Biofluid, Imaging, Physiological and Functional Biomarkers of Mild Traumatic Brain Injury and Subconcussive Head Impacts.

Post-concussive symptoms are frequently reported by individuals who sustain mild traumatic brain injuries (mTBIs) and subconcussive head impacts, even when evidence of intracranial pathology is lacking. Current strategies used to evaluate head injuries, which primarily rely on self-report, have a limited ability to predict the incidence, severity, and duration of post-concussive symptoms that will develop in an individual patient. Additionally, these self-report measures have little association with the underlying mechanisms of pathology that may contribute to persisting symptoms, impeding advancement in precision treatment for TBI. Emerging evidence suggests that biofluid, imaging, physiological, and functional biomarkers associated with mTBI and subconcussive head impacts may address these shortcomings by providing more objective measures of injury severity and underlying pathology. Interest in the use of biomarker data has rapidly accelerated, which is reflected by the recent efforts of organizations such as the National Institute of Neurological Disorders and Stroke and the National Academy of Sciences, Engineering, and Medicine to prioritize the collection of biomarker data during TBI characterization in acute care settings. Thus, this review aims to describe recent progress in the identification and development of biomarkers of mTBI and subconcussive head impacts and to discuss important considerations for the implementation of these biomarkers in clinical practice.

Perspectives on Development of Measures to Estimate Career Blast Exposure History in Service Members and Veterans.

The Department of Defense (DOD) has recently prioritized the investigation of the acute and chronic adverse brain health and performance effects of low-level blast (LLB) generated by the use of weapons systems. While acute exposure can be quantified by sensor technology, career exposure has no widely accepted and validated measure for characterization. Currently, distinct research groups are developing and validating four promising measures to estimate career blast exposure history: the Salisbury Blast Interview, Blast Exposure Threshold Survey, Blast Ordnance and Occupational Exposure Measure, and the Blast Frequency and Symptom Severity. Each measure offers an assessment of blast history that is uniquely beneficial to addressing specific research questions. However, use of divergent strategies is not efficient to accelerate the field's understanding of the impact of career exposure and Service-connected health outcomes. As a DOD-wide solution, collaboration across these groups is required to develop a tool(s) that can be standardized across research studies and, ultimately, pared down to be implemented in clinical settings. Here, we overview the current four measures and provide a perspective on the way forward for optimization and/or combination in support of this solution.

High-frequency head impact causes chronic synaptic adaptation and long-term cognitive impairment in mice.

Repeated head impact exposure can cause memory and behavioral impairments. Here, we report that exposure to non-damaging, but high frequency, head impacts can alter brain function in mice through synaptic adaptation. High frequency head impact mice develop chronic cognitive impairments in the absence of traditional brain trauma pathology, and transcriptomic profiling of mouse and human chronic traumatic encephalopathy brain reveal that synapses are strongly affected by head impact. Electrophysiological analysis shows that high frequency head impacts cause chronic modification of the AMPA/NMDA ratio in neurons that underlie the changes to cognition. To demonstrate that synaptic adaptation is caused by head impact-induced glutamate release, we pretreated mice with memantine prior to head impact. Memantine prevents the development of the key transcriptomic and electrophysiological signatures of high frequency head impact, and averts cognitive dysfunction. These data reveal synapses as a target of high frequency head impact in human and mouse brain, and that this physiological adaptation in response to head impact is sufficient to induce chronic cognitive impairment in mice.

Military TBI-What civilian primary care providers should know.

