Circuits and Biomarkers of the Central Nervous System Relating to Astronaut Performance: Summary Report for a NASA-Sponsored Technical Interchange Meeting.

Biomarkers, ranging from molecules to behavior, can be used to identify thresholds beyond which performance of mission tasks may be compromised and could potentially trigger the activation of countermeasures. Identification of homologous brain regions and/or neural circuits related to operational performance may allow for translational studies between species. Three discussion groups were directed to use operationally relevant performance tasks as a driver when identifying biomarkers and brain regions or circuits for selected constructs. Here we summarize small-group discussions in tables of circuits and biomarkers categorized by (a) sensorimotor, (b) behavioral medicine and (c) integrated approaches (e.g., physiological responses). In total, hundreds of biomarkers have been identified and are summarized herein by the respective group leads. We hope the meeting proceedings become a rich resource for NASA's Human Research Program (HRP) and the community of researchers.

Hyperpolarized 13C metabolic imaging detects long-lasting metabolic alterations following mild repetitive traumatic brain injury.

Career athletes, active military, and head trauma victims are at increased risk for mild repetitive traumatic brain injury (rTBI), a condition that contributes to the development of epilepsy and neurodegenerative diseases. Standard clinical imaging fails to identify rTBI-induced lesions, and novel non-invasive methods are needed. Here, we evaluated if hyperpolarized 13C magnetic resonance spectroscopic imaging (HP 13C MRSI) could detect long-lasting changes in brain metabolism 3.5 months post-injury in a rTBI mouse model. Our results show that this metabolic imaging approach can detect changes in cortical metabolism at that timepoint, whereas multimodal MR imaging did not detect any structural or contrast alterations. Using Machine Learning, we further show that HP 13C MRSI parameters can help classify rTBI vs. Sham and predict long-term rTBI-induced behavioral outcomes. Altogether, our study demonstrates the potential of metabolic imaging to improve detection, classification and outcome prediction of previously undetected rTBI.

Postsynaptic density radiation signature following space irradiation.

Introduction: The response of the brain to space radiation is an important concern for astronauts during space missions. Therefore, we assessed the response of the brain to 28Si ion irradiation (600 MeV/n), a heavy ion present in the space environment, on cognitive performance and whether the response is associated with altered DNA methylation in the hippocampus, a brain area important for cognitive performance. Methods: We determined the effects of 28Si ion irradiation on object recognition, 6-month-old mice irradiated with 28Si ions (600 MeV/n, 0.3, 0.6, and 0.9 Gy) and cognitively tested two weeks later. In addition, we determined if those effects were associated with alterations in hippocampal networks and/or hippocampal DNA methylation. Results: At 0.3 Gy, but not at 0.6 Gy or 0.9 Gy, 28Si ion irradiation impaired cognition that correlated with altered gene expression and 5 hmC profiles that mapped to specific gene ontology pathways. Comparing hippocampal DNA hydroxymethylation following proton, 56Fe ion, and 28Si ion irradiation revealed a general space radiation synaptic signature with 45 genes that are associated with profound phenotypes. The most significant categories were glutamatergic synapse and postsynaptic density. Discussion: The brain's response to space irradiation involves novel excitatory synapse and postsynaptic remodeling.

Combined space stressors induce independent behavioral deficits predicted by early peripheral blood monocytes.

Interplanetary space travel poses many hazards to the human body. To protect astronaut health and performance on critical missions, there is first a need to understand the effects of deep space hazards, including ionizing radiation, confinement, and altered gravity. Previous studies of rodents exposed to a single such stressor document significant deficits, but our study is the first to investigate possible cumulative and synergistic impacts of simultaneous ionizing radiation, confinement, and altered gravity on behavior and cognition. Our cohort was divided between 6-month-old female and male mice in group, social isolation, or hindlimb unloading housing, exposed to 0 or 50 cGy of 5 ion simplified simulated galactic cosmic radiation (GCRsim). We report interactions and independent effects of GCRsim exposure and housing conditions on behavioral and cognitive performance. Exposure to GCRsim drove changes in immune cell populations in peripheral blood collected early after irradiation, while housing conditions drove changes in blood collected at a later point. Female mice were largely resilient to deficits observed in male mice. Finally, we used principal component analysis to represent total deficits as principal component scores, which were predicted by general linear models using GCR exposure, housing condition, and early blood biomarkers.

