Bruton's tyrosine kinase drives neuroinflammation and anxiogenic behavior in mouse models of stress.

BACKGROUND: Current therapies targeting several neurotransmitter systems are only able to partially mitigate the symptoms of stress- and trauma-related disorder. Stress and trauma-related disorders lead to a prominent inflammatory response in humans, and in pre-clinical models. However, mechanisms underlying the induction of neuroinflammatory response in PTSD and anxiety disorders are not clearly understood. The present study investigated the mechanism underlying the activation of proinflammatory NLRP3 inflammasome and IL1ß in mouse models of stress. METHODS: We used two mouse models of stress, i.e., mice subjected to physical restraint stress with brief underwater submersion, and predator odor stress. Mice were injected with MCC950, a small molecule specific inhibitor of NLRP3 activation. To pharmacologically inhibit BTK, a specific inhibitor ibrutinib was used. To validate the observation from ibrutinib studies, a separate group of mice was injected with another BTK-specific inhibitor LFM-A13. Seven days after the induction of stress, mice were examined for anxious behavior using open field test (OFT), light-dark test (LDT), and elevated plus maze test (EPM). Following the behavior tests, hippocampus and amygdale were extracted and analyzed for various components of NLRP3-caspase 1-IL1ß pathway. Plasma and peripheral blood mononuclear cells were also used to assess the induction of NLRP3-Caspase 1-IL-1ß pathway in stressed mice. RESULTS: Using two different pre-clinical models of stress, we demonstrate heightened anxious behavior in female mice as compared to their male counterparts. Stressed animals exhibited upregulation of proinflammatory IL1ß, IL-6, Caspase 1 activity and NLRP3 inflammasome activation in brain, which were significantly higher in female mice. Pharmacological inhibition of NLRP3 inflammasome activation led to anxiolysis as well as attenuated neuroinflammatory response. Further, we observed induction of activated Bruton's tyrosine kinase (BTK), an upstream positive-regulator of NLRP3 inflammasome activation, in hippocampus and amygdala of stressed mice. Next, we conducted proof-of-concept pharmacological BTK inhibitor studies with ibrutinib and LFM-A13. In both sets of experiments, we found BTK inhibition led to anxiolysis and attenuated neuroinflammation, as indicated by significant reduction of NLRP3 inflammasome and proinflammatory IL-1ß in hippocampus and amygdala. Analysis of plasma and peripheral blood mononuclear cells indicated peripheral induction of NLRP3-caspase 1-IL1ß pathway in stressed mice. CONCLUSION: Our study identified BTK as a key upstream regulator of neuroinflammation, which drives anxiogenic behavior in mouse model of stress. Further, we demonstrated the sexually divergent activation of BTK, providing a clue to heightened neuroinflammation and anxiogenic response to stress in females as compared to their male counterparts. Our data from the pharmacological inhibition studies suggest BTK as a novel target for the development of potential clinical treatment of PTSD and anxiety disorders. Induction of pBTK and NLRP3 in peripheral blood mononuclear cells of stressed mice suggest the potential effect of stress on systemic inflammation.

Causal roles of stress kinase JNK2 in DNA methylation and binge alcohol withdrawal-evoked behavioral deficits.

