-like action of Bromelain by preventing neurobehavioral, biochemical, and monoamine alterations.

Effects of Empagliflozin on a Model of Chronic Depression and Brain Toll-Like Receptors Gene Expression in Male Rats

Background: Empagliflozin is an antidiabetic medication having anti-inflammatory and antioxidant properties. Objectives: To develop a chronic unpredictable stress (CUS) model in male rats, evaluate empagliflozin’s antidepressant effects, and describe the link between stress, an antioxidant enzyme, and Toll-like receptor (TLR)-4 gene expression in male rats. Materials and Methods: In this experiment, 50 rats were divided into five groups: G1–G5. The forced swimming test (FST) was used to examine the antidepressant effects of fluoxetine and empagliflozin, and real-time polymerase chain reaction was used to measure TLR-4 gene expression. FST was performed on each rat on days 0, 10, and 25. CUS was performed on each rat for 24 days. Results: By the end of day 10, all animals subjected to the CUS program had a substantial (P &lt; 0.05) increase in immobility duration compared with day 0. The immobility duration in the CUS group was statistically significantly greater than the baseline. Stressed rats demonstrated a statistically significant reduction in immobility duration compared with CUS group G2. In comparison to the CUS group, fluoxetine significantly reduced immobility duration (P &lt; 0.05). In terms of gene expression, the mean of fold changes in TLR-4 mRNA level in group 2 was considerably greater than in group 1. When compared with group 2, the means of the fold changes in TLR-4 mRNA level were considerably lower in groups 3, 4, and 5. Conclusions: Empagliflozin has antidepressant-like effects and can counteract the impact of stress-induced TLR-4 overexpression in the hippocampus and elevate the activity of antioxidant enzymes in the brains of depressed rats.

Stress-induced myelin damage in the hippocampal formation in a rat model of depression

BackgroundAccording to previous studies, myelin damage may be involved in the occurrence of depression. However, to date, no study has quantitatively investigated the changes in myelinated fibers and myelin sheaths in the hippocampal formation (HF) and hippocampal subfields in the context of depression. Methods: Male Sprague-Dawley (SD) rats (aged 4–5 weeks) were evenly divided into the control group and chronic unpredictable stress (CUS) group. Behavioral tests were performed, and then changes in myelinated fibers and myelin ultrastructure in hippocampal subfields in depression model rats were investigated using modern stereological methods and transmission electron microscopy techniques. Results: After a four-week CUS protocol, CUS rats showed depressive-like and anxiety-like behaviors. The total length and total volume of myelinated fibers were reduced in the CA1 region and DG in the CUS group compared with the control group. The total volumes of myelin sheaths and axons in the CA1 region but not in the DG were significantly lower in the CUS group than in the control group. The decrease in the total length of myelinated nerve fibers in the CA1 region in CUS rats was mainly due to a decrease in the length of myelinated fibers with a myelin sheath thickness of 0.15 μm–0.20 μm. Limitations: The exact relationship between the degeneration of myelin sheaths and depression-like, anxiety-like behaviors needs to be further investigated. Conclusions: CUS induces depression- and anxiety-like behaviors, and the demyelination in the CA1 region induced by 4 weeks of CUS might be an important structural basis for these behaviors.

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

ObjectiveAlthough 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). MethodsTotal 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 2 groups 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. ResultsMC 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. ConclusionMC 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.

The effects of neonatal maternal deprivation and chronic unpredictable stresses on migraine-like behaviors in adult rats