A single-cell atlas of glioblastoma evolution under therapy reveals cell-intrinsic and cell-extrinsic therapeutic targets.

Recent longitudinal studies of glioblastoma (GBM) have demonstrated a lack of apparent selection pressure for specific DNA mutations in recurrent disease. Single-cell lineage tracing has shown that GBM cells possess a high degree of plasticity. Together this suggests that phenotype switching, as opposed to genetic evolution, may be the escape mechanism that explains the failure of precision therapies to date. We profiled 86 primary-recurrent patient-matched paired GBM specimens with single-nucleus RNA, single-cell open-chromatin, DNA and spatial transcriptomic/proteomic assays. We found that recurrent GBMs are characterized by a shift to a mesenchymal phenotype. We show that the mesenchymal state is mediated by activator protein 1. Increased T-cell abundance at recurrence was prognostic and correlated with hypermutation status. We identified tumor-supportive networks of paracrine and autocrine signals between GBM cells, nonmalignant neuroglia and immune cells. We present cell-intrinsic and cell-extrinsic targets and a single-cell multiomics atlas of GBM under therapy.

Aberrant cortical spine dynamics after concussive injury are reversed by integrated stress response inhibition.

Traumatic brain injury (TBI) is a leading cause of long-term neurological disability in the world and the strongest environmental risk factor for the development of dementia. Even mild TBI (resulting from concussive injuries) is associated with a greater than twofold increase in the risk of dementia onset. Little is known about the cellular mechanisms responsible for the progression of long-lasting cognitive deficits. The integrated stress response (ISR), a phylogenetically conserved pathway involved in the cellular response to stress, is activated after TBI, and inhibition of the ISR-even weeks after injury-can reverse behavioral and cognitive deficits. However, the cellular mechanisms by which ISR inhibition restores cognition are unknown. Here, we used longitudinal two-photon imaging in vivo after concussive injury in mice to study dendritic spine dynamics in the parietal cortex, a brain region involved in working memory. Concussive injury profoundly altered spine dynamics measured up to a month after injury. Strikingly, brief pharmacological treatment with the drug-like small-molecule ISR inhibitor ISRIB entirely reversed structural changes measured in the parietal cortex and the associated working memory deficits. Thus, both neural and cognitive consequences of concussive injury are mediated in part by activation of the ISR and can be corrected by its inhibition. These findings suggest that targeting ISR activation could serve as a promising approach to the clinical treatment of chronic cognitive deficits after TBI.

Stem Cell Therapy for Sequestration of Traumatic Brain Injury-Induced Inflammation.

Traumatic brain injury (TBI) is one of the leading causes of long-term neurological disabilities in the world. TBI is a signature disease for soldiers and veterans, but also affects civilians, including adults and children. Following TBI, the brain resident and immune cells turn into a "reactive" state, characterized by the production of inflammatory mediators that contribute to the development of cognitive deficits. Other injuries to the brain, including radiation exposure, may trigger TBI-like pathology, characterized by inflammation. Currently there are no treatments to prevent or reverse the deleterious consequences of brain trauma. The recognition that TBI predisposes stem cell alterations suggests that stem cell-based therapies stand as a potential treatment for TBI. Here, we discuss the inflamed brain after TBI and radiation injury. We further review the status of stem cells in the inflamed brain and the applications of cell therapy in sequestering inflammation in TBI.

Selective Inhibitory Circuit Dysfunction after Chronic Frontal Lobe Contusion.