Excessive binge alcohol intake is a common drinking pattern in humans, especially during holidays. Cessation of the binge drinking often leads to aberrant withdrawal behaviors, as well as serious heart rhythm abnormalities (clinically diagnosed as Holiday Heart Syndrome (HHS)). In our HHS mouse model with well-characterized binge alcohol withdrawal (BAW)-induced heart phenotypes, BAW leads to anxiety-like behaviors and cognitive impairment. We have previously reported that stress-activated c-Jun NH(2)-terminal kinase (JNK) plays a causal role in BAW-induced heart phenotypes. In the HHS brain, we found that activation of JNK2 (but not JNK1 and JNK3) in the prefrontal cortex (PFC), but not hippocampus and amygdala, led to anxiety-like behaviors and impaired cognition. DNA methylation mediated by a crucial DNA methylation enzyme, DNA methyltransferase1 (DNMT1), is known to be critical in alcohol-associated behavioral deficits. In HHS mice, JNK2 in the PFC (but not hippocampus and amygdala) causally enhanced total genomic DNA methylation via increased DNMT1 expression, which was regulated by enhanced binding of JNK downstream transcriptional factor c-JUN to the DNMT1 promoter. JNK2-specific inhibition either by an inhibitor JNK2I or JNK2 knockout completely offset c-JUN-regulated DNMT1 upregulation and restored the level of DNA methylation in HHS PFC to the baseline levels seen in sham controls. Strikingly, either JNK2-specific inhibition or genetic JNK2 depletion or DNMT1 inhibition (by an inhibitor 5-Azacytidine) completely abolished BAW-evoked behavioral deficits. In conclusion, our studies revealed a novel mechanism by which JNK2 drives BAW-evoked behavioral deficits through a DNMT1-regulated DNA hypermethylation. JNK2 could be a novel therapeutic target for alcohol withdrawal treatment and/or prevention.

Effect of rutin on anxiety-like behavior and activity of acetylcholinesterase isoforms in specific brain regions of pentylenetetrazol-treated mice.

The aim of the present study was to investigate the effect of rutin administration (100 mg/kg/day) to pentylenetetrazol (PTZ)-treated Balb-c mice (60 mg/kg/day), with respect to anxiety-like behavior using both open-field and elevated plus-maze (EPM) tests, and acetylcholinesterase (AChE) activity in salt-soluble (SS) fraction and detergent-soluble (DS) fraction of the cerebral cortex, hippocampus, striatum, midbrain, and diencephalon. Our results demonstrated that the administration of PTZ in 3 doses and the induction of seizures increased significantly anxiety behavior of mice and reduced significantly DS-AChE activity in all brain regions examined, while the reduction in the SS fraction was brain region-specific. Rutin administration to normal mice did not affect their behavior, while it induced a brain region-specific reduction in SS-AChE and a significant decrease in DS-AChE in all brain regions. We demonstrated for the first time that pretreatment of PTZ-mice with rutin (PTZ + Rutin group) prevented the manifestation of anxiety and induced interestingly a further significant reduction on the SS- and DS-AChE activities only in the cerebral cortex and striatum, in comparison with PTZ group. Our results show that rutin exhibits an important anxiolytic effect and an anticholinesterase activity in specific brain areas in the seizure model of PTZ.

Pilot Study on the Effect of Biophysical Therapy on Salivary Alpha-Amylase as a Surrogate Measure of Anxiety/Stress: In Search of a Novel Noninvasive Molecular Approach for the Management of Stress.

Anxiety and depression impact dramatically on public health, underlying the importance of alternative cost-effective treatments. Previous studies have shown that biophysical treatment can significantly reduce anxiety symptoms and recently, salivary alpha-amylase (SAA) has been identified as an objective correlate of the sympathetic-parasympathetic imbalance related to increased stress burden, defined as allostatic load. The aim of this study was to evaluate the effect of biophysical therapy on SAA levels, in addition to the Depression Anxiety Stress Scale (DASS)-21 questionnaire. Twenty-four workers (sales representatives) presenting with mild anxiety/stress symptoms (Generalized Anxiety Disorder 7-item scale of > 5) were randomized to biophysical treatment (N = 12) or placebo control (N = 12). The biophysical group underwent electromagnetic information transfer through an aqueous system procedure, with daily self-administration for one month. SAA collection and the DASS-21 questionnaire were undertaken at baseline and after one month in all patients. Clinical characteristics and baseline DASS-21 subscale scores were similar between placebo and biophysical group at baseline. After one month, patients receiving biophysical therapy had significantly reduced SAA levels compared to the placebo group (27.8 ± 39.4 vs. 116.8 ± 114.9 U/mL, p = 0.019). All three DASS-21 subscales, depression (9.3 ± 5.1 vs. 5.7 ± 5.5, p = 0.1), anxiety (6.7 ± 25 vs. 3.7 ± 2.2, p = 0.0049) and stress (10.8 ± 4.2 vs. 7.3 ± 3.7, p = 0.041) were also decreased after biophysical treatment compared to placebo after one month. Our findings suggest that biophysical therapy can benefit workers with mild (subclinical) anxiety/stress. These results were also validated by the concomitant reduction of SAA levels and an improvement in DASS-21 subscales. The underlying molecular mechanisms of this therapy remain to be characterized.

