Fish oil or its major constituents, namely omega-3 poly-unsaturated fatty acid (n3-PUFA), are popular supplements to improve neurogenesis, neuroprotection, and overall brain functions. Our objective was to probe the implications of fat enriched diet with variable PUFAs supplements in ameliorating social stress (SS). We fed mice on either of the three diet types, namely the n-3 PUFA-enriched diet (ERD, n3:n6= 7:1), a balanced diet (BLD, n3:n6= 1:1) or a standard lab diet (STD, n3:n6= 1:6). With respect to the gross fat contents, the customized special diets, namely ERD and BLD were extreme diet, not reflecting the typical human dietary composition. Aggressor-exposed SS (Agg-E SS) model triggered behavioral deficiencies that lingered for 6 weeks (6w) post-stress in mice on STD. ERD and BLD elevated bodyweights but potentially helped in building the behavioral resilience to SS. STD adversely affected the gene networks of brain transcriptomics associated with the cell mortality, energy homeostasis and neurodevelopment disorder. Diverging from the ERD's influences on these networks, BLD showed potential long-term benefits in combatting Agg-E SS. The gene networks linked to cell mortality and energy homeostasis, and their subfamilies, such as cerebral disorder and obesity remained at the baseline level of Agg-E SS mice on BLD 6w post-stress. Moreover, neurodevelopment disorder network and its subfamilies like behavioral deficits remained inhibited in the cohort fed on BLD 6w post Agg-E SS.

Although Post-Traumatic Stress Disorder (PTSD) can be understood as dysregulation in fear extinction circuitry, it is not a unitary, homogeneous disorder; some symptoms of PTSD do not have direct fear-related correlates (anhedonia, emotional numbing, associated substance abuse). This study examines the relationship between the severity of PTSD symptom sub-clusters and changes in gray matter volume (GMV). 11 patients who had experienced interpersonal violence (females, mean age 38.4 yrs) participated in a MRI study and were assessed using the PTSD Checklist (PCL) (Weathers 1993). Individuals with current substance dependence, lifetime or current psychosis, bipolar disorder, current suicidality, and current psychoactive medications were excluded. A T1-weighted 3D MPRAGE volume was acquired (Siemens 3T Tim Trio, 176 sagittal slices, TR/TE=1900/2.52ms, effective resolution 1mm 3). Voxel-Based Morphometry (VBM) analysis was conducted in SPM5 (Ashburner 2000). Symptom subclusters of Avoidance, Dysphoria, Hyperarousal, and Intrusions were calculated according to Krause et al. (2008). Associations between GMV and subclusters were tested with linear regression, with age as covariate of no interest. Results were thresholded at p=0.0025, with cluster-level correction. Overall, symptom subcluster severity negatively correlated with GMV in regions previously implicated in PTSD. Severity of Avoidance symptoms (avoidant thinking, behavior) was negatively correlated with bilateral mid-cingulate (BA32) and anterior cingulate (BA24) cortices. Severity of Intrusions (flashbacks, nightmares, reliving) was negatively correlated with left-lateralized insula (BA13, BA47), inferior orbitofrontal cortex (BA47), postcentral gyrus (BA3), lingual gyrus, and calcarine sulcus (BA17). Hyperarousal (hypervigilant, “jumpy”) severity was not associated with GMV. Dysphoria symptom severity exhibited a trend of positive correlation with GMV in middle frontal gyrus (p=0.012). Total PCL score tended to correlate negatively (p=0.08) with GMV in right parahippocampal gyrus. These results support the hypothesis that PTSD is an imbalance of brain systems involved in approach and avoidance (Stein and Paulus 2009). Use of symptom sub-clusters revealed, to a greater degree than total PCL score, subtle changes in GMV. Specifically, severity of Intrusions was associated with decreased GMV in visual, motor, and pain processing areas while Dysphoria was positively correlated with GMV in middle frontal gyrus as observed in depressed patients. Greater knowledge of the structural correlates of symptom clusters may enhance understanding and treatment of PTSD.