Traumatic brain injury (TBI) is a leading cause of neurologic disability; the most common deficits affect prefrontal cortex-dependent functions such as attention, working memory, social behavior, and mental flexibility. Despite this prevalence, little is known about the pathophysiology that develops in frontal cortical microcircuits after TBI. We investigated whether alterations in subtype-specific inhibitory circuits are associated with cognitive inflexibility in a mouse model of frontal lobe contusion in both male and female mice that recapitulates aberrant mental flexibility as measured by deficits in rule reversal learning. Using patch-clamp recordings and optogenetic stimulation, we identified selective vulnerability in the non-fast-spiking and somatostatin-expressing (SOM+) subtypes of inhibitory neurons in layer V of the orbitofrontal cortex 2 months after injury. These subtypes exhibited reduced intrinsic excitability and a decrease in their synaptic output onto pyramidal neurons, respectively. By contrast, the fast-spiking and parvalbumin-expressing interneurons did not show changes in intrinsic excitability or synaptic output, respectively. Impairments in non-fast-spiking/SOM+ inhibitory circuit function were also associated with network hyperexcitability. These findings provide evidence for selective disruptions within specific inhibitory microcircuits that may guide the development of novel therapeutics for TBI.SIGNIFICANCE STATEMENT TBI frequently leads to chronic deficits in cognitive and behavioral functions that involve the prefrontal cortex, yet the maladaptive changes that occur in these cortical microcircuits are unknown. Our data indicate that alterations in subtype-specific inhibitory circuits, specifically vulnerability in the non-fast-spiking/somatostatin-expressing interneurons, occurs in the orbitofrontal cortex in the context of chronic deficits in reversal learning. These neurons exhibit reduced excitability and synaptic output, whereas the other prominent inhibitory population in layer V, the fast-spiking/parvalbumin-expressing interneurons as well as pyramidal neurons are not affected. Our work offers mechanistic insight into the subtype-specific function of neurons that may contribute to mental inflexibility after TBI.

Empowering Data Sharing and Analytics through the Open Data Commons for Traumatic Brain Injury Research.

Traumatic brain injury (TBI) is a major public health problem. Despite considerable research deciphering injury pathophysiology, precision therapies remain elusive. Here, we present large-scale data sharing and machine intelligence approaches to leverage TBI complexity. The Open Data Commons for TBI (ODC-TBI) is a community-centered repository emphasizing Findable, Accessible, Interoperable, and Reusable data sharing and publication with persistent identifiers. Importantly, the ODC-TBI implements data sharing of individual subject data, enabling pooling for high-sample-size, feature-rich data sets for machine learning analytics. We demonstrate pooled ODC-TBI data analyses, starting with descriptive analytics of subject-level data from 11 previously published articles (N = 1250 subjects) representing six distinct pre-clinical TBI models. Second, we perform unsupervised machine learning on multi-cohort data to identify persistent inflammatory patterns across different studies, improving experimental sensitivity for pro- versus anti-inflammation effects. As funders and journals increasingly mandate open data practices, ODC-TBI will create new scientific opportunities for researchers and facilitate multi-data-set, multi-dimensional analytics toward effective translation.

Microglia are involved in the protection of memories formed during sleep deprivation.

Sleep deprivation can generate inflammatory responses in the central nervous system. In turn, this inflammation increases sleep drive, leading to a rebound in sleep duration. Microglia, the innate immune cells found exclusively in the CNS, have previously been found to release inflammatory signals and exhibit altered characteristics in response to sleep deprivation. Together, this suggests that microglia may be partially responsible for the brain's response to sleep deprivation through their inflammatory activity. In this study, we ablated microglia from the mouse brain and assessed resulting sleep, circadian, and sleep deprivation phenotypes. We find that microglia are dispensable for both homeostatic sleep and circadian function and the sleep rebound response to sleep deprivation. However, we uncover a phenomenon by which microglia appear to be essential for the protection of fear-conditioning memories formed during the recovery sleep period following a period of sleep deprivation. This phenomenon occurs potentially through the upregulation of synaptic-homeostasis related genes to protect nascent dendritic spines that may be otherwise removed or downscaled during recovery sleep. These findings further expand the list of known functions for microglia in synaptic modulation.

The impact of deep space radiation on cognitive performance: From biological sex to biomarkers to countermeasures.

In the coming decade, astronauts will travel back to the moon in preparation for future Mars missions. Exposure to galactic cosmic radiation (GCR) is a major obstacle for deep space travel. Using multivariate principal components analysis, we found sex-dimorphic responses in mice exposed to accelerated charged particles to simulate GCR (GCRsim); males displayed impaired spatial learning, whereas females did not. Mechanistically, these GCRsim-induced learning impairments corresponded with chronic microglia activation and synaptic alterations in the hippocampus. Temporary microglia depletion shortly after GCRsim exposure mitigated GCRsim-induced deficits measured months after the radiation exposure. Furthermore, blood monocyte levels measured early after GCRsim exposure were predictive of the late learning deficits and microglia activation measured in the male mice. Our findings (i) advance our understanding of charged particle­induced cognitive challenges, (ii) provide evidence for early peripheral biomarkers for identifying late cognitive deficits, and (iii) offer potential therapeutic strategies for mitigating GCR-induced cognitive loss.

