Non-stressed Mice Research Articles

Real-Time Imaging Assessment of Stress-Induced Vascular Inflammation Using Heartbeat-Synchronized Motion Compensation.

Chronic mental stress accelerates atherosclerosis through complicated neuroimmune pathways, needing for advanced imaging techniques to delineate underlying cellular mechanisms. While histopathology, ex vivo imaging, and snapshots of in vivo images offer promising evidence, they lack the ability to capture real-time visualization of blood cell dynamics within pulsatile arteries in longitudinal studies. An electrically tunable lens was implemented in intravital optical microscopy, synchronizing the focal plane with heartbeats to follow artery movements. ApoE-/- mice underwent 2 weeks of restraint stress before baseline imaging followed by 2 weeks of stress exposure in the longitudinal imaging, while nonstressed mice remained undisturbed. The progression of vascular inflammation was assessed in the carotid arteries through intravital imaging and histological analyses. A 4-fold reduction of motion artifact, assessed by interframe SD, and an effective temporal resolution of 25.2 Hz were achieved in beating murine carotid arteries. Longitudinal intravital imaging showed chronic stress led to a 6.09-fold (P=0.017) increase in myeloid cell infiltration compared with nonstressed mice. After 3 weeks, we observed that chronic stress intensified vascular inflammation, increasing adhered myeloid cells by 2.45-fold (P=0.031), while no significant changes were noted in nonstressed mice. Microcirculation imaging revealed increased circulating, rolling, and adhered cells in stressed mice's venules. Histological analysis of the carotid arteries confirmed the in vivo findings that stress augmented plaque area, myeloid cell and macrophage accumulation, and necrotic core volume while reducing fibrous cap thickness indicating accelerated plaque formation. We visualized the 3-dimensional structure of the carotid artery and 4-dimensional dynamics of the venules in the cremaster muscle. Dynamic focusing motion compensation intravital microscopy enabled subcellular resolution in vivo imaging of blood cell dynamics in beating arteries under chronic restraint stress in real time. This novel technique emphasizes the importance of advanced in vivo imaging for understanding cardiovascular disease.

Transcriptome signatures of the medial prefrontal cortex underlying GABAergic control of resilience to chronic stress exposure.

Analyses of postmortem human brains and preclinical studies of rodents have identified somatostatin (SST)-positive interneurons as key elements that regulate the vulnerability to stress-related psychiatric disorders. Conversely, genetically induced disinhibition of SST neurons or brain region-specific chemogenetic activation of SST neurons in mice results in stress resilience. Here, we used RNA sequencing of mice with disinhibited SST neurons to characterize the transcriptome changes underlying GABAergic control of stress resilience. We found that stress resilience of male but not female mice with disinhibited SST neurons is characterized by resilience to chronic stress-induced transcriptome changes in the medial prefrontal cortex. Interestingly, the transcriptome of non-stressed stress-resilient male mice resembled the transcriptome of chronic stress-exposed stress-vulnerable mice. However, the behavior and the serum corticosterone levels of non-stressed stress-resilient mice showed no signs of physiological stress. Most strikingly, chronic stress exposure of stress-resilient mice was associated with an almost complete reversal of their chronic stress-like transcriptome signature, along with pathway changes indicating stress-induced enhancement of mRNA translation. Behaviorally, the mice with disinhibited SST neurons were not only resilient to chronic stress-induced anhedonia - they also showed an inversed anxiolytic-like response to chronic stress exposure that mirrored the chronic stress-induced reversal of the chronic stress-like transcriptome signature. We conclude that GABAergic dendritic inhibition by SST neurons exerts bidirectional control over behavioral vulnerability and resilience to chronic stress exposure that is mirrored in bidirectional changes in expression of putative stress resilience genes, through a sex-specific brain substrate.

Endogenous opioids facilitate stress-induced binge eating via an insular cortex-claustrum pathway.