INTRODUCTION:The glia-operated glymphatic system, analogous to but separate from the lymphatics in the periphery, is unique to brain and retina, where it is very closely aligned with the arteriolar system. This intimate relationship leads to a "blood vessel like" distribution pattern of glymphatic vessels in the brain. The spatial relationship of glymphatics, including their essential component aquaporin-4 with vascular pericytes of brain arterioles is critical to functionality and is termed "polarization".MATERIALS AND METHODS:We review the available literature on the factors affecting the resting state of glymphatics under normal conditions, including the important role of sleep in supporting normal glymphatic function (including waste removal) as well as the critical role of "polarization" under normal conditions. We then examine the effects of traumatic brain injury (TBI) or seizures on the glymphatic system and its state of "polarization".RESULTS:Injury, such as TBI, can disrupt polarization resulting in "depolarization" leading to brain edema.CONCLUSION:Damage to the glymphatic system might explain the brain edema so often seen following TBI or other insult. Moreover, similar damage should be expected in response to seizures, which can often be associated with chemical exposures as well as with TBI. Military operations, whether night operations or continuous operations, quite often impose limitations on sleep. As glymphatic function is sleep-dependent, sleep deprivation alone could compromise glymphatic function, as well, and might in addition, explain some of the well-known performance deficits associated with sleep deprivation. Possible effects of submarine and diving operations, chemical agents (including seizures), as well as high altitude exposure and other threats should be considered. In addition to the brain, the retina is also served and protected by the glymphatic system. Accordingly, the effect of military-related risks (e.g., exposure to laser or other threats) to retinal glymphatic function should also be considered. An intact glymphatic system is absolutely essential to support normal central nervous system functionality, including cognition. This effects a broad range of military threats on brain and retinal glymphatics should be explored. Possible preventive and therapeutic measures should be proposed and evaluated, as well.

The brain is the command center for the mammalian nervous system and an organ with enormous structural complexity. Protected within the skull, the brain consists of an outer covering of grey matter over the hemispheres known as the cerebral cortex. Underneath this layer reside many other specialized structures that are essential for multiple phenomenon important for existence. Acquiring samples of specific gross brain regions requires quick and precise dissection steps. It is understood that at the microscopic level, many sub-regions exist and likely cross the arbitrary regional boundaries that we impose for the purpose of this dissection. Mouse models are routinely used to study human brain functions and diseases. Changes in gene expression patterns may be confined to specific brain areas targeting a particular phenotype depending on the diseased state. Thus, it is of great importance to study regulation of transcription with respect to its well-defined structural organization. A complete understanding of the brain requires studying distinct brain regions, defining connections, and identifying key differences in the activities of each of these brain regions. A more comprehensive understanding of each of these distinct regions may pave the way for new and improved treatments in the field of neuroscience. Herein, we discuss a step-by-step methodology for dissecting the mouse brain into sixteen distinct regions. In this procedure, we have focused on male mouse C57Bl/6J (6-8 week old) brain removal and dissection into multiple regions using neuroanatomical landmarks to identify and sample discrete functionally-relevant and behaviorally-relevant brain regions. This work will help lay a strong foundation in the field of neuroscience, leading to more focused approaches in the deeper understanding of brain function.

Acute and chronic effects of exposure to the potent organophosphate acetylcholinesterase inhibitor soman are an increasing concern for military and civilian populations. High dose exposure results in increased levels of acetylcholine and cholinergic crisis. This cholinergic crisis induces convulsions and seizures and can result in death if left untreated. Some individuals do not experience seizures on high dose exposure although they experience long‐term neurological injury. Understanding the downstream effects of soman at the molecular level is vital to developing a methods for prognosis and treatment and for explaining inter‐individual differences in response. We performed an integrated analysis of miRNA and mRNA expression in brain regions associated with soman exposure and resultant seizure activity in rats. Adult male Sprague‐Dawley rats were exposed subcutaneously to soman. Samples from seizing and non‐seizing animals were collected from the heart, kidney, liver, lungs, spleen, and brain, including the amygdala, hippocampus, hypothalamus, piriform, medial prefrontal cortex, parietal cortex, and thalamus at 72 hrs and 90 days following exposure. Initial data from the piriform cortex demonstrates significant differences in expression patterns between animals that seized and those that did not. The observed miRNA expression differences across brain tissue types indicate the hypothalamus has a distinct response from other regions. Functional annotation analysis of miRNA‐mRNA interaction pairs identified many that are likely to be involved in the seizing response.Ongoing work will fully characterize the gene networks and pathways altered by soman exposure and associated with soman‐induced seizure responses across different brain regions and should lead to useful assays for diagnosis, prognosis, or exposure surveillance.Disclaimers:Research was conducted in compliance with the Animal Welfare Act, and all other Federal requirements. The views expressed are those of the authors and do not constitute endorsement by the U.S. Army.Support or Funding InformationSupport was provided by interagency agreements between BARDA, the Geneva Foundation (ARO agreement no. W911NF‐13‐1‐0376), and the U.S. Army Medical Research Institute of Chemical Defense (USAMRICD) as well as a memorandum of agreement between USAMRICD and the U.S. Army Center of Environmental Health (USACEHR).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