BackgroundStress is known to cause migraine. This study investigates the effects of neonatal maternal deprivation (MD) and chronic unpredictable stress (CUS) on migraine in rats. MethodsSeventy rats were randomly divided into ten groups (five groups of each sex, and seven rats/group). The groups included: untreated intact, nitroglycerin (NTG) only, NTG + MD, NTG + CUS (10 weeks after birth), and NTG + MD + CUS. For the induction of MD, pups were separated from their mothers from postnatal day 2 to day 14. The CUS was conducted by daily exposure to different stressors for 2 weeks. For the induction of migraine after stress, NTG (5 mg/kg/IP) was administered every second day for 9 days. Afterward, NTG-related symptoms, including climbing behavior, facial rubbing, body grooming, freezing behavior, and head-scratching, were recorded for 90 min. Statistical differences between the groups were analyzed by one-way and two-way ANOVA followed by the Newman–Keuls test. ResultsMigraine symptoms, including increased head-scratching, facial rubbing, and decreased climbing behavior, were more significant in females than in males. Head scratching and facial rubbing increased in stressed females, but not in males as compared to NTG-treated rats. Body grooming was significantly decreased in MD males compared to the NTG group. The effects of NTG in MD + CUS on the rats did not differ from those in the MD or CUS groups. ConclusionsMD and CUS had a sex-related aggravating effect on the development of migraine, while the combination of MD and CUS had no additive migraine-aggravating effect.

Chronic unpredictable stress disturbs the blood–testis barrier affecting sperm parameters in mice

Research questionDoes chronic stress affect the key proteins and sperm parameters of the blood–testis barrier (BTB)? DesignC57Bl/6 mice were divided into two groups: a non-treated control group and a chronic unpredictable stress (CUS) applied group. The stress status of the animals was confirmed with behavioural tests. Histopathologic evaluation was conducted by haematoxylin and eosin staining and electron microscope. Malondialdehyde, corticosterone and testosterone levels were evaluated in peripheral blood. Expression levels of BTB proteins, namely zonula occludens-1 (ZO-1), claudin-11 (CLDN11) and clathrin in Sertoli cells, were assessed by Western blotting and immunofluorescence techniques. Sperm samples were collected from cauda epididymis, and sperm parameters analysed. ResultsThe stress model was confirmed by behavioural tests. Histopathological evaluation of the testes demonstrated a mild degeneration in seminiferous tubules. Malondialdehyde (P = 0.008) and corticosterone levels increased (P = 0.004) and testosterone levels decreased (P = 0.005) in the CUS group. Electron microscopic evaluation confirmed the damage in BTB integrity in the CUS group. Western blot analysis showed that ZO-1 and CLDN11 levels were significantly decreased, although clathrin levels were unchanged. Although sperm concentration and total motility rate were not significantly different between the groups, progressive motility (P = 0.03), normal sperm morphology (P = 0.04), chromatin integrity (toluidine blue) (P = 0.002) and the acrosomal reaction rate (P = 0.002) were significantly decreased, and acrosomal abnormality rate was dramatically increased (P = 0.04) in the CUS group. ConclusionsIn mice, CUS disrupted BTB integrity and impaired sperm parameters. A decrease in ZO-1 and CLDN11 expression levels may be proposed as the causative factor.

Study on construction of rat model of acute pancreatitis with syndrome of liver depression and Qi stagnation

The aim of this paper was to construct a rat model of acute pancreatitis(AP) with syndrome of liver depression and Qi stagnation, and provide evaluation tools for pharmacodynamic research and efficacy network verification of related traditional Chinese medicine in view of the etiology, pathogenesis and clinical manifestations of AP. According to the Chinese and Western medicine diagnosis and treatment guidelines for AP with syndrome of liver depression and Qi stagnation, etiology, pathogenesis and clinical syndromes in TCM, Meta-analysis results, and evaluation strategy of establishing an animal model combining disease and syndrome in our laboratory, the biological surrogate outcomes suitable for the evaluation of animal models of AP with syndrome of liver depression and Qi stagnation were extracted then. The chronic unpredictable stress(CUS) method and chronic unpredictable stress +L-arginine(CUS +L-Arg) method were used to construct the rat model, and the above biological surrogate outcomes were used to evaluate whether an AP rat model was established. During the experiment, the weight and syndrome scores of the rats were observed and recorded. At the end of the experiment, the rats' serum, organs and tissues were collected from the operation to detect the various indicators. As compared with the normal group, the syndrome scores of the CUS group and CUS +L-Arg group were significantly increased(P&lt;0.01); the anti-syndrome medicine Chaihu Shugan Pills could significantly reduce the syndrome scores of the two groups of rats(P&lt;0.01), indicating that both modeling methods can replicate the syndrome of liver depression and Qi stagnation in the rat model. As compared with the normal group, the serum amylase(AMY) activity level was increased by 3 times in the CUS +L-Arg group(P&lt;0.01), and the AMY activity level was also increased in CUS group, but not up to 3 times of the normal value. As compared with the normal group, the le-vels of TNF-α and IL-6 mRNA in the pancreatic tissues of the CUS +L-Arg group were significantly increased(P&lt;0.01); the levels of TNF-α mRNA in the pancreatic tissues of the CUS group were significantly increased(P&lt;0.05), but IL-6 mRNA level only showed a rising trend, indicating that only the CUS + L-Arg method can be used to replicate the AP damage in the disease-syndrome combination model. The CUS + L-Arg method can be used for continuous modeling for 4 weeks to establish a disease-syndrome combination model of AP rats with syndrome of liver depression and Qi stagnation. The model has the characteristics of repeatability, stability after mode-ling, low animal mortality, and similar clinical pathogenesis. It can be used for the evaluation of traditional Chinese medicine efficacy and the verification of efficacy network.

