Circulating myeloid-derived MMP8 in stress susceptibility and depression.

Psychosocial stress has profound effects on the body, including the immune system and the brain1,2. Although a large number of pre-clinical and clinical studies have linked peripheral immune system alterations to stress-related disorders such as major depressive disorder (MDD)3, the underlying mechanisms are not well understood. Here we show that expression of a circulating myeloid cell-specific proteinase, matrix metalloproteinase 8 (MMP8), is increased in the serum of humans with MDD as well as in stress-susceptible mice following chronic social defeat stress (CSDS). In mice, we show that this increase leads to alterations in extracellular space and neurophysiological changes in the nucleus accumbens (NAc), as well as altered social behaviour. Using a combination of mass cytometry and single-cell RNA sequencing, we performed high-dimensional phenotyping of immune cells in circulation and in the brain and demonstrate that peripheral monocytes are strongly affected by stress. In stress-susceptible mice, both circulating monocytes and monocytes that traffic to the brain showed increased Mmp8 expression following chronic social defeat stress. We further demonstrate that circulating MMP8 directly infiltrates the NAc parenchyma and controls the ultrastructure of the extracellular space. Depleting MMP8 prevented stress-induced social avoidance behaviour and alterations in NAc neurophysiology and extracellular space. Collectively, these data establish a mechanism by which peripheral immune factors can affect central nervous system function and behaviour in the context of stress. Targeting specific peripheral immune cell-derived matrix metalloproteinases could constitute novel therapeutic targets for stress-related neuropsychiatric disorders.

Circulating senescent myeloid cells infiltrate the brain and cause neurodegeneration in histiocytic disorders.

Neurodegenerative diseases (ND) are characterized by progressive loss of neuronal function. Mechanisms of ND pathogenesis are incompletely understood, hampering the development of effective therapies. Langerhans cell histiocytosis (LCH) is an inflammatory neoplastic disorder caused by hematopoietic progenitors expressing mitogen-activated protein kinase (MAPK)-activating mutations that differentiate into senescent myeloid cells that drive lesion formation. Some individuals with LCH subsequently develop progressive and incurable neurodegeneration (LCH-ND). Here, we showed that LCH-ND was caused by myeloid cells that were clonal with peripheral LCH cells. Circulating BRAFV600E+ myeloid cells caused the breakdown of the blood-brain barrier (BBB), enhancing migration into the brain parenchyma where they differentiated into senescent, inflammatory CD11a+ macrophages that accumulated in the brainstem and cerebellum. Blocking MAPK activity and senescence programs reduced peripheral inflammation, brain parenchymal infiltration, neuroinflammation, neuronal damage and improved neurological outcome in preclinical LCH-ND. MAPK activation and senescence programs in circulating myeloid cells represent targetable mechanisms of LCH-ND.

Stress-activated brain-gut circuits disrupt intestinal barrier integrity and social behaviour.

Chronic stress underlies the etiology of both major depressive disorder (MDD) and irritable bowel syndrome (IBS), two highly prevalent and debilitating conditions with high rates of co-morbidity. However, it is not fully understood how the brain and gut bi-directionally communicate during stress to impact intestinal homeostasis and stress-relevant behaviours. Using the chronic social defeat stress (CSDS) model, we find that stressed mice display greater intestinal permeability and circulating levels of the endotoxin lipopolysaccharide (LPS) compared to unstressed control (CON) mice. Interestingly, the microbiota in the colon also exhibit elevated LPS biosynthesis gene expression following CSDS. Additionally, CSDS triggers an increase in pro-inflammatory colonic IFNγ+ Th1 cells and a decrease in IL4+ Th2 cells compared to CON mice, and this gut inflammation contributes to stress-induced intestinal barrier permeability and social avoidance behaviour. We next investigated the role of enteric neurons and identified that noradrenergic dopamine beta-hydroxylase (DBH)+ neurons in the colon are activated by CSDS, and that their ablation protects against gut pathophysiology and disturbances in social behaviour. Retrograde tracing from the colon identified a population of corticotropin-releasing hormone-expressing (CRH+) neurons in the paraventricular nucleus of the hypothalamus (PVH) that innervate the colon and are activated by stress. Chemogenetically activating these PVH CRH+ neurons is sufficient to induce gut inflammation, barrier permeability, and social avoidance behaviour, while inhibiting these cells prevents these effects following exposure to CSDS. Thus, we define a stress-activated brain-to-gut circuit that confers colonic inflammation, leading to impaired intestinal barrier function, and consequent behavioural deficits.

Chemokine receptor 5 signaling in PFC mediates stress susceptibility in female mice.