In June 2019, the Department of Veterans Affairs (VA) launched the VA Mission Act, which expanded veterans' health-care access to the private sector. Since civilian primary care providers may see more veterans in their practice, it will be important to understand the unique experiences, comorbidities, and culture of this population in order to provide optimal care. Military service members (SMs) are at an increased risk for traumatic brain injury (TBI), and comorbidities, such as post traumatic stress disorder (PTSD), increasing the likelihood of prolonged symptoms. Military training and repetitive low-level blast exposure may cause symptoms similar to TBI or increase long-term negative effects in SMs. Military culture often has a strong influence in this population. Those who serve in the military identify with military values and have a strong team mentality, which places emphasis on the mission above all else, not accepting defeat, and not ever leaving a fellow SM behind. These values can impact the way a SM/veteran seeks care and/or communicates with his or her health-care provider. Taking a detailed history to understand how these factors apply, as well as screening for mental health comorbidities, are recommended. Understanding the military cultural influences can assist in promoting a stronger therapeutic alliance and encourage more open communication. Ultimately, it is the trusting and respectful relationship between the SM/veteran and the provider that will determine the most effective treatment and result in the most effective resolution of TBI and comorbid symptoms.

High-Frequency Head Impact Disrupts Hippocampal Neural Ensemble Dynamics.

We have recently shown that the cognitive impairments in a mouse model of high-frequency head impact (HFHI) are caused by chronic changes to synaptic physiology. To better understand these synaptic changes occurring after repeat head impact, we used Thy1-GcCAMP6f mice to study intracellular and intercellular calcium dynamics and neuronal ensembles in HFHI mice. We performed simultaneous calcium imaging and local field potential (LFP) recordings of the CA1 field during an early-LTP paradigm in acute hippocampal slice preparations 24 h post-impact. As previously reported, HFHI causes a decrease in early-LTP in the absence of any shift in the input-output curve. Calcium analytics revealed that HFHI hippocampal slices have similar numbers of active ROIs, however, the number of calcium transients per ROI was significantly increased in HFHI slices. Ensembles consist of coordinated activity between groups of active ROIs. We exposed the CA1 ensemble to Schaffer-collateral stimulation in an abbreviated LTP paradigm and observed novel coordinated patterns of post stimulus calcium ensemble activity. HFHI ensembles displayed qualitatively similar patterns of post-stimulus ensemble activity to shams but showed significant changes in quantitative ensemble inactivation and reactivation. Previous in vivo and in vitro reports have shown that ensemble activity frequently occurs through a similar set of ROIs firing in a repeating fashion. HFHI slices showed a decrease in such coordinated firing patterns during post stimulus ensemble activity. The present study shows that HFHI alters synaptic activity and disrupts neuronal organization of the ensemble, providing further evidence of physiological synaptic adaptation occurring in the brain after a high frequency of non-pathological head impacts.

Cellular Changes in Injured Rat Spinal Cord Following Electrical Brainstem Stimulation.

Spinal cord injury (SCI) is a major cause of disability and pain, but little progress has been made in its clinical management. Low-frequency electrical stimulation (LFS) of various anti-nociceptive targets improves outcomes after SCI, including motor recovery and mechanical allodynia. However, the mechanisms of these beneficial effects are incompletely delineated and probably multiple. Our aim was to explore near-term effects of LFS in the hindbrain's nucleus raphe magnus (NRM) on cellular proliferation in a rat SCI model. Starting 24 h after incomplete contusional SCI at C5, intermittent LFS at 8 Hz was delivered wirelessly to NRM. Controls were given inactive stimulators. At 48 h, 5-bromodeoxyuridine (BrdU) was administered and, at 72 h, spinal cords were extracted and immunostained for various immune and neuroglial progenitor markers and BrdU at the level of the lesion and proximally and distally. LFS altered cell marker counts predominantly at the dorsal injury site. BrdU cell counts were decreased. Individually and in combination with BrdU, there were reductions in CD68 (monocytes) and Sox2 (immature neural precursors) and increases in Blbp (radial glia) expression. CD68-positive cells showed increased co-staining with iNOS. No differences in the expression of GFAP (glia) and NG2 (oligodendrocytes) or in GFAP cell morphology were found. In conclusion, our work shows that LFS of NRM in subacute SCI influences the proliferation of cell types implicated in inflammation and repair, thus providing mechanistic insight into deep brain stimulation as a neuromodulatory treatment for this devastating pathology.

Apolipoprotein E4 impairs spontaneous blood brain barrier repair following traumatic brain injury.