A social-stress mouse model was used to simulate features of post-traumatic stress disorder (PTSD). The model involved exposure of an intruder (male C57BL/6) mouse to a resident aggressor (male SJL) mouse for 5 or 10 consecutive days. Transcriptome changes in brain regions (hippocampus, amygdala, medial prefrontal cortex and hemibrain), blood and spleen as well as epigenome changes in the hemibrain were assayed after 1- and 10-day intervals following the 5-day trauma or after 1- and 42-day intervals following the 10-day trauma. Analyses of differentially expressed genes (common among brain, blood and spleen) and differentially methylated promoter regions revealed that neurogenesis and synaptic plasticity pathways were activated during the early responses but were inhibited after the later post-trauma intervals. However, inflammatory pathways were activated throughout the observation periods, except in the amygdala in which they were inhibited only at the later post-trauma intervals. Phenotypically, inhibition of neurogenesis was corroborated by impaired Y-maze behavioral responses. Sustained neuroinflammation appears to drive the development and maintenance of behavioral manifestations of PTSD, potentially via its inhibitory effect on neurogenesis and synaptic plasticity. By contrast, peripheral inflammation seems to be directly responsible for tissue damage underpinning somatic comorbid pathologies. Identification of overlapping, differentially regulated genes and pathways between blood and brain suggests that blood could be a useful and accessible brain surrogate specimen for clinical translation.

The term seizure refers to certain physical or behavioral actions that may occur after an episode of abnormal electrical activity in the brain. Epileptic seizure, one of the major seizure types, affects about 1% of the population currently and has affected about 4% of the population at some point in time. Most of those affected by seizures (nearly 80%) live in developing countries. Soman, also known as GD, is an extremely toxic man‐made organophosphorous chemical. Exposure to GD frequently results in seizures, as well as neuropathological and neurochemical abnormalities in various regions of the rat brain. Small microRNAs (miRNA) are (~22 nt) non‐coding RNAs that regulate the expression of target mRNAs at the post‐transcriptional level. Experimental evidence has demonstrated that miRNAs are involved in heterogeneous brain functions, including neuro‐inflammation, synaptic remodeling, and neuronal death. The purpose of the current study was to search potential therapeutic targets and biomarkers for seizure by sequencing genome‐wide small RNA and identifying differentially expressed (DE) miRNA of the rat brain following exposure to GD. Rats were exposed to either 0.8 LD 50 GD or 1.0 LD 50 GD versus a saline control group and then evaluated for seizure activity with EEG and also visually. We conducted miRNA profiling studies in brain tissues (hippocampus‐HP, hypothalamus‐HT, piriform cortex‐PIR, thalamus‐TH, amygdala‐AY and medial prefrontal cortex‐mPFC) collected from rats exposed to GD at 72 hours post‐exposure. Principal component analysis indicated good separation in relation to seizure activity. The principal component analysis showed two distinct clusters as per expression profile for all brain regions and AY, PIR and mPFC brain region were clustered together than the other brain tissues. To identify the expression profile of significantly different miRNAs in these brain regions, we performed the moderated t‐test to call differentially expressed (DE) miRNAs. The DE analysis showed total of 276 significant DE miRNAs in rat brain (87 in AY, 166 in PIR and 23 in mPFC) at the 1.0 LD 50 of GD exposure concentrations as compared to the control group. After comparison of the seizure activity group to “none” seizure activity (DNS) group, we observed a total of 533 DE miRNAs (123 in AY, 58 in HP, 147 in HT, 249 in PIR, 185 in TH and 19 in mPFC) at 1 LD 50 and 0.8 LD 50 GD exposures. In conclusion, we identified changes in brain miRNAs (DE miRNA) that clearly distinguish seizures in rats from the control group, “none” seizure activity group and seizure group. These alterations in DE miRNA expression correlate with neuropathological progression and, potentially, with response to experimental treatment. This study provides potential therapeutic targets and biomarkers for the control of seizures. In addition, these data provide important insights into the molecular mechanisms involved in soman‐induced seizure activity and a basis for generating hypotheses about the mechanisms of seizure.