Eugenia uniflora fruit extract exerts neuroprotective effect on chronic unpredictable stress-induced behavioral and neurochemical changes.

The aim of the current study was to evaluate the effect of chronic administration of Eugenia uniflora fruit extract on behavioral parameters, oxidative stress markers, and acetylcholinesterase activity in an animal model of depression, which was induced by chronic unpredictable stress (CUS). Mice were divided into six groups as follows: control/vehicle (water), control/fluoxetine (20mg/kg), control/extract (200mg/kg), CUS/vehicle, CUS/fluoxetine (20mg/kg), and CUS/extract (200mg/kg). Animals of the CUS group were exposed to a series of stressors for a period of 21days. Vehicle, fluoxetine, and hydroalcoholic extract were administered daily by gavage. Results showed that E. uniflora treatment: (a) prevented the depressant-like effect induced by CUS; (b) regulated the activity of acetylcholinesterase; (c) reduced oxidative damage to lipids and reactive oxygen species production, in the prefrontal cortex and hippocampus; and (d) prevented the reduction of glutathione peroxidase in the hippocampus of animals subjected to CUS protocol. Taken together, our findings suggested that E. uniflora extract exerts a neuroprotective effect by preventing oxidative damage and decreasing CUS-induced acetylcholinesterase activity, thus, ameliorating depressive-type behavior. PRACTICAL APPLICATIONS: E. uniflora fruit extract revealed an antidepressant-like effect and prevented the oxidative damage as well as cholinergic alterations caused by chronic stress in mice. Therefore, we believe that the results obtained in this study can be used to develop an alternative therapy for the management of depressive disorders.

Antidepressant Effects of (S)-Ketamine through a Reduction of Hyperpolarization-Activated Current Ih.

SummaryCompelling evidence suggests that a single sub-anesthetic dose of (R,S)-ketamine exerts rapid and robust antidepressant effects. However, the cellular mechanisms underlying the antidepressant effects of (R,S)-ketamine remain unclear. Here, we show that (S)-ketamine reduced dendritic but not somatic hyperpolarization-activated current Ih of dorsal CA1 neurons in unstressed rats, whereas (S)-ketamine decreased both somatic and dendritic Ih in chronic unpredictable stress (CUS) rats. The reduction of Ih by (S)-ketamine was independent of NMDA receptors, barium-sensitive conductances, and cAMP-dependent signaling pathways in both unstressed and CUS groups. (S)-ketamine pretreatment before the onset of depression prevented CUS-induced behavioral phenotypes and neuropathological changes of dorsal CA1 neurons. Finally, in vivo infusion of thapsigargin-induced anxiogenic- and anhedonic-like behaviors and upregulation of functional Ih, but these were reversed by (S)-ketamine. Our results suggest that (S)-ketamine reduces or prevents Ih from being increased following CUS, which contributes to the rapid antidepressant effects and resiliency to CUS.