BACKGROUND: Traumatic Brain Injury (TBI) is a major cause of disability and mortality, to which there is currently no comprehensive treatment. Blood Brain Barrier (BBB) dysfunction is well documented in human TBI patients, yet the molecular mechanisms that underlie this neurovascular unit (NVU) pathology remains unclear. The apolipoprotein-E (apoE) protein has been implicated in controlling BBB integrity in an isoform dependent manner, via suppression of Cyclophilin A (CypA)-Matrix metallopeptidase-9 (MMP-9) signaling cascades, however the contribution of this pathway in TBI-induced BBB permeability is not fully investigated. METHODS: We exposed C57Bl/6 mice to controlled cortical impact and assessed NVU and BBB permeability responses up to 21 days post-injury. We pharmacologically probed the role of the CypA-MMP-9 pathway in BBB permeability after TBI using Cyclosporin A (CsA, 20 mg/kg). Finally, as the apoE4 protein is known to be functionally deficient compared to the apoE3 protein, we used humanized APOE mice as a clinically relevant model to study the role of apoE on BBB injury and repair after TBI. RESULTS: In C57Bl/6 mice there was an inverse relationship between soluble apoE and BBB permeability, such that damaged BBB stabilizes as apoE levels increase in the days following TBI. TBI mice displayed acute pericyte loss, increased MMP-9 production and activity, and reduced tight-junction expression. Treatment with the CypA antagonist CsA in C57Bl/6 mice attenuates MMP-9 responses and enhances BBB repair after injury, demonstrating that MMP-9 plays an important role in the timing of spontaneous BBB repair after TBI. We also show that apoe mRNA is present in both astrocytes and pericytes after TBI. We report that APOE3 and APOE4 mice have similar acute BBB responses to TBI, but APOE3 mice display faster spontaneous BBB repair than APOE4 mice. Isolated microvessel analysis reveals delayed pericyte repopulation, augmented and sustained MMP-9 expression at the NVU, and impaired stabilization of Zonula Occludens-1, Occludin and Claudin-5 expression at tight junctions in APOE4 mice after TBI compared to APOE3 mice. CONCLUSIONS: These data confirm apoE as an important modulator of spontaneous BBB stabilization following TBI, and highlights the APOE4 allele as a risk factor for poor outcome after TBI.

Some Autonomic Deficits of Acute or Chronic Cervical Spinal Contusion Reversed by Interim Brainstem Stimulation.

Prolonged electrical stimulation of the hindbrain's nucleus raphe magnus (NRM) or of its major midbrain input region, the periaqueductal gray (PAG), was previously found in rats to promote recovery from sensory-motor and histological deficits of acute thoracic spinal cord injury (SCI). Here, some visceral deficits of acute and chronic midline cervical (C5) contusion are similarly examined. Cranially implanted wireless stimulators delivered intermittent 8 Hz, 30-70 µA cathodal pulse trains to a brainstem microelectrode. Injured controls were given inactive stimulators; rats without injuries or implants were also compared. Rectal distension or squeezing of the forepaws caused an exaggerated rise in mean arterial pressure in injured, untreated rats under anesthesia on post-injury week 6, probably reflecting autonomic dysreflexia (AD). These pressor responses became normal when 7 days of unilateral PAG stimulation was started on the injury day. Older untreated injuries (weeks 18-19) showed normal pressor responses, but unexpectedly had significant resting and nociceptive bradycardia, which was reversed by 3 weeks of PAG stimulation started on weeks 7 or 12. Subsequent chronic studies examined gastric emptying (GE), as indicated by intestinal transit of gavaged dye, and serum chemistry. GE and fasting serum insulin were reduced on injury weeks 14-15, and were both normalized by ∼5 weeks of PAG stimulation begun in weeks 7-8. Increases in calcitonin gene-related peptide, a prominent visceral afferent neurotransmitter, measured near untreated injuries (first thoracic segment) in superficial dorsal laminae were reversed by acutely or chronically initiated PAG stimulation. The NRM, given 2-3 weeks of stimulation beginning 2 days after SCI, prevented abnormalities in both pressor responses and GE on post-injury week 9, consistent with its relaying of repair commands from the PAG. The descending PAG-NRM axis thus exhibits broadly restorative influences on visceral as well as sensory-motor deficits, improving chronic as well as acute signs of injury.