Functional role of brain-engrafted macrophages against brain injuries.

BACKGROUND: Brain-resident microglia have a distinct origin compared to macrophages in other organs. Under physiological conditions, microglia are maintained by self-renewal from the local pool, independent of hematopoietic progenitors. Pharmacological depletion of microglia during whole-brain radiotherapy prevents synaptic loss and long-term recognition memory deficits. However, the origin or repopulated cells and the mechanisms behind these protective effects are unknown. METHODS: CD45low/int/CD11b+ cells from naïve brains, irradiated brains, PLX5622-treated brains and PLX5622 + whole-brain radiotherapy-treated brains were FACS sorted and sequenced for transcriptomic comparisons. Bone marrow chimeras were used to trace the origin and long-term morphology of repopulated cells after PLX5622 and whole-brain radiotherapy. FACS analyses of intrinsic and exotic synaptic compartments were used to measure phagocytic activities of microglia and repopulated cells. In addition, concussive brain injuries were given to PLX5622 and brain-irradiated mice to study the potential protective functions of repopulated cells after PLX5622 + whole-brain radiotherapy. RESULTS: After a combination of whole-brain radiotherapy and microglia depletion, repopulated cells are brain-engrafted macrophages that originate from circulating monocytes. Comparisons of transcriptomes reveal that brain-engrafted macrophages have an intermediate phenotype that resembles both monocytes and embryonic microglia. In addition, brain-engrafted macrophages display reduced phagocytic activity for synaptic compartments compared to microglia from normal brains in response to a secondary concussive brain injury. Importantly, replacement of microglia by brain-engrafted macrophages spare mice from whole-brain radiotherapy-induced long-term cognitive deficits, and prevent concussive injury-induced memory loss. CONCLUSIONS: Brain-engrafted macrophages prevent radiation- and concussion-induced brain injuries and cognitive deficits.

Novel microglia-mediated mechanisms underlying synaptic loss and cognitive impairment after traumatic brain injury.

Traumatic brain injury (TBI) is one of the leading causes of long-term neurological disability in the world. Currently, there are no therapeutics for treating the deleterious consequences of brain trauma; this is in part due to a lack of complete understanding of cellular processes that underlie TBI-related pathologies. Following TBI, microglia, the brain resident immune cells, turn into a "reactive" state characterized by the production of inflammatory mediators that contribute to the development of cognitive deficits. Utilizing multimodal, state-of-the-art techniques that widely span from ultrastructural analysis to optogenetic interrogation of circuit function, we investigated the reactive microglia phenotype one week after injury when learning and memory deficits are also measured. Microglia displayed increased: (i) phagocytic activity in vivo, (ii) synaptic engulfment, (iii) increased neuronal contact, including with dendrites and somata (termed 'satellite microglia'). Functionally, satellite microglia might impact somatic inhibition as demonstrated by the associated reduction in inhibitory synaptic drive. Cumulatively, here we demonstrate novel microglia-mediated mechanisms that may contribute to synaptic loss and cognitive impairment after traumatic brain injury.

Roadmap for Advancing Pre-Clinical Science in Traumatic Brain Injury.

Pre-clinical models of disease have long played important roles in the advancement of new treatments. However, in traumatic brain injury (TBI), despite the availability of numerous model systems, translation from bench to bedside remains elusive. Integrating clinical relevance into pre-clinical model development is a critical step toward advancing therapies for TBI patients across the spectrum of injury severity. Pre-clinical models include in vivo and ex vivo animal work-both small and large-and in vitro modeling. The wide range of pre-clinical models reflect substantial attempts to replicate multiple aspects of TBI sequelae in humans. Although these models reveal multiple putative mechanisms underlying TBI pathophysiology, failures to translate these findings into successful clinical trials call into question the clinical relevance and applicability of the models. Here, we address the promises and pitfalls of pre-clinical models with the goal of evolving frameworks that will advance translational TBI research across models, injury types, and the heterogenous etiology of pathology.

Microglia: Ally and Enemy in Deep Space.