Early-stage paired housing improves social interaction in neuronal Uba6-deficient mice.

UBA6 is an alternative enzyme for ubiquitin activation in vertebrates that plays a pivotal role in early mouse development. Previously, we reported that the Uba6 brain-specific knockout (NKO) mouse is a novel autism spectrum disorder (ASD) mouse model that displays decreased social behavior and communication. To determine the therapeutic impact of environmental stimulation in ASDs, we investigated the behavioral and molecular changes of the NKO and control mice after exposure to environmental enrichment and paired housing in different developmental phases. Our results demonstrated that early paired housing could diminish the ASD phenotypes of NKO mice such as impaired nest building and social interaction and anxiety. Additionally, increased histone acetylation in the amygdala was observed in NKO mice after paired housing without a change in Ube3a levels. Our data suggest that paired housing at an early time point can play a crucial role in ameliorating ASD behavior and can be applied in other ASD animal models or clinical settings.

The GCs-SGK1-ATP Signaling Pathway in Spinal Astrocytes Underlied Presurgical Anxiety-Induced Postsurgical Hyperalgesia.

BACKGROUND: Patients undergoing surgery often feel anxious. Accumulating evidence indicated that presurgical anxiety was related to the more severe postsurgical pain. An animal model was established that exposed Sprague-Dawley rats to a single-prolonged stress (SPS) procedure to induce presurgical anxiety-like behaviors. The experiment revealed that presurgical anxiety not only aggravated but also prolonged postsurgical pain. However, the underlying mechanisms were unknown. METHODS: The rats in group C + Cort, group I + Cort, group A + Cort, and group AI + Cort were injected with corticosterone. The rats in group C + RU486, group I + RU486, group A + RU486, and group AI + RU486 were injected with mifepristone (RU486). The rats in group C + GSK650394 and group AI + GSK650394 were injected with GSK650394. The rats in group C + FC1 and group AI + FC1 were injected with fluorocitrate (FC) 30 minutes before SPS, 30 minutes before incision, and on postoperative days 1, 2, 3, 4, and 5. The rats in group C + FC2 and group AI + FC2 were injected with FC on postoperative days 7, 8, 9, 10, 11, 12, and 13. The paw withdrawal mechanical threshold was assessed 24 hours before SPS and from postoperative days 1 to 28. The level of corticosterone was determined by enzyme-linked immunosorbent assay. The expression of serum/glucocorticoid regulated kinase 1 (SGK1), interleukin-1ß, and tumor necrosis factor-α was visualized by Western blot. The concentrations of adenosine triphosphate (ATP) were measured by ATP assay kit. RESULTS: This study showed SPS elevated plasma glucocorticoids and ATP release from astrocytes, which meant the mechanical pain hypersensitivity in presurgical anxiety-induced postsurgical hyperalgesia was dependent on GCs-SGK1-ATP signaling pathway. SGK1 protein level in astrocytes was increased in response to the glucocorticoid stimuli and enhanced the extracellular release of ATP. Furthermore, spinal astrocytes played a key role in the maintenance. Targeting spinal astrocytes in maintenance phase prevented the pathological progression. CONCLUSIONS: These data suggested an important signaling pathway that affected the pain sensitivity after operation caused by presurgical anxiety.

High serum levels of malondialdehyde and antioxidant enzymes are associated with post-stroke anxiety.

