Enhanced fear memory after social defeat in mice is dependent on interleukin-1 receptor signaling in glutamatergic neurons.

Chronic stress is associated with increased anxiety, cognitive deficits, and post-traumatic stress disorder. Repeated social defeat (RSD) in mice causes long-term stress-sensitization associated with increased microglia activation, monocyte accumulation, and enhanced interleukin (IL)-1 signaling in endothelia and neurons. With stress-sensitization, mice have amplified neuronal, immune, and behavioral responses to acute stress 24 days later. This is clinically relevant as it shares key aspects with post-traumatic stress disorder. The mechanisms underlying stress-sensitization are unclear, but enhanced fear memory may be critical. The purpose of this study was to determine the influence of microglia and IL-1R1 signaling in neurons in the development of sensitization and increased fear memory after RSD. Here, RSD accelerated fear acquisition, delayed fear extinction, and increased cued-based freezing at 0.5 day. The enhancement in contextual fear memory after RSD persisted 24 days later. Next, microglia were depleted with a CSF1R antagonist prior to RSD and several parameters were assessed. Microglia depletion blocked monocyte recruitment to the brain. Nonetheless, neuronal reactivity (pCREB) and IL-1ß RNA expression in the hippocampus and enhanced fear memory after RSD were microglial-independent. Because IL-1ß RNA was prominent in the hippocampus after RSD even with microglia depletion, IL-1R1 mediated signaling in glutamatergic neurons was assessed using neuronal Vglut2+/IL-1R1-/- mice. RSD-induced neuronal reactivity (pCREB) in the hippocampus and enhancement in fear memory were dependent on neuronal IL-1R1 signaling. Furthermore, single-nuclei RNA sequencing (snRNAseq) showed that RSD influenced transcription in specific hippocampal neurons (DG neurons, CA2/3, CA1 neurons) associated with glutamate signaling, inflammation and synaptic plasticity, which were neuronal IL-1R1-dependent. Furthermore, snRNAseq data provided evidence that RSD increased CREB, BDNF, and calcium signaling in DG neurons in an IL-1R1-dependent manner. Collectively, increased IL-1R1-mediated signaling (monocytes/microglia independent) in glutamatergic neurons after RSD enhanced neuronal reactivity and fear memory.

Antagonism of the brain P2X7 ion channel attenuates repeated social defeat induced microglia reactivity, monocyte recruitment and anxiety-like behavior in male mice.

Chronic stress is linked to increased anxiety. Repeated social defeat (RSD) in mice causes anxiety that is dependent on activated neurons, reactive microglia, and accumulation of monocytes in the brain. This response requires interactions between the immune system and central nervous system (CNS). Neuronal activation within threat appraisal regions is a key response to RSD, however, it is unclear how microglia become activated. One potential explanation is that microglia express a purinergic non-selective ligand gated adenosine-triphosphate (ATP) receptor 7 (P2X7). Activation of P2X7 promotes the release of chemokines and cytokines, and recruitment of monocytes to the brain. Thus, the purpose of this study was to determine if a novel P2X7 antagonist blocked neuronal and microglia interactions and the corresponding anxiety following RSD. Male mice were administered (i.p.) a P2X7 antagonist, JNJ-54471300, prior to each cycle of RSD. Fourteen hours after RSD, behavioral deficits including social avoidance and anxiety-like were determined. Moreover, several immune parameters were assessed. RSD caused neuronal activation in stress-responsive regions, monocyte production and release, splenomegaly, and social avoidance. These parameters were unaffected by P2X7 antagonism. RSD-associated proportional area of Iba-1+ microglia, monocyte accumulation in the brain, IL-1ß mRNA expression in enriched myeloid cells, plasma IL-6, and anxiety-like behavior were ameliorated by P2X7 antagonism. Gene expression analysis in the hippocampus and amygdala showed regional specific responses to RSD and some were reversed with P2X7 antagonism. Overall, blocking P2X7 activation attenuated RSD-induced microglia reactivity with corresponding reduction in neuroinflammation, monocyte accumulation, and anxiety-like behavior in male mice.

CD8+ T cells contribute to diet-induced memory deficits in aged male rats.