Stress has been shown to promote the development and persistence of binge eating behaviors. However, the neural circuit mechanisms for stress-induced binge-eating behaviors are largely unreported. The endogenous dynorphin (dyn)/kappa opioid receptor (KOR) opioid neuropeptide system has been well established to be a crucial mediator of the anhedonic component of stress. Here, we aimed to dissect the basis of dynorphinergic control of stress-induced binge-like eating behavior. We first established a mouse behavioral model for stress-induced binge-like eating behaviors. We found that mice exposed to stress increased their food intake of familiar palatable food (high fat, high sugar, HPD) compared to non-stressed mice. Following a brain-wide analysis, we isolated robust cFos-positive cells in the Claustrum (CLA), a subcortical structure with highly abundant KOR expression, following stress-induced binge-eating behavior. We report that KOR signaling in CLA is necessary for this elevated stress-induced binge eating behavior using local pharmacology and local deletion of KOR. In vivo calcium recordings using fiber photometry revealed a disinhibition circuit structure in the CLA during the initiation of HPD feeding bouts. We further established the dynamics of endogenous dynorphinergic control of this behavior using a genetically encoded dynorphin biosensor, Klight. Combined with 1-photon single-cell calcium imaging, we report significant heterogeneity with the CLA population during stress-induced binge eating and such behavior attenuates local dynorphin tone. Furthermore, we isolate the anterior Insular cortex (aIC) as the potential source of endogenous dynorphin afferents in the CLA. By characterizing neural circuits and peptidergic mechanisms within the CLA, we uncover a pathway that implicates endogenous opioid regulation stress-induced binge eating.

Omega-3 alleviates behavioral and molecular changes in a mouse model of stress-induced juvenile depression

IntroductionDepression is increasingly diagnosed in adolescence, necessitating specific prevention and treatment methods. However, there is a lack of animal models mimicking juvenile depression. This study explores a novel model using ultrasound (US) stress in juvenile mice. MethodsWe employed the US stress model in one-month-old C57/BL6 mice, exposing them to alternating ultrasound frequencies (20–25 kHz and 25–45 kHz) for three weeks. These frequencies correspond to negative and neutral emotional states in rodents and can induce a depressive-like syndrome. Concurrently, mice received either an omega-3 food supplement (FS) containing eicosapentaenoic acid (EPA; 0.55 mg/kg/day) and docosahexaenoic acid (DHA; 0.55 mg/kg/day) or a vehicle. Post-stress, we evaluated anxiety- and depressive-like behaviors, blood corticosterone levels, brain expression of pro-inflammatory cytokines, and conducted metabolome analysis of brain, liver and blood plasma. ResultsUS-exposed mice treated with vehicle exhibited decreased sucrose preference, a sign of anhedonia, a key feature of depression, increased anxiety-like behavior, elevated corticosterone levels, and enhanced TNF and IL-1β gene expression in the brain. In contrast, US-FS mice did not display these changes. Omega-3 supplementation also reduced anxiety-like behavior in non-stressed mice. Metabolomic analysis revealed US-induced changes in brain energy metabolism, with FS increasing brain sphingomyelin. Liver metabolism was affected by both US and FS, while plasma metabolome changes were exclusive to FS. Brain glucose levels correlated positively with activity in anxiety tests. ConclusionChronic omega-3 intake counteracted depressive- and anxiety-like behaviors in a US model of juvenile depression in mice. These effects likely stem from the anti-inflammatory properties of the supplement, suggesting potential therapeutic applications in juvenile depression.

Sex- Specific Effects of Chronic Unpredictable Stress on Renal Mitochondrial Function in C57BL/6 Mice