In 2024 the first female astronaut will land on the moon, advancing our preparations for human missions to Mars. While on Earth we are protected from space radiation by our planet's magnetic field, on such deep space voyages astronauts will be exposed to high energy particles from solar flares and galactic cosmic rays (GCR). This exposure carries risks to the central nervous system (CNS) that could jeopardize the mission and astronaut health. Earth-bound studies have employed a variety of single-beam and sequential radiation exposures to simulate the effects of GCR exposure in rodents. Multiple studies have shown that GCR simulation induces a maladaptive activation of microglia - the brain-resident immune cells. GCR simulation also induced synaptic changes resulting in lasting cognitive and behavioral defects. Female and male mice show different susceptibilities to GCR exposure, and evidence suggests this sexually dimorphic response is linked to microglia. Manipulating microglia can prevent the development of cognitive deficits in male mice exposed to components of GCR. This discovery may provide clues towards how to protect astronauts' cognitive and behavioral health both during deep space missions and upon return to Earth.

Microglia depletion and cognitive functions after brain injury: From trauma to galactic cosmic ray.

Microglia are the resident immune cells of the central nervous system (CNS). In physiological conditions, microglia contribute to maintaining brain homeostasis by scanning the surrounding parenchyma and acting as scavenger cells. Following different insults to the CNS, microglia turn into a "reactive" state characterized by the production of inflammatory mediators that promote tissue repair to restore homeostasis. Brain insults such as traumatic brain injury, therapeutic brain irradiation and galactic cosmic ray exposure are associated with chronic microglia activation. Chronic microglia activation contributes to injury-related impairments in cognitive functions. Microglia depletion achieved either by pharmacological or genetic techniques represents not only a useful tool for more extensive investigations of microglia roles, but also a potential therapeutic approach to ameliorate or prevent cognitive dysfunctions following brain injury.

Small molecule cognitive enhancer reverses age-related memory decline in mice.

With increased life expectancy, age-associated cognitive decline becomes a growing concern, even in the absence of recognizable neurodegenerative disease. The integrated stress response (ISR) is activated during aging and contributes to age-related brain phenotypes. We demonstrate that treatment with the drug-like small-molecule ISR inhibitor ISRIB reverses ISR activation in the brain, as indicated by decreased levels of activating transcription factor 4 (ATF4) and phosphorylated eukaryotic translation initiation factor eIF2. Furthermore, ISRIB treatment reverses spatial memory deficits and ameliorates working memory in old mice. At the cellular level in the hippocampus, ISR inhibition (i) rescues intrinsic neuronal electrophysiological properties, (ii) restores spine density and (iii) reduces immune profiles, specifically interferon and T cell-mediated responses. Thus, pharmacological interference with the ISR emerges as a promising intervention strategy for combating age-related cognitive decline in otherwise healthy individuals.

The Integrated Stress Response: A Central Memory Switch in Down Syndrome.

Genetic and pharmacological evidence causally demonstrate that the integrated stress response (ISR) is a central molecular switch for long-term memory formation across different species. Zhu et al. (2019) recently demonstrated that persistent activation of the ISR could explain the long-term memory and synaptic plasticity deficits in a mouse model of Down syndrome, the most common genetic cause of intellectual disability.

Integrated Stress Response Inhibitor Reverses Sex-Dependent Behavioral and Cell-Specific Deficits after Mild Repetitive Head Trauma.

Mild repetitive traumatic brain injury (rTBI) induces chronic behavioral and cognitive alterations and increases the risk for dementia. Currently, there are no therapeutic strategies to prevent or mitigate chronic deficits associated with rTBI. Previously we developed an animal model of rTBI that recapitulates the cognitive and behavioral deficits observed in humans. We now report that rTBI results in an increase in risk-taking behavior in male but not female mice. This behavioral phenotype is associated with chronic activation of the integrated stress response and cell-specific synaptic alterations in the type A subtype of layer V pyramidal neurons in the medial prefrontal cortex. Strikingly, by briefly treating animals weeks after injury with ISRIB, a selective inhibitor of the integrated stress response (ISR), we (1) relieve ISR activation, (2) reverse the increased risk-taking behavioral phenotype and maintain this reversal, and (3) restore cell-specific synaptic function in the affected mice. Our results indicate that targeting the ISR even at late time points after injury can permanently reverse behavioral changes. As such, pharmacological inhibition of the ISR emerges as a promising avenue to combat rTBI-induced behavioral dysfunction.