We have previously shown that short-term (3-day) high fat diet (HFD) consumption induces a neuroinflammatory response and subsequent impairment of long-term memory in aged, but not young adult, male rats. However, the immune cell phenotypes driving this proinflammatory response are not well understood. Previously, we showed that microglia isolated from young and aged rats fed a HFD express similar levels of priming and proinflammatory transcripts, suggesting that additional factors may drive the exaggerated neuroinflammatory response selectively observed in aged HFD-fed rats. It is established that T cells infiltrate both the young and especially the aged central nervous system (CNS) and contribute to immune surveillance of the parenchyma. Thus, we investigated the modulating role of short-term HFD on T cell presence in the CNS in aged rats using bulk RNA sequencing and flow cytometry. RNA sequencing results indicate that aging and HFD altered the expression of genes and signaling pathways associated with T cell signaling, immune cell trafficking, and neuroinflammation. Moreover, flow cytometry data showed that aging alone increased CD4+ and CD8+ T cell presence in the brain and that CD8+, but not CD4+, T cells were further increased in aged rats fed a HFD. Based on these data, we selectively depleted circulating CD8+ T cells via an intravenous injection of an anti-CD8 antibody in aged rats prior to 3 days of HFD to infer the functional role these cells may be playing in long-term memory and neuroinflammation. Results indicate that peripheral depletion of CD8+ T cells lowered hippocampal cytokine levels and prevented the HFD-induced i) increase in brain CD8+ T cells, ii) memory impairment, and iii) alterations in pre- and post-synaptic structures in the hippocampus and amygdala. Together, these data indicate a substantial role for CD8+ T cells in mediating diet-induced memory impairments in aged male rats.

The neuroimmunology of social-stress-induced sensitization.

Myriad clinical findings provide links between chronic stressors, inflammation, and mood disorders. Furthermore, traumatic or chronic exposure to psychological stressors may promote stress sensitization, in which individuals have long-term complications, including increased vulnerability to subsequent stressors. Post-traumatic stress disorder (PTSD) is a clinically relevant example of stress sensitization. PTSD alters neuronal circuitry and mood; however, the mechanisms underlying long-term stress sensitization within this disorder are unclear. Rodent models of chronic social defeat recapitulate several key physiological, immunological, and behavioral responses associated with psychological stress in humans. Repeated social defeat (RSD) uniquely promotes the convergence of neuronal, central inflammatory (microglial), and peripheral immune (monocyte) pathways, leading to prolonged anxiety, social withdrawal, and cognitive impairment. Moreover, RSD promotes stress sensitization, in which mice are highly sensitive to subthreshold stress exposure and recurrence of anxiety weeks after the cessation of stress. Therefore, the purpose of this Review is to discuss the influence of social-defeat stress on the immune system that may underlie stress sensitization within three key cellular compartments: neurons, microglia, and monocytes. Delineating the mechanisms of stress sensitization is critical in understanding and treating conditions such as PTSD.

IL-1 Receptor-1 on Vglut2 + neurons in the hippocampus is critical for neuronal and behavioral sensitization after repeated social stress.

Myriad findings connect stress and inflammation to mood disorders. Social defeat in mice promotes the convergence of neuronal, central inflammatory (microglia), and peripheral immune (monocytes) pathways causing anxiety, social avoidance, and "stress-sensitization." Stress-sensitization results in augmented inflammation and the recurrence of anxiety after re-exposure to social stress. Different cell compartments, including neurons, may be uniquely sensitized by social defeat-induced interleukin-1 (IL-1) signaling. Therefore, the aim of this study was to determine if glutamatergic neuronal IL-1 receptor signaling was essential in promoting stress-sensitization after social defeat. Here, wild-type (IL-1R1+/+) mice and mice with IL-1 receptor-1 deleted selectively in glutamatergic neurons (Vglut2-IL-1R1-/-) were stress-sensitized by social defeat (6-cycles) and then exposed to acute defeat (1-cycle) at day 30. Acute defeat-induced neuronal activation (ΔFosB and phospo-CREB) in the hippocampus of stress-sensitized mice was dependent on neuronal IL-1R1. Moreover, acute defeat-induced social withdrawal and working memory impairment in stress-sensitized mice were also dependent on neuronal IL-1R1. To address region and time dependency, an AAV2-IL-1 receptor antagonist construct was administered into the hippocampus after sensitization, but prior to acute defeat at day 30. Although stress-sensitized mice had increased hippocampal pCREB and decreased working memory after stress re-exposure, these events were not influenced by AAV2-IL-1 receptor antagonist. Hippocampal ΔFosB induction and corresponding social withdrawal in stress-sensitized mice after stress re-exposure were prevented by the AAV2-IL-1 receptor antagonist. Collectively, IL-1 signaling in glutamatergic neurons of the hippocampus was essential in neuronal-sensitization after social defeat and the recall of social withdrawal.