Post-traumatic stress disorder (PTSD) is a debilitating illness that imposes significant emotional and financial burdens on military families. The understanding of PTSD etiology remains elusive; nonetheless, it is clear that PTSD is manifested by a cluster of symptoms including hyperarousal, reexperiencing of traumatic events, and avoidance of trauma reminders. With these characteristics in mind, several rodent models have been developed eliciting PTSD-like features. Animal models with social dimensions are of particular interest, since the social context plays a major role in the development and manifestation of PTSD.For civilians, a core trauma that elicits PTSD might be characterized by a singular life-threatening event such as a car accident. In contrast, among war veterans, PTSD might be triggered by repeated threats and a cumulative psychological burden that coalesced in the combat zone. In capturing this fundamental difference, the aggressor-exposed social stress (Agg-E SS) model imposes highly threatening conspecific trauma on naïve mice repeatedly and randomly.There is abundant evidence that suggests the potential role of genetic contributions to risk factors for PTSD. Specific observations include putatively heritable attributes of the disorder, the cited cases of atypical brain morphology, and the observed neuroendocrine shifts away from normative. Taken together, these features underscore the importance of multi-omics investigations to develop a comprehensive picture. More daunting will be the task of downstream analysis with integration of these heterogeneous genotypic and phenotypic data types to deliver putative clinical biomarkers. Researchers are advocating for a systems biology approach, which has demonstrated an increasingly robust potential for integrating multidisciplinary data. By applying a systems biology approach here, we have connected the tissue-specific molecular perturbations to the behaviors displayed by mice subjected to Agg-E SS. A molecular pattern that links the atypical fear plasticity to energy deficiency was thereby identified to be causally associated with many behavioral shifts and transformations.PTSD is a multifactorial illness sensitive to environmental influence. Accordingly, it is essential to employ the optimal animal model approximating the environmental condition that elicits PTSD-like symptoms. Integration of an optimal animal model with a systems biology approach can contribute to a more knowledge-driven and efficient next-generation care management system and, potentially, prevention of PTSD.

Background: PCP use is prevalent in Washington DC. There has been fluctuation in PCP use in DC areas over time; from the 1980s to 1990s the use of PCP in DC area decreased. Unfortunately, the trend has been increasing. 10% of arrested adults in DC has active PCP urine toxicology screen. Howard University is a unique institution serving mostly underserved African American population. In the current study, we sought to identify the correlates of PCP use in our patient population. Methods: We obtained data from QI Project that psychiatry resident conduct continuously at Howard University Hospital. We included patients admitted to the inpatient psychiatric services. Of the 1241 patients, we included 132 patients. We reviewed the charts and laboratory workup including UDS results. Tabaco and synthetic cannabinoid use were self-reported. We used chi-square to analyze the association between PCP use and other variables and used linear regression to correct for confounding factors. Results: Of 132 patients, 40% are males and 7% of the patients has UDS positive for PCP. Our analysis showed that PCP use was positively associated with cannabis and cocaine use (P,0.02 and p,0.01 respectively). Regression model taking in account other substances and gender, the relationship between PCP use remained statistically significant. Conclusions: Use of PCP has been unfortunately consistently higher in DC compared to the rest of the US. In our previous study, cocaine predicts poor outcome for patients maintained in buprenorphine treatment. These results indicate that treatment strategies addressing substance use in DC should take in account co-occurring substance use pattern.