Post-stroke anxiety (PSA) is a common neuropsychiatric affective disorder occurring after stroke. The purpose of this study was to investigate the association between anxiety and the serum levels of oxidative stress markers at admission. First-ever or recurrent ischemic stroke patients were consecutively recruited into the study and followed up 1 month. Patients were divided into PSA and non-PSA group according DSM-IV criteria for anxiety due to stroke. Overall, 49 patients (24.1%) were diagnosed anxiety. Serum GPX (glutathione peroxidase), CAT (catalase), SOD (superoxide dismutase), and MDA (malondialdehyde) were significantly higher in patients with anxiety than patients without anxiety. The HAM-A scores had a significant positive association with MDA levels. In multivariate logistic regression analysis, serum antioxidant enzymes and MDA were independent predictors of PSA. An increased risk of PSA was associated with serum MDA levels ≥ 3.0 nmol/ml (adjusted OR 8.68, 95% CI 3.02-24.95; P < 0.001) after adjusting for relevant confounders such as social support and treatments at admission. Elevated serum levels of lipid oxidation products and antioxidant enzymes at admission were associated with anxiety 1 month after stroke, suggesting that these alterations might participate in the pathophysiology of anxiety symptoms in stroke patients.

Activation of Extracellular Signal-Regulated Kinases (ERK 1/2) in the Locus Coeruleus Contributes to Pain-Related Anxiety in Arthritic Male Rats.

Background: There is increasing evidence suggesting that the Locus Coeruleus plays a role in pain-related anxiety. Indeed, we previously found that prolonged arthritis produces anxiety-like behavior in rats, along with enhanced expression of phosphorylated extracellular signal-regulated kinase 1/2 (a marker of plasticity) in the Locus Coeruleus. However, it is unknown how this effect correlates with the electrophysiological activity of Locus Coeruleus neurons or pain-related anxiety. Methods: Using the complete Freund's adjuvant model of monoarthritis in male Sprague-Dawley rats, we studied the behavioral attributes of pain and anxiety as well as Locus Coeruleus electrophysiology in vivo 1 (MA1W) and 4 weeks (MA4W) after disease induction. Results: The manifestation of anxiety in MA4W was accompanied by dampened tonic Locus Coeruleus activity, which was coupled to an exacerbated evoked Locus Coeruleus response to noxious stimulation of the inflamed and healthy paw. When a mitogen-activating extracellular kinase inhibitor was administered to the contralateral Locus Coeruleus of MA4W, the phosphorylated extracellular signal-regulated kinase 1/2 levels in the Locus Coeruleus were restored and the exaggerated evoked response was blocked, reversing the anxiogenic-like behavior while pain hypersensitivity remained unaltered. Conclusion: As phosphorylated extracellular signal-regulated kinase 1/2 blockade in the Locus Coeruleus relieved anxiety and counteracted altered LC function, we propose that phosphorylated extracellular signal-regulated kinase 1/2 activation in the Locus Coeruleus plays a crucial role in pain-related anxiety.

Xiao Yao San Improves the Anxiety-Like Behaviors of Rats Induced by Chronic Immobilization Stress: The Involvement of the JNK Signaling Pathway in the Hippocampus.