Background: The kidneys play a central role in long term control of blood pressure. Chronic psychological stress has been linked to hypertension, inflammation, and is associated with both an increased prevalence and progression of renal disease. However, the molecular and cellular mechanisms by which chronic stress alters renal function are not fully understood, and data on sex specific differences in renal and blood pressure responses to stress are limited. The impact of stress to cause local inflammation and mitochondrial dysfunction in the kidneys is not clear and represents a potential mechanism leading to renal dysfunction and hypertension. The chronic unpredictable stress (CUS) model induces psychological stress in rodents thus allowing the study of integrated physiological changes to the kidney in response to stress. Hypothesis: Exposing mice to CUS will lead to increased blood pressure that is associated with decreased renal mitochondrial function and renal inflammation. Methods: To investigate the link between stress, renal mitochondrial function, inflammation, and blood pressure, we exposed adult male and female C57BL/6 mice to CUS for 28 consecutive days. Blood pressure was measured via tail cuff on the 28th day of stress. 24 hours later, mice were euthanized and estrous cycle phase in females was determined by vaginal cytology. Kidney mitochondrial oxygen consumption was assessed using the Oroboros respirometer on the pole ends of the left kidneys. NLR family pyrin domain containing 3 (NLRP3) expression was measured in the right kidney using Real-Time TaqMan PCR. Results: Mean arterial pressure was elevated in both stressed males (in mmHg, 106±4 vs 87±4, n<11, p<0.01) and females (131± 6 vs 94±11 n<4, p<0.01) compared to non-stressed mice. While maximal electron transport capacity (ETS), a measure of mitochondrial function, was unchanged in CUS male kidneys, stressed females in the receptive (estrus and proestrus) phase of the estrous cycle exhibited a significant decrease in renal ETS compared to non-stressed females (in pmol O2/mg protein, 14.4±0.9 vs 11.4±0.7, n=11, p<0.05). Renal mRNA expression of NLRP3 was increased in CUS males (35%± 7.7, n=8, p<0.01) but unchanged in CUS females. Conclusion: These data suggest potential mechanistic differences in stress-induced hypertension in renal mitochondria (decreased ETS in stressed females) and inflammasome level (increased NLRP3 in stressed males) between male and female mice. This work was supported by P20GM109091 (F.H.), R01HL130972-01A1, R01HL5949, BX000168-10A1, BX005320 (F.G.S.), R01DK132948 (C.W. & F.P.) R01 MH129798 (SKW); VA VISN7 RDA (F.H.),, 1U54HL169191-01 & BX002604 (M.J.R.). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the abstract. This is the full abstract presented at the American Physiology Summit 2024 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.

The Influence of Stress and Binge-Patterned Alcohol Drinking on Mouse Skeletal Muscle Protein Synthesis and Degradation Pathways.

Adverse experiences (e.g., acute stress) and alcohol misuse can both impair skeletal muscle homeostasis, resulting in reduced protein synthesis and greater protein breakdown. Exposure to acute stress is a significant risk factor for engaging in alcohol misuse. However, little is known about how these factors together might further affect skeletal muscle health. To that end, this study investigated the effects of acute stress exposure followed by a period of binge-patterned alcohol drinking on signaling factors along mouse skeletal muscle protein synthesis (MPS) and degradation (MPD) pathways. Young adult male C57BL/6J mice participated in the Drinking in the Dark paradigm, where they received 2-4 h of access to 20% ethanol (alcohol group) or water (control group) for four days to establish baseline drinking levels. Three days later, half of the mice in each group were either exposed to a single episode of uncontrollable tail shocks (acute stress) or remained undisturbed in their home cages (no stress). Three days after stress exposure, mice received 4 h of access to 20% ethanol (alcohol) to model binge-patterned alcohol drinking or water for ten consecutive days. Immediately following the final episode of alcohol access, mouse gastrocnemius muscle was extracted to measure changes in relative protein levels along the Akt-mTOR MPS, as well as the ubiquitin-proteasome pathway (UPP) and autophagy MPD pathways via Western blotting. A single exposure to acute stress impaired Akt singling and reduced rates of MPS, independent of alcohol access. This observation was concurrent with a potent increase in heat shock protein seventy expression in the muscle of stressed mice. Alcohol drinking did not exacerbate stress-induced alterations in the MPS and MPD signaling pathways. Instead, changes in the MPS and MPD signaling factors due to alcohol access were primarily observed in non-stressed mice. Taken together, these data suggest that exposure to a stressor of sufficient intensity may cause prolonged disruptions to signaling factors that impact skeletal muscle health and function beyond what could be further induced by periods of alcohol misuse.

Short- and Long-Term Effects of Subchronic Stress Exposure in Male and Female Brain-Derived Neurotrophic Factor Knock-In Val66Met Mice.