Chronic stress induces changes in the periphery and the central nervous system (CNS) that contribute to neuropathology and behavioral abnormalities associated with psychiatric disorders. In this study, we examined the impact of peripheral and central inflammation during chronic social defeat stress (CSDS) in female mice. Compared to male mice, we found that female mice exhibited heightened peripheral inflammatory response and identified C-C motif chemokine ligand 5 (CCL5), as a stress-susceptibility marker in females. Blocking CCL5 signaling in the periphery promoted resilience to CSDS. In the brain, stress-susceptible mice displayed increased expression of C-C chemokine receptor 5 (CCR5), a receptor for CCL5, in microglia in the prefrontal cortex (PFC). This upregulation was associated with microglia morphological changes, their increased migration to the blood vessels, and enhanced phagocytosis of synaptic components and vascular material. These changes coincided with neurophysiological alterations and impaired blood-brain barrier (BBB) integrity. By blocking CCR5 signaling specifically in the PFC were able to prevent stress-induced physiological changes and rescue social avoidance behavior. Our findings are the first to demonstrate that stress-mediated dysregulation of the CCL5-CCR5 axis triggers excessive phagocytosis of synaptic materials and neurovascular components by microglia, resulting in disruptions in neurotransmission, reduced BBB integrity, and increased stress susceptibility. Our study provides new insights into the role of cortical microglia in female stress susceptibility and suggests that the CCL5-CCR5 axis may serve as a novel sex-specific therapeutic target for treating psychiatric disorders in females.

A critical role for cortical amygdala circuitry in shaping social encounters.

Aggression is an evolutionarily conserved behavior that controls social hierarchies and protects valuable resources like mates, food, and territory. In mice, aggressive behaviour can be broken down into an appetitive phase, which involves approach and investigation, and a consummatory phase, which involves biting, kicking, and wrestling. By performing an unsupervised weighted correlation network analysis on whole-brain c-Fos expression, we identified a cluster of brain regions including hypothalamic and amygdalar sub-regions and olfactory cortical regions highly co-activated in male, but not female aggressors (AGG). The posterolateral cortical amygdala (COApl), an extended olfactory structure, was found to be a hub region based on the number and strength of correlations with other regions in the cluster. Our data further show that estrogen receptor 1 (ESR1)-expressing cells in the COApl exhibit increased activity during attack behaviour, and during bouts of investigation which precede an attack, in male mice only. Chemogenetic or optogenetic inhibition of COApl ESR1 cells in AGG males reduces aggression and increases pro-social investigation without affecting social reward/reinforcement behavior. We further confirmed that COApl ESR1 projections to the ventrolateral portion of the ventromedial hypothalamus and central amygdala are necessary for these behaviours. Collectively, these data suggest that in aggressive males, COApl ESR1 cells respond specifically to social stimuli, thereby enhancing their salience and promoting attack behaviour.

Peripheral immune-derived matrix metalloproteinase promotes stress susceptibility.

Psychosocial stress has profound effects on the body, including the peripheral immune system and the brain1,2. Although a large number of pre-clinical and clinical studies have linked peripheral immune system alterations to stress-related disorders such as major depressive disorder (MDD)3,4,5, the underlying mechanisms are not well understood. Here we show that a peripheral myeloid cell-specific proteinase, matrix metalloproteinase 8 (MMP8), is elevated in serum of subjects with MDD as well as in stress-susceptible (SUS) mice following chronic social defeat stress (CSDS). In mice, we show that this increase leads to alterations in extracellular space and neurophysiological changes in the nucleus accumbens (NAc), thereby altering social behaviour. Using a combination of mass cytometry and single-cell RNA-sequencing, we performed high-dimensional phenotyping of immune cells in circulation and brain and demonstrate that peripheral monocytes are strongly affected by stress. Both peripheral and brain-infiltrating monocytes of SUS mice showed increased Mmp8 expression following CSDS. We further demonstrate that peripheral MMP8 directly infiltrates the NAc parenchyma to control the ultrastructure of the extracellular space. Depleting MMP8 prevented stress-induced social avoidance behaviour and alterations in NAc neurophysiology and extracellular space. Collectively, these data establish a novel mechanism by which peripheral immune factors can affect central nervous system function and behaviour in the context of stress. Targeting specific peripheral immune cell-derived matrix metalloproteinases could constitute novel therapeutic targets for stress-related neuropsychiatric disorders.

Social trauma engages lateral septum circuitry to occlude social reward.