Effects of vagus nerve stimulation on inflammatory factors and apoptosis in depression model rats

Objective To explore the influence of vagus nerve stimulation (VNS) on inflammatory factors in serum and nucleus of the solitary tract, and hippocampal apoptosis in rats with depression induced by chronic unpredictable stress(CUS). Methods Totally 32 male Sprague Dawley (SD) rats aged 8-9 weeks were selected.Eight rats were chosen as control group, and the other 24 rats were treated as the depression model with CUS. The rats were randomly divided into CUS, fluoxetine and VNS group, with 8 rats in each group after successful modeling. The control group and CUS group were induced by normal saline. Fluoxetine group and VNS group were implanted with VNS stimulation electrode. The VNS stimulation lasted for 28 d. On the time points of before experiment, after modeling and after treatment, the sucrose consumption test and open-field test (OFT) were performed to observe the behavioral changes of rats. Elisa was used to detect the levels of TNF-α, IL-6 and IL-1β in serum and nucleus of the solitary tract. Cell apoptosis was observed with TUNEL staining in hippocampal CA1 region. Results (1) Sucrose consumption experiment and OFT showed that, compared with the CUS group, the consumption of sucrose, percentage of sucrose consumption, scores of vertical and horizontal movement increased significantly in the VNS group (consumption of sucrose: (11.78±2.67) ml, (8.06±2.85) ml; percentage of sucrose consumption: (72.31±9.98)%, (63.67±8.95)%; score of vertical movement: (16.61±3.98), (10.31±3.86); score of horizontal movement: (44.25±9.59), (36.21±7.21)) (t=4.87, 7.98, 5.87, 9.12, all P 0.05). (2) Compared with the CUS group, levels of TNF-α, IL-6 and IL-1β in serum decreased in the VNS group (serum TNF-α: (46.72±11.63) pg/ml, (125.47±15.18) pg/ml; serum IL-6: (243.65±38.90) pg/ml, (441.39±83.31) pg/ml; serum IL-1β: (209.31±32.45) pg/ml, (339.21±76.37) pg/ml) (t=-70.38, -196.25, -131.13, all P 0.05). Conclusion VNS can improve the depression behavior in rats with depression induced by CUS and the mechanism maybe related to inhibiting the expression of TNF-α, IL-6 and IL-1β in serum and nucleus of the solitary tract and cell apoptosis in hippocampal CA1 region. Key words: Chronic unpredictable stress; Vagus nerve stimulation; Nucleus of the solitary Tract; Inflammatory factors; Apoptosis; Rats

Low-Frequency Pulsed Magnetic Field Improves Depression-Like Behaviors and Cognitive Impairments in Depressive Rats Mainly via Modulating Synaptic Function.

Transcranial magnetic stimulation (TMS) has shown great promise as a medical treatment of depression. The effectiveness of TMS treatment at high frequency has been well investigated; however, low-frequency TMS in depression treatment has rarely been investigated in depression-induced cognitive deficits. Herein, this study was carried out to assess the possible modulatory role of low-frequency pulsed magnetic field (LFPMF) on reversing cognitive impairment in a model of depression induced by chronic unpredictable stress (CUS). Wistar rats were randomly allocated into four groups as follows: a control group (CON), a control applied with LFPMF (CON + LFPMF), a CUS group, and a CUS treated with LFPMF (CUS + LFPMF) group. During 8 weeks of CUS, compared to those in the CON group, animals not only gained less weight but also exhibited anhedonia, anxiety, and cognitive decline in behavioral tests. After 2-week treatment of LFPMF, a 20 mT, 1 Hz magnetic stimulation, it reversed the impairment of spatial cognition as well as hippocampal synaptic function including long-term potentiation and related protein expression. Thus, LFPMF has shown effectively improvements on depressant behavior and cognitive dysfunction in CUS rats, possibly via regulating synaptic function.