Stress is an important risk factor for the onset of anxiety and depression. The ability to cope with stressful events varies among different subjects, probably depending on different genetic variants, sex and previous life experiences. The Val66Met variant of Brain-Derived Neurotrophic Factor (BDNF), which impairs the activity-dependent secretion of BDNF, has been associated with increased susceptibility to the development of various neuropsychiatric disorders. Adult male and female wild-type Val/Val (BDNFV/V) and heterozygous Val/Met (BDNFV/M) mice were exposed to two sessions of forced swimming stress (FSS) per day for two consecutive days. The mice were behaviorally tested 1 day (short-term effect) or 11 days (long-term effect) after the last stress session. Protein and mRNA levels were measured in the hippocampus 16 days after the end of stress exposure. Stressed mice showed a higher anxiety-like phenotype compared to non-stressed mice, regardless of the sex and genotype, when analyzed following the short period of stress. In the prolonged period, anxiety-like behavior persisted only in male BDNFV/M mice (p < 0.0001). Interestingly, recovery in male BDNFV/V mice was accompanied by an increase in pCREB (p < 0.001) and Bdnf4 (p < 0.01) transcript and a decrease in HDAC1 (p < 0.05) and Dnmt3a (p = 0.01) in the hippocampus. Overall, our results show that male and female BDNF Val66Met knock-in mice can recover from subchronic stress in different ways.

NSC689857, an inhibitor of Skp2, produces antidepressant-like effects in mice.

We have previously reported that two inhibitors of an E3 ligase S-phase kinase-associated protein 2 (Skp2), SMIP004 and C1, have an antidepressant-like effect in non-stressed and chronically stressed mice. This prompted us to ask whether other Skp2 inhibitors could also have an antidepressant effect. Here, we used NSC689857, another Skp2 inhibitor, to investigate this hypothesis. The results showed that administration of NSC689857 (5 mg/kg) produced an antidepressant-like effect in a time-dependent manner in non-stressed male mice, which started 8 days after drug administration. Dose-dependent analysis showed that administration of 5 and 10 mg/kg, but not 1 mg/kg, of NSC689857 produced antidepressant-like effects in both non-stressed male and female mice. Administration of NSC689857 (5 mg/kg) also induced antidepressant-like effects in non-stressed male mice when administered three times within 24 h (24, 5, and 1 h before testing) but not when administered acutely (1 h before testing). In addition, NSC689857 and fluoxetine coadministration produced additive antidepressant-like effects in non-stressed male mice. These effects of NSC689857 were not associated with the changes in locomotor activity. Administration of NSC689857 (5 mg/kg) also attenuated depression-like behaviors in male mice induced by chronic social defeat stress, suggesting therapeutic potential of NSC689857 in depression. Overall, these results suggest that NSC689857 is capable of exerting antidepressant-like effects in both non-stressed and chronically stressed mice.

Characterization of CD4+ t cell subsets during Chlamydia muridarum genital infection

Characterization of CD4+ T cell subsets in a stressed mouse model during Chlamydia muridarum genital infection. Katrina Davis, Tesfaye Belay. Department of Applied Sciences and Mathematics. Chlamydia genital infection caused by Chlamydia trachomatis is highly prevalent in low socioeconomic status populations. Whether stress is a risk factor for the high incidence of chlamydia remains to be investigated. CD4+ T cells are the major immune cells that fight chlamydia genital infection; however, the dynamics of CD4+ T cells during stressful conditions and chlamydia infection are unclear. The profiles of CD4+ T cell signature cytokines: T helper 1 (IL-12, Interferon-gamma (IFN-γ), Thelper2 (IL-4, IL-5, IL-13), Thelper17(IL-17), and Treg (IL-10). Mice were stressed for 5 minutes daily for 21 days and infected with 1x 105 IFU/ml of Chlamydia muridarum intravaginally. Non-stressed mice were infected as controls. Mice were sacrificed 7 days post-infection, the spleen and lymph nodes were harvested, and T cells were purified and proliferated for 72 h. Determination of the levels of cytokines in culture supernatants of T cells from stressed and non-stressed mice are underway for poster presentation. Supported by the NIH grant # 1R15AI124156-01 awarded to BSU