In humans, traumatic social experiences can contribute to psychiatric disorders1. It is suggested that social trauma impairs brain reward function such that social behaviour is no longer rewarding, leading to severe social avoidance2,3. In rodents, the chronic social defeat stress (CSDS) model has been used to understand the neurobiology underlying stress susceptibility versus resilience following social trauma, yet little is known regarding its impact on social reward4,5. Here we show that, following CSDS, a subset of male and female mice, termed susceptible (SUS), avoid social interaction with non-aggressive, same-sex juvenile C57BL/6J mice and do not develop context-dependent social reward following encounters with them. Non-social stressors have no effect on social reward in either sex. Next, using whole-brain Fos mapping, in vivo Ca2+ imaging and whole-cell recordings, we identified a population of stress/threat-responsive lateral septum neurotensin (NTLS) neurons that are activated by juvenile social interactions only in SUS mice, but not in resilient or unstressed control mice. Optogenetic or chemogenetic manipulation of NTLS neurons and their downstream connections modulates social interaction and social reward. Together, these data suggest that previously rewarding social targets are possibly perceived as social threats in SUS mice, resulting from hyperactive NTLS neurons that occlude social reward processing.

Sex differences in appetitive and reactive aggression.

Aggression is an evolutionarily conserved, adaptive component of social behavior. Studies in male mice illustrate that aggression is influenced by numerous factors including the degree to which an individual finds aggression rewarding and will work for access to attack and subordinate mice. While such studies have expanded our understanding of the molecular and circuit mechanisms of male aggression very little is known about female aggression, within these established contexts. Here we use an ethologically relevant model of male vs. female aggression by pair housing adult male and female outbred CFW mice with opposite sex cage mates. We assess reactive (defensive) aggression in the resident intruder (RI) test and appetitive (rewarding) aggression in the aggression conditioned place preference (CPP) and operant self-administration (SA) tests. Our results show dramatic sex differences in both qualitative and quantitative aspects of reactive vs. appetitive aggression. Males exhibit more wrestling and less investigative behavior during RI, find aggression rewarding, and will work for access to a subordinate to attack. Females exhibit more bites, alternate between aggressive behaviors and investigative behaviors more readily during RI, however, they do not find aggression to be rewarding or reinforcing. These results establish sex differences in aggression in mice, providing an important resource for the field to better understand the circuit and molecular mechanisms of aggression in both sexes.

Neuromodulatory effect of interleukin 1ß in the dorsal raphe nucleus on individual differences in aggression.

Heightened aggressive behavior is considered as one of the central symptoms of many neuropsychiatric disorders including autism, schizophrenia, and dementia. The consequences of aggression pose a heavy burden on patients and their families and clinicians. Unfortunately, we have limited treatment options for aggression and lack mechanistic insight into the causes of aggression needed to inform new efforts in drug discovery and development. Levels of proinflammatory cytokines in the periphery or cerebrospinal fluid were previously reported to correlate with aggressive traits in humans. However, it is still unknown whether cytokines affect brain circuits to modulate aggression. Here, we examined the functional role of interleukin 1ß (IL-1ß) in mediating individual differences in aggression using a resident-intruder mouse model. We found that nonaggressive mice exhibit higher levels of IL-1ß in the dorsal raphe nucleus (DRN), the major source of forebrain serotonin (5-HT), compared to aggressive mice. We then examined the effect of pharmacological antagonism and viral-mediated gene knockdown of the receptors for IL-1 within the DRN and found that both treatments consistently increased aggressive behavior of male mice. Aggressive mice also exhibited higher c-Fos expression in 5-HT neurons in the DRN compared to nonaggressive mice. In line with these findings, deletion of IL-1 receptor in the DRN enhanced c-Fos expression in 5-HT neurons during aggressive encounters, suggesting that modulation of 5-HT neuronal activity by IL-1ß signaling in the DRN controls expression of aggressive behavior.

Region-specific myelin differences define behavioral consequences of chronic social defeat stress in mice.

Exposure to stress increases the risk of developing mood disorders. While a subset of individuals displays vulnerability to stress, others remain resilient, but the molecular basis for these behavioral differences is not well understood. Using a model of chronic social defeat stress, we identified region-specific differences in myelination between mice that displayed social avoidance behavior ('susceptible') and those who escaped the deleterious effect to stress ('resilient'). Myelin protein content in the nucleus accumbens was reduced in all mice exposed to stress, whereas decreased myelin thickness and internodal length were detected only in the medial prefrontal cortex (mPFC) of susceptible mice, with fewer mature oligodendrocytes and decreased heterochromatic histone marks. Focal demyelination in the mPFC was sufficient to decrease social preference, which was restored following new myelin formation. Together these data highlight the functional role of mPFC myelination as critical determinant of the avoidance response to traumatic social experiences. Editorial note: This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that all the issues have been addressed (see decision letter).

A diet enriched with curcumin promotes resilience to chronic social defeat stress.