Soman (GD) is a highly toxic man‐made substance and a potential chemical weapon. It belongs to a class of chemicals known as organophosphorus (OP) nerve agents, which may cause seizures, respiratory failure, and death. Soman undergoes aging and is one of the more difficult nerve agent exposures to treat since field available oxime therapy is not effective against soman. Understanding the effects of soman is vital to developing a method of detection and treatment for exposure. Exposure to GD can consequently result in neuropathological abnormalities in various regions of the brain, including the hippocampus, piriform cortex, amygdala, thalamus, and hypothalamus. In this study, rats were exposed to either 0.8 LD50 GD or 1.0 LD50 GD versus a saline control group and evaluated for seizure activity. Rats were sacrificed and brain regions were harvested at 72 hours post‐exposure. Brain tissues were homogenized and RNA was isolated using TRIzol reagent. In order to explore the pathways and biological processes associated with GD exposure, transcripts in the aforementioned brain regions were assayed using Agilent's rat gene expression arrays. Detailed analysis is currently underway to compare the expression profile generated from different regions of the brain and to understand the differences in response to the variation in the concentration of GD exposure.DISCLAIMER “Research was conducted in compliance with the Animal Welfare Act and all other Federal requirements. The views expressed are those of the authors and do not constitute endorsement by the U.S. Army.”

Soman, or GD (systematic name: O ‐pinacolyl methylphosphonofluoridate), is an extremely toxic man‐made chemical substance. It belongs to a class of chemicals known as organophosphorus (OP) nerve agents, which may cause seizures, respiratory failure, and death. GD interferes with normal functioning of the mammalian nervous system by inhibiting the cholinesterase enzyme. Exposure to GD can consequently result in neuropathological abnormalities in various regions of the rat brain. Small microRNAs are (~22 nt) non‐coding RNAs that regulate the expression of target mRNAs at the post‐transcriptional level. Experimental evidence has demonstrated that miRNAs are involved in heterogeneous brain functions including neuro‐inflammation, synaptic remodeling, and neuronal death. The purpose of the current study was to investigate if exposure to soman results in significant molecular changes in different brain tissues on the miRNA level. The experiments were conducted at the U.S. Army Medical Research Institute of Chemical Defense where rats were exposed to either 0.8 LD 50 GD or 1.0 LD 50 GD versus a saline control group and then evaluated for seizure activity. We carried out miRNA profiling studies in brain tissues (hippocampus, hypothalamus, piriform, thalamus, amygdala and medial prefrontal cortex) collected from rats exposed to GD at 72 hours post‐exposure. Compared to the control group, there were 74 differentially expressed (DE) miRNAs (54 downregulated and 20 upregulated) in the hypothalamus at a dose of 0.8 LD 50 of GD exposure. In addition, we found 105 DE miRNAs (62 downregulated and 43 upregulated) in the hypothalamus at a dose of 1.0 LD 50 of GD exposure when compared to the control group. We used false discovery rate (FDR) correction for DE miRNAs data and then filtered for experimentally validated miRNAs in ingenuity pathway analysis (IPA). We observed 12 miRNA with 357 gene targets, some of which play roles in neurotransmitter and nervous system signaling pathways. IPA revealed 31 miRNAs passing the experimental validated filter with 591 gene targets. Twenty‐eight miRNAs of these target 101 genes were involved in neurological pathways. Among the pathways targeted by differentially expressed miRNAs are: IL‐8 signaling, axonal guidance signaling, NF‐kB signaling, PTEN signaling and B‐cell receptor signaling. Among top toxicity pathways, aryl‐hydro carbon receptor was highly activated. Changes in miRNAs expression profile of other brain regions were also observed and will be discussed. These data provide important insights into the molecular mechanisms involved in soman‐induced neuropathology and a basis for generating hypotheses about the mechanisms of soman‐induced brain damage. Comprehensive investigations are currently underway to further elucidate the exposure related changes in different tissues of the brain. Support or Funding Information DISCLAIMERS: Research was conducted in compliance with the Animal Welfare Act and all other Federal Requirements. The views expressed are those of the authors and do not constitute endorsement by the U.S. Army.