Growth Hormone Secretagogue Receptor (GHSR) Signaling in the Ventral Tegmental Area (VTA) Mediates Feeding Produced by Chronic Social Defeat Stress in Male Mice

Ghrelin, a hormone secreted by the stomach, binds to the growth hormone secretagogue receptor (GHSR) in various brain regions to produce a number of behavioral effects that include increased feeding motivation. During social defeat stress, ghrelin levels rise in correlation with increased feeding and potentially play a role in attenuating the anxiogenic effects of social defeat. One region implicated in the feeding effects of ghrelin is the ventral tegmental area (VTA), a region implicated in reward seeking behaviors, and linked to social defeat in mice. Here we examined the role of GHSR signaling in the VTA in feeding behavior in mice exposed to social defeat stress. Male C57BL/J6 mice that were socially defeated once daily for 3 weeks ate more, had higher plasma ghrelin level and increased GHSR expression in the VTA compared to non-stressed mice. Socially defeated GHSR KO mice failed to increase their caloric intake in response to this stressor but rescue of GHSR expression in the VTA restored feeding responses. Finally, we pharmacologically blocked VTA GHSR signalling with JMV2959 infused via an indwelling VTA cannula connected to a minipump. Vehicle-treated mice increased their caloric intake during social defeat, but JMV2959-infusions attenuated feeding responses and increased anxiety-like behaviors. The data suggest that GHSR signalling in the VTA is critical for the increases in appetite observed during chronic social defeat stress. Furthermore, these data support the idea that GHSR signaling in the VTA may also have anxiolytic effects, and blocking GHSR in this region may result in an anxiety-like phenotype.

Early life stress aggravates disease pathogenesis in mice with experimental autoimmune encephalomyelitis: Support for a two-hit hypothesis of multiple sclerosis etiology

Vision problems are one of the earliest diagnosed symptoms of multiple sclerosis (MS). The onset and progression of vision loss and the underlying pathogenesis in MS may be influenced by cumulative psychophysiological stress. Here, we used a two-hit model of stress in female mice to determine if early life stress (ELS, the first hit) influences the response to an immunization that induces experimental autoimmune encephalomyelitis (EAE, the second hit) later in life. We hypothesized that ELS caused by animal transportation from a vendor during early postnatal development represents a co-factor which can exacerbate the clinical severity of EAE. Indeed, adult EAE mice with a history of ELS displayed more severe clinical signs and delayed recovery compared to non-stressed EAE mice. ELS also diminished visual acuity measured by optokinetic responses, as well as locomotion and exploratory behaviours in EAE mice. Notably, ELS accelerated vision loss and caused earlier onset of visual impairments in EAE. Exacerbated functional impairments in stressed EAE mice were highly correlated with circulating corticosterone levels. The findings show that the progression of induced EAE in adulthood can be significantly impacted by adverse early life experiences. These observations emphasize the importance of comprehensive behavioural testing, including non-motor functions, to enhance the translational value of preclinical animal models of MS. Moreover, shipment stress of laboratory animals should be considered a necessary variable in preclinical MS research. The consideration of cumulative lifetime stresses provides a new perspective of MS pathogenesis within a personalized medicine framework.

Early tactile stimulation influences the development of Alzheimer's disease in gestationally stressed APP NL-G-F adult offspring NL-G-F/NL-G-F mice

Alzheimer's disease (AD) is associated with cerebral plaques and tangles, reduced synapse number, and shrinkage in several brain areas and these morphological effects are associated with the onset of compromised cognitive, motor, and anxiety-like behaviours. The appearance of both anatomical and behavioural symptoms is worsened by stress. The focus of this study was to examine the effect of neonatal tactile stimulation on AD-like behavioural and neurological symptoms on APP NL-G-F/NL-G-F mice, a mouse model of AD, who have been gestationally stressed. Our findings indicate that neonatal tactile stimulation improves cognition, motor skills, and anxiety-like symptoms in both gestationally stressed and non-stressed adult APP mice and that these alterations are associated with reduced Aβ plaque formation. Thus, tactile stimulation appears to be a promising non-invasive preventative strategy for slowing the onset of dementia in aging animals.