Chronic exposure to stress is a well-known risk factor for the development of mood and anxiety disorders. Promoting resilience to stress may prevent the development of these disorders, but resilience-enhancing compounds are not yet clinically available. One compound that has shown promise in the clinical setting is curcumin, a polyphenol compound found in the rhizome of the turmeric plant (Curcuma longa) with known anti-inflammatory and antidepressant properties. Here, we tested the efficacy of 1.5% dietary curcumin at promoting resilience to chronic social defeat stress (CSDS) in 129/SvEv mice, a strain that we show is highly susceptible to this type of stress. We found that administration of curcumin during CSDS produced a 4.5-fold increase in stress resilience, as measured by the social interaction test. Although the overall effects of curcumin were striking, we identified two distinct responses to curcumin. While 64% of defeated mice on curcumin were resilient (responders), the remaining 36% of mice were susceptible to the effects of stress (non-responders). Interestingly, responders released less corticosterone following acute restraint stress and had lower levels of peripheral IL-6 than nonresponders, implicating a role for the NF-κB pathway in treatment response. Importantly, curcumin also prevented anxiety-like behavior in both responders and non-responders in the elevated-plus maze and open field test. Collectively, our findings provide the first preclinical evidence that curcumin promotes resilience to CSDS and suggest that curcumin may prevent the emergence of a range of anxiety-like symptoms when given to individuals during exposure to chronic social stress.

Social defeat stress induces depression-like behavior and alters spine morphology in the hippocampus of adolescent male C57BL/6 mice.

Social stress, including bullying during adolescence, is a risk factor for common psychopathologies such as depression. To investigate the neural mechanisms associated with juvenile social stress-induced mood-related endophenotypes, we examined the behavioral, morphological, and biochemical effects of the social defeat stress model of depression on hippocampal dendritic spines within the CA1 stratum radiatum. Adolescent (postnatal day 35) male C57BL/6 mice were subjected to defeat episodes for 10 consecutive days. Twenty-four h later, separate groups of mice were tested on the social interaction and tail suspension tests. Hippocampi were then dissected and Western blots were conducted to quantify protein levels for various markers important for synaptic plasticity including protein kinase M zeta (PKMζ), protein kinase C zeta (PKCζ), the dopamine-1 (D1) receptor, tyrosine hydroxylase (TH), and the dopamine transporter (DAT). Furthermore, we examined the presence of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-receptor subunit GluA2 as well as colocalization with the post-synaptic density 95 (PSD95) protein, within different spine subtypes (filopodia, stubby, long-thin, mushroom) using an immunohistochemistry and Golgi-Cox staining technique. The results revealed that social defeat induced a depression-like behavioral profile, as inferred from decreased social interaction levels, increased immobility on the tail suspension test, and decreases in body weight. Whole hippocampal immunoblots revealed decreases in GluA2, with a concomitant increase in DAT and TH levels in the stressed group. Spine morphology analyses further showed that defeated mice displayed a significant decrease in stubby spines, and an increase in long-thin spines within the CA1 stratum radiatum. Further evaluation of GluA2/PSD95 containing-spines demonstrated a decrease of these markers within long-thin and mushroom spine types. Together, these results indicate that juvenile social stress induces GluA2- and dopamine-associated dysregulation in the hippocampus - a neurobiological mechanism potentially underlying the development of mood-related syndromes as a consequence of adolescent bullying.

Molecular Mechanisms of Stress-Induced Increases in Fear Memory Consolidation within the Amygdala.

Stress can significantly impact brain function and increase the risk for developing various psychiatric disorders. Many of the brain regions that are implicated in psychiatric disorders and are vulnerable to the effects of stress are also involved in mediating emotional learning. Emotional learning has been a subject of intense investigation for the past 30 years, with the vast majority of studies focusing on the amygdala and its role in associative fear learning. However, the mechanisms by which stress affects the amygdala and amygdala-dependent fear memories remain unclear. Here we review the literature on the enhancing effects of acute and chronic stress on the acquisition and/or consolidation of a fear memory, as measured by auditory Pavlovian fear conditioning, and discuss potential mechanisms by which these changes occur in the amygdala. We hypothesize that stress-mediated activation of glucocorticoid receptors (GR) and norepinephrine release within the amygdala leads to the mobilization of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors to the synapse, which underlies stress-induced increases in fear memory. We discuss the implications of this hypothesis for evaluating the effects of stress on extinction and for developing treatments for anxiety disorders. Understanding how stress-induced changes in glucocorticoid and norepinephrine signaling might converge to affect emotional learning by increasing the trafficking of AMPA receptors and enhancing amygdala excitability is a promising area for future research.