PKA-mediated phosphorylation of CREB and NMDA receptor 2B in the hippocampus of offspring rats is involved in transmission of mental disorders across a generation

This study is aimed at the mechanism of transmission of mental disorders across a generation. We used 10 different stressors to establish an animal model of chronic unpredictable stress (CUS) before pregnancy. Forced swimming test (FST) and open field test (OFT) were used to analyze the behavior of 30-day-old adolescent offspring rats born to stress mothers. Magnetic resonance spectroscopy was used to measure glutamate, gamma-aminobutyric acid (GABA), and glutamine. Phosphate-activated glutaminase (PAG), glutamate decarboxylase (GAD), GABA-transaminase (GABA-T), protein kinase A (PKA), cAMP response element-binding protein (CREB), and N-methyl-D-aspartate (NMDA) receptor 2B (NR2B) were detected by western blot. Adolescent offspring rats in the CUS group exhibited depressive-like behavior in the FST and anxious behavior in the OFT. GAD was increased and GABA-T was decreased, which resulted in an increase in GABA levels and decrease of the glutamate/GABA ratio in the hippocampus of CUS offspring rats. Disruption of the glutamate/GABA–glutamine cycle was related to decrease PKA-mediated phosphorylation of CREB and NR2B in the hippocampus. These findings highlight the importance of mental health of females before pregnancy and suggest that CUS before pregnancy reduces p-CREB and p-NR2B in the offspring hippocampus, which could be responsible for behavioral disorders in the adolescent offspring.

Abstract 2890: Paternal pre-conception stress as a potential risk factor for neuroblastoma

Abstract Neuroblastomas (NBs) are pediatric tumors that lack adequate treatment for their aggressive form, despite many efforts focusing on novel therapies. Although the disease is known to have a genetic background, its etiology cannot be explained solely by genetics. Yet, the role of environment in NB development is understudied. Thus, there are no established risk factors for NB aside from rare genetic predispositions and therefore no preventative measures. NBs arise due to defects in sympathetic neuron (SN) differentiation, which occurs in the fetus. Thus, we hypothesize that prenatal or pre-conception exposures may interfere with this process and promote NB. Maternal stress during pregnancy and paternal pre-conception stress induce neurodevelopmental defects in offspring by direct effects on fetal development or epigenetic changes, respectively. Indeed, we have previously shown that in a mouse model of NB prenatal stress interferes with SN differentiation in offspring, accelerating NB development and leading to its increased frequency and malignancy. The goal of the current study was to determine if a similar increase in risk of NB development can result from paternal pre-conception stress. To test the effect of paternal stress on NB, we subjected TH-MYCN males to 6 weeks of chronic unpredictable stress (CUS) prior to mating with wild type females and assessed the impact of stress on 1) miRNA content in the germline (RNAseq); 2) fertility and fetal survival; 3) NB frequency, latency, growth and metastasis in their hemizygous TH-MYCN offspring (periodic MRI, histological analyses). CUS resulted in significant changes in non-coding RNA content of the germline, including miRNAs and tsRNAs. Of particular interest was a stress-induced down-regulation of sperm miRNAs, including miR-200b and miR-128, which are involved in neuroblast differentiation, inhibit NB invasiveness and are inactivated in NB tumors. Paternal stress decreased the rate of successful pregnancies from 75% in controls to 37.5% and increased frequency of death in utero from 5.5% to 29.2% (p=0.02). Moreover, offspring of stressed fathers exhibited accelerated NB development (60% vs 0% of mice with tumors at 6 weeks of age for CUS and control groups, respectively; p=0.03) and growth (p=0.3), higher tumor frequency (60% vs 25% at 14 weeks of age, p=0.03), as well as a trend toward a more metastatic phenotype. However, these effects on NB formation and phenotype were observed solely in male offspring. In summary, our data provide the first evidence for the role of paternal preconception stress in NB development. If confirmed, our findings may open new avenues for NB prevention and identifying biomarkers of increased NB risk. Citation Format: Sung-Hyeok Hong, Susana Galli, Raquel Santana da Cruz, Lu Jin, Larissa Wetlisbach, Shiya Zhu, Abir Malik, Jason Tilan, Sonia de Assis, Joanna B. Kitlinska. Paternal pre-conception stress as a potential risk factor for neuroblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2890.