Cathepsin S deficiency improves muscle mass loss and dysfunction via the modulation of protein metabolism in mice under pathological stress conditions.

Cathepsin S (CTSS) is a widely expressed cysteinyl protease that has garnered attention because of its enzymatic and non-enzymatic functions under inflammatory and metabolic pathological conditions. Here, we examined whether CTSS participates in stress-related skeletal muscle mass loss and dysfunction, focusing on protein metabolic imbalance. Eight-week-old male wildtype (CTSS+/+ ) and CTSS-knockout (CTSS-/- ) mice were randomly assigned to non-stress and variable-stress groups for 2 weeks, and then processed for morphological and biochemical studies. Compared with non-stressed mice, stressed CTSS+/+ mice showed significant losses of muscle mass, muscle function, and muscle fiber area. In this setting, the stress-induced harmful changes in the levels of oxidative stress-related (gp91phox and p22phox ,), inflammation-related (SDF-1, CXCR4, IL-1β, TNF-α, MCP-1, ICAM-1, and VCAM-1), mitochondrial biogenesis-related (PPAR-γ and PGC-1α) genes and/or proteins and protein metabolism-related (p-PI3K, p-Akt, p-FoxO3α, MuRF-1, and MAFbx1) proteins; and these alterations were rectified by CTSS deletion. Metabolomic analysis revealed that stressed CTSS-/- mice exhibited a significant improvement in the levels of glutamine metabolism pathway products. Thus, these findings indicated that CTSS can control chronic stress-related skeletal muscle atrophy and dysfunction by modulating protein metabolic imbalance, and thus CTSS was suggested to be a promising new therapeutic target for chronic stress-related muscular diseases.

Systematic evaluation of a predator stress model of depression in mice using a hierarchical 3D-motion learning framework

Investigation of the neurobiology of depression in humans depends on animal models that attempt to mimic specific features of the human disorder. However, frequently-used paradigms based on social stress cannot be easily applied to female mice which has led to a large sex bias in preclinical studies of depression. Furthermore, most studies focus on one or only a few behavioral assessments, with time and practical considerations prohibiting a comprehensive evaluation. In this study, we demonstrate that predator stress effectively induced depression-like behaviors in both male and female mice. By comparing predator stress and social defeat models, we observed that the former elicited a higher level of behavioral despair and the latter elicited more robust social avoidance. Furthermore, the use of machine learning (ML)-based spontaneous behavioral classification can distinguish mice subjected to one type of stress from another, and from non-stressed mice. We show that related patterns of spontaneous behaviors correspond to depression status as measured by canonical depression-like behaviors, which illustrates that depression-like symptoms can be predicted by ML-classified behavior patterns. Overall, our study confirms that the predator stress induced phenotype in mice is a good reflection of several important aspects of depression in humans and illustrates that ML-supported analysis can simultaneously evaluate multiple behavioral alterations in different animal models of depression, providing a more unbiased and holistic approach for the study of neuropsychiatric disorders.

Implications of gut microbiota dysbiosis and fecal metabolite changes in psychologically stressed mice.

Psychological stress can induce affective disorders. Gut microbiota plays a vital role in emotional function regulation; however, the association between gut microbiota and psychological stress is poorly understood. We investigated effects of psychological stress on the gut microbiome and fecal metabolites and assessed the relationship between affective disorder behavior and altered fecal microbiota. A psychological stress model was established in C57BL/6J mice using a communication box. Sucrose preference test, forced swim test, and open field test helped assess anxiety- and depression-like behaviors. Fecal microbiota transplantation (FMT) was conducted using fecal samples from stressed and non-stressed mice. Moreover, 16S rRNA gene sequencing and untargeted metabolomics were performed. After stress exposure for 14 days, a significant increase in anxiety- and depression-like behaviors was observed. FMT of "affective disorder microbiota" from psychologically stressed mice increased stress sensitivity relative to FMT of "normal microbiota" from non-stressed mice. 16S rRNA gene sequencing revealed decreased abundance of Bacteroides, Alistipes, and Lactobacillus and increased abundance of Parasutterella and Rikenellaceae_RC9_gut_group in stressed mice; furthermore, stressed mice showed differential metabolite profiles. KEGG pathway analysis indicated that differential metabolites were chiefly involved in the downregulated pathways of α-linolenic acid metabolism, taste transduction, and galactose metabolism. Alistipes and Bacteroides were mainly positively correlated and Parasutterella was mainly negatively correlated with diverse metabolites. Our findings suggest that gut microbiome dysbiosis contributes to affective disorder development in response to psychological stress.