The current study evaluated the effects of Xiao Yao San (XYS) on anxiety-like behaviors and sought to determine whether the c-Jun N-terminal kinase (JNK) signaling pathway is involved. A total of 40 rats were divided into 5 groups (n=8): the control group (deionized water, per os (p.o.)), the model group (deionized water, p.o.), the SP600125 group (surgery), the per se group (surgery), and the XYS group (3.9 g/kg/d, p.o.). A 1% dimethyl sulfoxide (DMSO) citrate buffer solution (2 µL/ventricle/d) and SP600125 (10 µg/ventricle, 2 µL/ventricle/d) were separately and bilaterally injected into the rats of the two surgery groups via the ventricular system of the brain. All but the control group underwent 14 d of chronic immobilization stress (CIS; 3 h/d). On day 15, the body weights of all of the rats were measured; additionally, the rats were subjected to the elevated plus maze (EPM) and novelty suppressed feeding (NSF) tests. Finally, JNK signaling pathway indices, including phosphorylated JNK (P-JNK), JNK, phosphorylated c-Jun (P-c-Jun) and cytochrome C (Cyt-C), were examined. After modeling, the body weight and behavioral analyses of the model rats indicated that this modeling method induced anxiety-like behaviors. P-JNK, JNK, and P-c-Jun were altered in the hippocampus of the model rats. After 14 d of treatment with XYS and SP600125, rat body weight and behaviors as well as P-JNK, JNK, and P-c-Jun had changed. However, no significant difference in Cyt-C was found. XYS improves the anxiety-like behaviors induced by CIS, which might be related to the JNK signaling pathway in the hippocampus.

Effects of environmental enrichment in aged mice on anxiety-like behaviors and neuronal nitric oxide synthase expression in the brain.

Previous studies have shown that an enriched environment (EE) has an important effect on brain function via the neuronal nitric oxide synthase/nitric oxide (nNOS/NO) pathway in young and aged animals. However, whether EE induces its effect by altering nNOS expression levels and whether it lowers anxiety-like behaviors in aged mice remains unclear. Here, we show that nNOS expression levels increased with age in the hippocampus and cerebellum in aged mice, but not in the cortex. Moreover, EE reduced anxiety-like behaviors in aged mice and reduced nNOS expression levels in the cerebellum, but not in the cortex. The present study suggests that EE improves anxiety-like behaviors in aged mice by altering nNOS expression levels in the hippocampus or cerebellum.

The impact of mitochondrial aldehyde dehydrogenase (ALDH2) activation by Alda-1 on the behavioral and biochemical disturbances in animal model of depression.

The etiology of depression remains still unclear. Recently, it has been proposed, that mitochondrial dysfunction may be associated with development of mood disorders, such as depression, bipolar disorder and anxiety disorders. Mitochondrial aldehyde dehydrogenase (ALDH2), an enzyme responsible for the detoxification of reactive aldehydes, is considered to exert protective function in mitochondria. We investigated the influence of Alda-1, a small-molecule activator of ALDH2, on depressive- and anxiety-like behaviors in an animal model of depression - the prenatally stressed rats - using behavioral, molecular and proteomic methods. Prolonged Alda-1 administration significantly increased the climbing time, tended to reduce the immobility time and increased the swimming time of the prenatally stressed rats in the forced swim test. Moreover, treatment of prenatally stressed rats with Alda-1 significantly increased number of entries into the open arms of the maze and the time spent therein, as assessed by elevated plus-maze test. Such actions were associated with reduction of plasma 4-HNE-protein content, decrease of TNF-α mRNA and increase of PGC-1α (regulator of mitochondrial biogenesis) mRNA level in the frontal cortex and hippocampus of the prenatally stressed rats as well as with normalization of peripheral immune parameters and significant changes in expression of 6 and 4 proteins related to mitochondrial functions in the frontal cortex and hippocampus, respectively. Collectively, ALDH2 activation by Alda-1 led to a significant attenuation of depressive- and anxiety-like behaviors in the prenatally stressed rats. The pattern of changes suggested mitoprotective effect of Alda-1, however the exact functional consequences of the revealed alterations require further investigation.

The Role of Protein Kinase A in Anxiety Behaviors.

This review focuses on the genetic and other evidence supporting the notion that the cyclic AMP (cAMP) signaling pathway and its mediator, the protein kinase A (PKA) enzyme, which respond to environmental stressors and regulate stress responses, are central to the pathogenesis of disorders related to anxiety. We describe the PKA pathway and review in vitro animal studies (mouse) and other evidence that support the importance of PKA in regulating behaviors that lead to anxiety. Since cAMP signaling and PKA have been pharmacologically exploited since the 1940s (even before the identification of cAMP as a second messenger with PKA as its mediator) for a number of disorders from asthma to cardiovascular diseases, there is ample opportunity to develop therapies using this new knowledge about cAMP, PKA, and anxiety disorders.