We collected hemi‐brain, hippocampus, amygdala, medial prefrontal cortex and ventral striatum from mice exposed to modified social‐stress for 1 to 12 days. We performed transcriptome, DNA‐methylome, miRNAome, and genome‐wide Creb1 transcription binding assays. Temporal and integrative analyses of the multi‐omics datasets were carried out using time‐dependent features extraction, and combinatorial algorithms. Our analyses showed unexpected shifts in relational time coordinates among the different omics datasets. Specifically, for a given gene or probe with significantly changed multi‐omics data points, we observed a temporal shift in its expression changes, methylation of its promoter, modulation of its transcript by upstream miRNAs, and binding of its promoter by transcription regulator. For many significantly changed features, the methylation changes occurred, unexpectedly, at earlier stress days while gene expression changes occurred at later time points. Many of the miRNA changes occurred at eight and later stress days. Our findings showed importance of temporal assays to capture time‐dependent multi‐omics changes as reliable signatures for a given condition. Integrative analyses of multi‐omics measurements based on a single time point may be deficient in many aspects, and may also be misleading if missed observations are interpreted as absence of changes.Disclaimers: Research was conducted in compliance with the Animal Welfare Act, and all other Federal requirements. The views expressed are those of the author(s) and do not constitute endorsement by the U.S. Army.Support or Funding InformationFunding: USAMRMC grant number 09284002

Systematically distinguishing genetic liability from other contributing factors is critical for designing a preventive strategy for post-traumatic stress disorder (PTSD). To address this issue, we investigated a murine model exposing C57BL/6j, DBA/2j and BALB/cj mice to repeated stress via exposure to conspecific aggressors (Agg-E). Naïve mice from each strain were subjected to the proximity of aggressor (Agg) mice for 6h using a 'cage-within-a-cage' paradigm, which was repeated for 5 or 10 days with intermittent and unpredictable direct contact with Agg mice. During the Agg-E stress, DBA/2j developed a different strategy to evade Agg mice, which potentially contributed to its phenotypic resilience to Agg-E stress. Although Agg mice inflicted C57BL/6j and BALB/cj with equivalent numbers of strikes, BALB/cj displayed a distinct behavioral phenotype with delayed exhibition of a number of PTSD-like features. By contrast, C57BL/6j mice displayed unique vulnerability to Agg-E stress induced myocardopathy, possibly attributable to their particular susceptibility to hypoxia. A group of genes (Bdnf, Ngf, Zwint, Cckbr, Slc6a4, Fkbp5) linked to PTSD and synaptic plasticity were significantly altered in C57BL/6j and BALB/cj Agg-E mice. Contributions of Agg-E stress history and genotypic heterogeneity emerged as the key mediators of PTSD-like features. Linking genetic components to specific phenotypic and pathological features could have potential clinical implications.

Post-traumatic stress disorder (PTSD) in humans and PTSD-like disorders in animal models tend to comorbid with inflammation, cardiovascular and metabolic disorders. Inflammation is implicated as the cause of chronic pain in PTSD patients. We assessed changes in the expression of genes known to mediate inflammation. We meta-analyzed our in-house generated data acquired from blood of PTSD patients, blood and brain parts of mouse models of PTSD, and similar data sets downloaded from public repositories. We did time-line stratifications of the metadata to identify molecular indicators of chronological severity of inflammation accompanying pathologic progression of PTSD. Expression levels of genes mediating inflammation significantly increased at longer time points; in mice 6 weeks post social-stress sessions, in rats after weeks of predator stress, in non-human primates in accordance with chronic stressors of social hierarchies, and in humans, in correlation with chronic nature of PTSD symptoms. Regulators of inflammatory molecules (transcription factors, epigenetic regulators and microRNAs) also indicated increased severity of inflammation with prolonged PTSD symptoms (longer trauma memories). Our findings suggest that intervention of chronic PTSD should include ways of mitigating inflammation to alleviate chronic pain of PTSD patients. We thank the grant support of USAMRMC NO: 09284002. Research was conducted in compliance with all Federal requirements.