Psychological stress triggers the accumulation of myeloid-derived suppressor cells in the liver through adrenergic receptor signaling

Abstract Chronic physiological stress affects tightly regulated crosstalk between the nervous and the immune system. However, the exact mechanism(s) of stress-induced immunomodulation is far from clear. Immature myeloid cells such as myeloid-derived suppressor cells (MDSCs) impair immune responses, which may in turn increase susceptibility to infections and cancer or their complications. In a mouse model of chronic psychological stress through physical restraint, we found mononuclear-MDSC (Mo-MDSC) cells, defined as CD11b +Ly6C highLy6G highcells, to accumulate in the liver. These cells expressed high levels of Mo-MDSC markers such as CD14, TLR4, IL-4Rα as well as markers indicative of anti-apoptotic and immunosuppressive functions (HIF-1α, G-CSFR, Bcl2, Arg2, iNOS). In addition, FACS sorted MDSCs suppressed production of IFN-ɣ by T cells in vitro. Increased levels of proinflammatory mediators were detectable in serum samples of stressed mice, with circulating levels of IL-6 and G-CSF being the most pronounced mediators. The observed intrahepatic accumulation of Mo-MDSC-like cells was mediated by IL-6 and G-CSF and could be reversed by treatment with corresponding receptor-blocking monoclonal antibodies. In addition, this phenomenon was driven by norepinephrine (NE), the main neurotransmitter released from sympathetic neurons, and could be recapitulated by in vivo administration of NE to non-stressed mice. Our work reveals a novel mechanism of immunosuppression that is governed by adrenergic and inflammatory cues and alters the cellular landscape of the liver with implications for immune surveillance against microbes and cancer. This work was funded by NSERC

Intact Recognition Memory and Altered Hippocampal Glucocorticoid Receptor Signaling in Fkbp5-deficient Mice Following Acute Uncontrollable Stress.

The FK506 binding protein 5 (FKBP5) is a co-chaperone that regulates the activity of the glucocorticoid receptor (GR) and has been reported to mediate stress resilience. This study aimed to determine the effects of Fkbp5 deletion on acute stress-induced recognition memory impairment and hippocampal GR signaling. Wild-type and Fkbp5-knockout mice were subjected to acute uncontrollable stress induced by restraint and electrical tail shock. First, we assessed the cognitive status of mice using a novel object recognition task. Next, we measured plasma corticosterone, GR levels, and the levels of GR phosphorylation at serine 211 in the hippocampus. Wild-type mice exhibited stress-induced memory impairments, whereas Fkbp5-knockout mice did not. Plasma corticosterone and GR levels did not differ between the non-stressed wild-type and Fkbp5-knockout mice, but the levels of phosphorylated GR were lower in Fkbp5-knockout mice than in wild-type mice. Wild-type and Fkbp5-knockout mice showed increased nuclear GR levels following stress, indicating GR translocation. However, cytosolic phosphorylated GR levels were lower in the hippocampi of Fkbp5-knockout mice following stress than in those of wild-type mice. These results suggest that FKBP5 deficiency increases resilience to acute stress by altering GR signaling.

Thelper1 and T helper2 cytokine production patterns in stressed beta2-adrenergic receptor-deficient mice during chlamydia genital infection.