Monoamine Oxidase A is Required for Rapid Dendritic Remodeling in Response to Stress.

BACKGROUND: Acute stress triggers transient alterations in the synaptic release and metabolism of brain monoamine neurotransmitters. These rapid changes are essential to activate neuroplastic processes aimed at the appraisal of the stressor and enactment of commensurate defensive behaviors. Threat evaluation has been recently associated with the dendritic morphology of pyramidal cells in the orbitofrontal cortex (OFC) and basolateral amygdala (BLA); thus, we examined the rapid effects of restraint stress on anxiety-like behavior and dendritic morphology in the BLA and OFC of mice. Furthermore, we tested whether these processes may be affected by deficiency of monoamine oxidase A (MAO-A), the primary enzyme catalyzing monoamine metabolism. METHODS: Following a short-term (1-4h) restraint schedule, MAO-A knockout (KO) and wild-type (WT) mice were sacrificed, and histological analyses of dendrites in pyramidal neurons of the BLA and OFC of the animals were performed. Anxiety-like behaviors were examined in a separate cohort of animals subjected to the same experimental conditions. RESULTS: In WT mice, short-term restraint stress significantly enhanced anxiety-like responses, as well as a time-dependent proliferation of apical (but not basilar) dendrites of the OFC neurons; conversely, a retraction in BLA dendrites was observed. None of these behavioral and morphological changes were observed in MAO-A KO mice. CONCLUSIONS: These findings suggest that acute stress induces anxiety-like responses by affecting rapid dendritic remodeling in the pyramidal cells of OFC and BLA; furthermore, our data show that MAO-A and monoamine metabolism are required for these phenomena.

Loss of CDKL5 disrupts kinome profile and event-related potentials leading to autistic-like phenotypes in mice.

Mutations in the X-linked cyclin-dependent kinase-like 5 (CDKL5) gene have been identified in neurodevelopmental disorders including atypical Rett syndrome (RTT), autism spectrum disorders (ASDs), and early infantile epileptic encephalopathy. The biological function of CDKL5 and its role in the etiology of these disorders, however, remain unclear. Here we report the development of a unique knockout mouse model of CDKL5-related disorders and demonstrate that mice lacking CDKL5 show autistic-like deficits in social interaction, as well as impairments in motor control and fear memory. Neurophysiological recordings reveal alterations in event-related potentials (ERPs) similar to those observed in RTT and ASDs. Moreover, kinome profiling uncovers disruption of multiple signal transduction pathways, including the AKT-mammalian target of rapamycin (mTOR) cascade, upon Cdkl5 loss-of-function. These data demonstrate that CDKL5 regulates signal transduction pathways and mediates autistic-like phenotypes and together establish a causal role for Cdkl5 loss-of-function in neurodevelopmental disorders.

Copy number variation of glyoxalase I.

The glyoxalase I gene GLO1 is a hotspot for copy number variation in the human and mouse genomes. The additional copies are often functional, giving rise to 2-4-fold increased glyoxalase I expression and activity. The prevalence of GLO1 copy number increase in the human population appears to be approximately 2% and may be linked to a risk of obesity, diabetes and aging. Increased GLO1 copy number has been found in human tumour cell lines and primary human tumours. The minimum common copy number increase region was approximately 1 Mb and it contained GLO1 and seven other genes. The increased copy number was generally functional, being associated with increased glyoxalase I protein and multidrug resistance in cancer chemotherapy. Glo1 duplication in the mouse genome is found within approximately 0.5 Mb of duplicated DNA. It was claimed to be linked to anxiety phenotypes, but other related discordant findings have doubted the association with glyoxalase I and further investigation is required.

Diet-induced obesity progressively alters cognition, anxiety-like behavior and lipopolysaccharide-induced depressive-like behavior: focus on brain indoleamine 2,3-dioxygenase activation.