BACKGROUND:Social-stress mouse model, based on the resident-intruder paradigm was used to simulate features of human post-traumatic stress disorder (PTSD). The model involved exposure of an intruder (subject) mouse to a resident aggressor mouse followed by exposure to trauma reminders with rest periods. C57BL/6 mice exposed to SJL aggressor mice exhibited behaviors suggested as PTSD-in-mouse phenotypes: intermittent freezing, reduced locomotion, avoidance of the aggressor-associated cue and apparent startled jumping. Brain tissues (amygdala, hippocampus, medial prefrontal cortex, septal region, corpus striatum and ventral striatum) from subject (aggressor exposed: Agg-E) and control C57BL/6 mice were collected at one, 10 and 42 days post aggressor exposure sessions. Transcripts in these brain regions were assayed using Agilent's mouse genome-wide arrays.RESULTS:Pathways and biological processes associated with differentially regulated genes were mainly those thought to be involved in fear-related behavioral responses and neuronal signaling. Expression-based assessments of activation patterns showed increased activations of pathways related to anxiety disorders (hyperactivity and fear responses), impaired cognition, mood disorders, circadian rhythm disruption, and impaired territorial and aggressive behaviors. In amygdala, activations of these pathways were more pronounced at earlier time-points, with some attenuation after longer rest periods. In hippocampus and medial prefrontal cortex, activation patterns were observed at later time points. Signaling pathways associated with PTSD-comorbid conditions, such as diabetes, metabolic disorder, inflammation and cardiac infarction, were also significantly enriched. In contrast, signaling processes related to neurogenesis and synaptic plasticity were inhibited.CONCLUSIONS:Our data suggests activations of behavioral responses associated with anxiety disorders as well as inhibition of neuronal signaling pathways important for neurogenesis, cognition and extinction of fear memory. These pathways along with comorbid-related signaling pathways indicate the pervasive and multisystem effects of aggressor exposure in mice, potentially mirroring the pathologic conditions of PTSD patients.

Acute responses to intense stressors can give rise to post-traumatic stress disorder (PTSD). PTSD diagnostic criteria include trauma exposure history and self-reported symptoms. Individuals who meet PTSD diagnostic criteria often meet criteria for additional psychiatric diagnoses. Biomarkers promise to contribute to reliable phenotypes of PTSD and comorbidities by linking biological system alterations to behavioral symptoms. Here we have analyzed unbiased plasma metabolomics and other stress effects in a mouse model with behavioral features of PTSD. In this model, C57BL/6 mice are repeatedly exposed to a trained aggressor mouse (albino SJL) using a modified, resident-intruder, social defeat paradigm. Our recent studies using this model found that aggressor-exposed mice exhibited acute stress effects including changed behaviors, body weight gain, increased body temperature, as well as inflammatory and fibrotic histopathologies and transcriptomic changes of heart tissue. Some of these acute stress effects persisted, reminiscent of PTSD. Here we report elevated proteins in plasma that function in inflammation and responses to oxidative stress and damaged tissue at 24 hrs post-stressor. Additionally at this acute time point, transcriptomic analysis indicated liver inflammation. The unbiased metabolomics analysis showed altered metabolites in plasma at 24 hrs that only partially normalized toward control levels after stress-withdrawal for 1.5 or 4 wks. In particular, gut-derived metabolites were altered at 24 hrs post-stressor and remained altered up to 4 wks after stress-withdrawal. Also at the 4 wk time point, hyperlipidemia and suppressed metabolites of amino acids and carbohydrates in plasma coincided with transcriptomic indicators of altered liver metabolism (activated xenobiotic and lipid metabolism). Collectively, these system-wide sequelae to repeated intense stress suggest that the simultaneous perturbed functioning of multiple organ systems (e.g., brain, heart, intestine and liver) can interact to produce injuries that lead to chronic metabolic changes and disorders that have been associated with PTSD.