Murine CD4+ T cells have two distinct cytokine secretion patterns to play different functions. We have demonstrated that cold-induced stress (CIS) suppresses the immune system leading to increased intensity of Chlamydia muridaum in mice. We have reported that beta2-adrenergic receptor (b2-AR) knockout (KO) mice resist chlamydia genital infection. However, the cytokine profile of CD4+ T cells is not well explored. This study aimed to determine the cytokine production of Th1 and Th2 types in stressed and non-stressed b2-AR KO mice. We investigated the cytokine production levels of stressed and non-stressed mice during C. muridarum genital infection. Significantly increased production of cytokines was observed in plates pre-coated with anti-CD3/CD28 or in the presence of Con A or LPS. A decrease in the production of IFN-g and IL-2, whereas an increase in the secretion of IL-10, IL-13, and IL-23 in CD4+ T cells of stressed wild-type mice was obtained; however, the secretion of IFN-g and IL-2 was restored in T cells of b2-AR KO mice. Moreover, in vitro proliferation of CD4 T cells in the presence of b2-AR antagonists, ICI 118, 551 stimulated the production of Th1 cytokines, whereas b2-AR agonist, Fenoterol, decreased the production of Th1-type cytokines. Overall, Th1 cytokine responses are reduced in stressed mice suggesting that the cytokine status was polarized toward a Th2 immune response that can be restored by removing b2-AR from immune cells.

Roles of dendritic cells and macrophages during Chlamydia muridarum genital infection in stressed mice

It is well known that stress has a significant impact on public health and more evidence shows that stressors generally are greater in populations of lower socioeconomic status. Cold-induced stress inhibits immune cell activity and has been shown to cause different expressions of cytokine profiles during Chlamydia muridarum genital infection. Dendritic cells (DCs) and macrophages(MØ) are immune cells that express beta2-adrenergic receptor (β2-AR) and with key roles during chlamydia genital infection. This study is aimed to investigate the role of β2-AR by using a β2-AR knockout mouse. Wildtype (WT) and β2-AR knockout(KO) mice were stressed for five minutes daily and infected with C. muridarum intravaginally. Non-stressed infected mice of the groups were used as controls. Bone-marrow-derived DCs and MØ were tested isolated, counted differentiated, and proliferated for cytokine production. The proliferation of DCs and macrophages in the presence/absence of β2-AR agonists and antagonists was tested. Data show that non-stressed mice had a higher production of cytokines than stressed mice. Increased production of TNF-α in LPS-treated DCs and MØ of WT and β2-AR KO was observed. The effect of fenoterol and ICI118,55 antagonist showed no significant difference in cytokine production. The data indicate that β2-AR KO and WT had a similar pattern of cytokine production suggesting that deficiency in β2-AR restores the function of immune cells during genital infection. Experiments are undergoing to fully understand the mechanisms involved in modulating the function of DC and MØ during chlamydia genital infection.

Analysis of immune response of stressed male mice during Chlamydia muridarum lung infection

Chlamydia lung infection caused by Chlamydia trachomatis is a serious lung infection particularly in infants but there are a few studies about lung infection. Chlamydial genital and lung infections in mice is frequently done with Chlamydia muridarum. The purpose of our study was to explore the effect of stress in lung chlamydia infection in male mice. We hypothesize that stress changes the level of cytokine production in monocytes during lung infection. As a The result from these studies include counting of live cells, differentiation of proliferation of bone marrow derived monocytes where the result showed that non-stressed dendritic cells (DCs) WT mice and stressed macrophages (MO) WT mice had sufficient live cells to study. No difference in IL-b production of macrophages of stressed and non-stress mice was observed. Beta2-adrenergic receptor (b2-AR agonist (fenoterol) and antagonist (ICI 118 551) treatment resulted in difference in cytokine production the effect of agonists and antagonists. TNF-alpha production was high in stressed mice compared to non-stress mice for MO. LPS stimulated TNF-alpha production in DC but showed no difference in stressed and non-stressed mice. However, TNF-alpha production in macrophages of non-stressed mice was decreased. The production of IFN-g, in Con A- and LPS-treated splenic T cells. In contrast IL-5, IL-10, and IL-23 production was high in T cells of stressed mice. T cells treated with norepinephrine and bet2-adrenergic receptor agonist, Fenoterol. Overall, the production of cytokines in stressed and non-stressed mice shows variation that may have roles in enhancing protection or increased lung infection.