Obesity is associated with a high prevalence of mood symptoms and cognitive dysfunctions that emerges as significant risk factors for important health complications such as cardiovascular diseases and type 2 diabetes. It is therefore important to identify the dynamic of development and the pathophysiological mechanisms underlying these neuropsychiatric symptoms. Obesity is also associated with peripheral low-grade inflammation and increased susceptibility to immune-mediated diseases. Excessive production of proinflammatory cytokines and the resulting activation of the brain tryptophan catabolizing enzyme indoleamine 2,3-dioxygenase (IDO) have been shown to promote neurobehavioral complications, particularly depression. In that context, questions arise about the impact of diet-induced obesity on the onset of neuropsychiatric alterations and the increased susceptibility to immune-mediated diseases displayed by obese patients, particularly through brain IDO activation. To answer these questions, we used C57Bl/6 mice exposed to standard diet or western diet (WD; consisting of palatable energy-dense food) since weaning and for 20 weeks. We then measured inflammatory and behavioral responses to a systemic immune challenge with lipopolysaccharide (LPS) in experimental conditions known to alter cognitive and emotional behaviors independently of any motor impairment. We first showed that in absence of LPS, 9 weeks of WD is sufficient to impair spatial recognition memory (in the Y-maze). On the other hand, 18 weeks of WD increased anxiety-like behavior (in the elevated plus-maze), but did not affect depressive-like behavior (in the tail-suspension and forced-swim tests). However, 20 weeks of WD altered LPS-induced depressive-like behavior compared to LPS-treated lean mice and exacerbated hippocampal and hypothalamic proinflammatory cytokine expression and brain IDO activation. Taken together, these results show that WD exposure alters cognition and anxiety in unstimulated conditions and enhances activation of neurobiological mechanisms underlying depression after immune stimulation. They suggest therefore that obesity, and possibly obesity-associated inflammatory priming, may represent a vulnerability state to immune-mediated depressive symptoms.

Disruption of fatty acid amide hydrolase activity prevents the effects of chronic stress on anxiety and amygdalar microstructure.

Hyperactivation of the amygdala following chronic stress is believed to be one of the primary mechanisms underlying the increased propensity for anxiety-like behaviors and pathological states; however, the mechanisms by which chronic stress modulates amygdalar function are not well characterized. The aim of the current study was to determine the extent to which the endocannabinoid (eCB) system, which is known to regulate emotional behavior and neuroplasticity, contributes to changes in amygdalar structure and function following chronic stress. To examine the hypothesis, we have exposed C57/Bl6 mice to chronic restraint stress, which results in an increase in fatty acid amide hydrolase (FAAH) activity and a reduction in the concentration of the eCB N-arachidonylethanolamine (AEA) within the amygdala. Chronic restraint stress also increased dendritic arborization, complexity and spine density of pyramidal neurons in the basolateral nucleus of the amygdala (BLA) and increased anxiety-like behavior in wild-type mice. All of the stress-induced changes in amygdalar structure and function were absent in mice deficient in FAAH. Further, the anti-anxiety effect of FAAH deletion was recapitulated in rats treated orally with a novel pharmacological inhibitor of FAAH, JNJ5003 (50 mg per kg per day), during exposure to chronic stress. These studies suggest that FAAH is required for chronic stress to induce hyperactivity and structural remodeling of the amygdala. Collectively, these studies indicate that FAAH-mediated decreases in AEA occur following chronic stress and that this loss of AEA signaling is functionally relevant to the effects of chronic stress. These data support the hypothesis that inhibition of FAAH has therapeutic potential in the treatment of anxiety disorders, possibly by maintaining normal amygdalar function in the face of chronic stress.

Enhanced hippocampus-dependent memory and reduced anxiety in mice over-expressing human catalase in mitochondria.