To gain insights into the toxicity induced by the nerve agent VX, an MS-based phosphoproteomic analysis was carried out on the piriform cortex region of brains from VX-treated rats. Using isobaric tag based TMT labeling followed by titanium dioxide enrichment strategy, we identified 9975 unique phosphosites derived from 3287 phosphoproteins. Temporal changes in the phosphorylation status of peptides were observed over a time period of 24 h in rats exposed to a 1× LD50, intravenous (i.v.) dose with the most notable changes occurring at the 1 h postexposure time point. Five major functional classes of proteins exhibited changes in their phosphorylation status: (i) ion channels/transporters, including ATPases, (ii) kinases/phosphatases, (iii) GTPases, (iv) structural proteins, and (v) transcriptional regulatory proteins. This study is the first quantitative phosphoproteomic analysis of VX toxicity in the brain. Understanding the toxicity and compensatory signaling mechanisms will improve the understanding of the complex toxicity of VX in the brain and aid in the elucidation of novel molecular targets that would be important for development of improved countermeasures. All MS data have been deposited in the ProteomeXchange with identifier PXD001184 (http://proteomecentral.proteomexchange.org/dataset/PXD001184).

The health benefits of fish oil enriched with high omega-3 polyunsaturated fatty acids (n-3 PUFA) are widely documented. Fish oil as dietary supplements, however, show moderate clinical efficacy, highlighting an immediate scope of systematic in vitro feedback. Our transcriptomic study was designed to investigate the genomic shift of murine brains fed on fish oil enriched diets. A customized fish oil enriched diet (FD) and standard lab diet (SD) were separately administered to two randomly chosen populations of C57BL/6J mice from their weaning age until late adolescence. Statistical analysis mined 1,142 genes of interest (GOI) differentially altered in the hemibrains collected from the FD- and SD-fed mice at the age of five months. The majority of identified GOI (∼40%) encodes proteins located in the plasma membrane, suggesting that fish oil primarily facilitated the membrane-oriented biofunctions. FD potentially augmented the nervous system's development and functions by selectively stimulating the Src-mediated calcium-induced growth cascade and the downstream PI3K-AKT-PKC pathways. FD reduced the amyloidal burden, attenuated oxidative stress, and assisted in somatostatin activation—the signatures of attenuation of Alzheimer's disease, Parkinson's disease, and affective disorder. FD induced elevation of FKBP5 and suppression of BDNF, which are often linked with the improvement of anxiety disorder, depression, and post-traumatic stress disorder. Hence we anticipate efficacy of FD in treating illnesses such as depression that are typically triggered by the hypoactivities of dopaminergic, adrenergic, cholinergic, and GABAergic networks. Contrastingly, FD's efficacy could be compromised in treating illnesses such as bipolar disorder and schizophrenia, which are triggered by hyperactivities of the same set of neuromodulators. A more comprehensive investigation is recommended to elucidate the implications of fish oil on disease pathomechanisms, and the result-driven repositioning of fish oil utilization may revitalize its therapeutic efficacy.

A utility criterion of a PTSD mouse model is recruiting a stressor with systematically variable intensity. An ideal stressor should maintain a ‘dose‐response relationship’ with the subjects’ behavioral shift. A typical PTSD model includes a brief exposure to foot or tail shock.We developed a model that involves repeated exposures (either 5‐day or 10‐day) to trained conspecific aggressors (Agg). A contextual reminder study evaluating a cluster of ethogram identified PTSD‐like (acute‐ and persistent‐) syndromes such as incubation, extinction and desensitization of fear responses. Pathophysiological consequences supported the model.This model reflects the combat‐like situation where life‐threatening events occur repeatedly and randomly. The direct relationship of PTSD‐risk with deployment frequency further justifies the model.Agg induced stress intensity can only be elevated by prolonging the exposure. The risk of habituation can thereby defeat the ‘dose‐response’ relationship. We addressed this concern by evaluating the time‐dependent behavioral shift of the subject mice (C57BL/6j). Subsets of subject mice were withdrawn from the Agg‐exposure (Agg‐E) schedule at regular intervals and their psycho‐patho‐physiological characters were evaluated. A regression model elucidated the temporal relationship of Agg‐E stress with the psychological alteration.