Unique brain endothelial profiles activated by social stress promote cell adhesion, prostaglandin E2 signaling, hypothalamic-pituitary-adrenal axis modulation, and anxiety.

Chronic stress may precipitate psychiatric disorders including anxiety. We reported that Repeated Social Defeat (RSD) in mice increased accumulation of inflammatory monocytes within the brain vasculature, which corresponded with increased interleukin (IL)-1 Receptor 1-mediated activation of endothelia, and augmented anxiety-like behavior. One unknown, however, is the role of immune-activated endothelia in regulating the physiological and behavioral responses to social stress. Thus, we sought to determine the RNA profile of activated endothelia and delineate the pathways by which these endothelia communicate within the brain to influence key responses to social stress. First, endothelial-specific RiboTag mice were exposed to RSD and brain endothelial mRNA profiles from the whole brain and prefrontal cortex were determined using RNAseq. RSD increased expression of cell adhesion molecules (Icam1), inflammatory genes (Lrg1, Lcn2, Ackr1, Il1r1), and cyclooxygenase-2 (Ptgs2/COX-2). In studies with IL-1R1KO mice, there was clear dependence on IL-1R1 on endothelia-associated transcripts including Lrg1, Icam1, Lcn2. Moreover, prostaglandin (PG)E2 was increased in the brain after RSD and Ptgs2 was localized to endothelia, especially within the hypothalamus. Next, a selective COX-2 inhibitor, Celecoxib (CCB), was used with social stress. RSD increased PGE2 in the brain and this was abrogated by CCB. Moreover, CCB reduced RSD-induced Hypothalamic-Pituitary-Adrenal (HPA) axis activation with attenuation of hypothalamic paraventricular neuron activation, hypothalamic Crh expression, and corticosterone in circulation. Production, release, and accumulation of inflammatory monocytes after RSD was COX-2 independent. Nonetheless, CCB blocked anxiety-like behavior in response to RSD. Collectively, social stress stimulated specific endothelia RNA profiles associated with increased cell adhesion, IL-1 and prostaglandin signaling, HPA axis activation, and anxiety.

Breaking Mental Barriers Promotes Recovery After Spinal Cord Injury.

Functional recovery after spinal cord injury (SCI) often proves difficult as physical and mental barriers bar survivors from enacting their designated rehabilitation programs. We recently demonstrated that adult mice administered gabapentinoids, clinically approved drugs prescribed to mitigate chronic neuropathic pain, recovered upper extremity function following cervical SCI. Given that rehabilitative training enhances neuronal plasticity and promotes motor recovery, we hypothesized that the combination of an aerobic-based rehabilitation regimen like treadmill training with gabapentin (GBP) administration will maximize recovery in SCI mice by strengthening synaptic connections along the sensorimotor axis. Whereas mice administered GBP recovered forelimb functions over the course of weeks and months following SCI, no additive forelimb recovery as the result of voluntary treadmill training was noted in these mice. To our surprise, we also failed to find an additive effect in mice administered vehicle. As motivation is crucial in rehabilitation interventions, we scored active engagement toward the rehabilitation protocol and found that mice administered GBP were consistently participating in the rehabilitation program. In contrast, mice administered vehicle exhibited a steep decline in participation, especially at chronic time points. Whereas neuroinflammatory gene expression profiles were comparable between experimental conditions, we discovered that mice administered GBP had increased hippocampal neurogenesis and exhibited less anxiety-like behavior after SCI. We also found that an external, social motivator effectively rescues participation in mice administered vehicle and promotes forelimb recovery after chronic SCI. Thus, not only does a clinically relevant treatment strategy preclude the deterioration of mental health after chronic SCI, but group intervention strategies may prove to be physically and emotionally beneficial for SCI individuals.