Oxidative stress (OS) and reactive oxygen species (ROS) play a modulatory role in synaptic plasticity and signaling pathways. Mitochondria (MT), a major source of ROS because of their involvement in energy metabolism, are important for brain function. MT-generated ROS are proposed to be responsible for a significant proportion of OS and are associated with developmental abnormalities and aspects of cellular aging. The role of ROS and MT function in cognition of healthy individuals is relatively understudied. In this study, we characterized behavioral and cognitive performance of 5- to 6-month-old mice over-expressing mitochondrial catalase (MCAT). MCAT mice showed enhancements in hippocampus-dependent spatial learning and memory in the water maze and contextual fear conditioning, and reduced measures of anxiety in the elevated zero maze. Catalase activity was elevated in MCAT mice in all brain regions examined. Measures of oxidative stress (glutathione, protein carbonyl content, lipid peroxidation, and 8-hydroxyguanine) did not significantly differ between the groups. The lack of differences in these markers of oxidative stress suggests that the differences observed in this study may be due to altered redox signaling. Catalase over-expression might be sufficient to enhance cognition and reduce measures of anxiety even in the absence of alteration in levels of OS.

Monoacylglycerol lipase inhibition-induced changes in plasma corticosterone levels, anxiety and locomotor activity in male CD1 mice.

The hypothalamus-pituitary-adrenal-axis is strongly controlled by the endocannabinoid system. The specific impact of enhanced 2-arachidonoylglycerol signaling on corticosterone plasma levels, however, was not investigated so far. Here we studied the effects of the recently developed monoacylglycerol lipase inhibitor JZL184 on basal and stress-induced corticosterone levels in male CD1 mice, and found that this compound dramatically increased basal levels without affecting stress responses. Since acute changes in corticosterone levels can affect behavior, JZL184 was administered concurrently with the corticosterone synthesis inhibitor metyrapone, to investigate whether the previously shown behavioral effects of JZL184 are dependent on corticosterone. We found that in the elevated plus-maze, the effects of JZL184 on "classical" anxiety-related measures were abolished by corticosterone synthesis blockade. By contrast, effects on the "ethological" measures of anxiety (i.e. risk assessment) were not affected by metyrapone. In the open-field, the locomotion-enhancing effects of the compound were not changed either. These findings show that monoacylglycerol lipase inhibition dramatically increases basal levels of corticosterone. This endocrine effect partly affects the anxiolytic, but not the locomotion-enhancing effects of monoacylglycerol lipase blockade.

Experimental gastritis leads to anxiety- and depression-like behaviors in female but not male rats.

Human and animals studies support the idea that there is a gender-related co-morbidity of pain-related and inflammatory gastrointestinal (GI) diseases with psychological disorders. This co-morbidity is the evidence for the existence of GI-brain axis which consists of immune (cytokines), neural (vagus nerve) and neuroendocrine (HPA axis) pathways. Psychological stress causes disturbances in GI physiology, such as altered GI barrier function, changes in motility and secretion, development of visceral hypersensitivity, and dysfunction of inflammatory responses. Whether GI inflammation would exert impact on psychological behavior is not well established. We examined the effect of experimental gastritis on anxiety- and depression-like behaviors in male and female Sprague-Dawley rats, and evaluated potential mechanisms of action. Gastritis was induced by adding 0.1% (w/v) iodoacetamide (IAA) to the sterile drinking water for 7 days. Sucrose preference test assessed the depression-like behavior, open field test and elevated plus maze evaluated the anxiety-like behavior. IAA treatment induced gastric inflammation in rats of either gender. No behavioral abnormality or dysfunction of GI-brain axis was observed in male rats with IAA-induced gastritis. Anxiety- and depression-like behaviors were apparent and the HPA axis was hyperactive in female rats with IAA-induced gastritis. Our results show that gastric inflammation leads to anxiety- and depression-like behaviors in female but not male rats via the neuroendocrine (HPA axis) pathway, suggesting that the GI inflammation can impair normal brain function and induce changes in psychological behavior in a gender-related manner through the GI-to-brain signaling.