A Mouse Model of Single and Repetitive Mild Traumatic Brain Injury.

Mild Traumatic Brain Injury (mTBI) can result in the acute loss of brain function, including a period of confusion, a loss of consciousness (LOC), focal neurological deficits and even amnesia. Athletes participating in contact sports are at high risk of exposure to large number of mTBIs. In terms of the level of injury in a sporting athlete, a mTBI is defined as a mild injury that does not cause gross pathological changes, but does cause short-term neurological deficits that are spontaneously resolved. Despite previous attempts to model mTBI in mice and rats, many have reported gross adverse effects including skull fractures, intracerebral bleeding, axonal injury and neuronal cell death. Herein, we describe our highly reproducible animal model of mTBI that reproduces clinically relevant symptoms. This model uses a custom made pneumatic impactor device to deliver a closed-head trauma. This impact is made under precise velocity and deformation parameters, creating a reliable and reproducible model to examine the mechanisms that contribute to effects of single or repetitive concussive mTBI.

Prolonged stimulation of a brainstem raphe region attenuates experimental autoimmune encephalomyelitis.

Multiple sclerosis (MS), a neuroinflammatory disease, has few treatment options, none entirely adequate. We studied whether prolonged electrical microstimulation of a hindbrain region (the nucleus raphe magnus) can attenuate experimental autoimmune encephalomyelitis, a murine model of MS induced by MOG35-55 injection. Eight days after symptoms emerged, a wireless electrical stimulator with an attached microelectrode was implanted cranially, and daily intermittent stimulation was begun in awake, unrestrained mice. The thoracic spinal cord was analyzed for changes in histology (on day 29) and gene expression (on day 37), with a focus on myelination and cytokine production. Controls, with inactive implants, showed a phase of disease exacerbation on days 19-25 that stimulation for >16days eliminated. Prolonged stimulation also reduced numbers of infiltrating immune cells and increased numbers of myelinated axons. It additionally lowered genetic expression of some pro-inflammatory cytokines (interferon gamma and tumor necrosis factor) and platelet-derived growth factor receptor alpha, a marker of oligodendrocyte precursors, while raising expression of myelin basic protein. Studies of restorative treatments for MS might profitably consider ways to stimulate the raphe magnus, directly or via its inputs, or to emulate its serotonergic and peptidergic output.

Proteomic analyses of zebra finch optic tectum and comparative histochemistry.

Proteomic analyses of zebra finch (Taeniopygia guttata) optic tectum resulted in identification of 176 proteins. In the Swissprot database, only 52 proteins were identified as bird homologs and only 71 proteins were identified in songbird transcriptome databases, reflecting a lack of completeness in the T. guttata genomic sequence. Analysis in Kyoto encyclopedia of genes and genome (KEGG) pathway database found that identified proteins most frequently belong to glucose, pyruvate, glyoxylate, dicarboxylate, alanine, and aspartate metabolism pathways. A number of identified proteins have been previously reported to exist in the avian optic tectum. The immunohistochemical localization of selected proteins showed their distribution in similar laminae of the owl (Tyto alba) and chicken (Gallus gallus) tectum. Immunohistochemical analysis of identified proteins can provide clues about cell types and circuit layout of the avian optic tectum in general. As the optic tectum of nonmammals is homologous to the superior colliculus of mammals, the analysis of the tectal and collicular proteome may provide clues about conserved cell and circuit layout, circuit function, and evolution.

Proteomic analyses of songbird (Zebra finch; Taeniopygia guttata) retina.

Proteomic analyses of male songbird (Zebra finch; Taeniopygia guttata; ZF) retina were performed resulting in identification of 129 proteins. Comparison of T. guttata retinal proteome with that of chicken found proteins detected in both retinas. Immunohistochemical analyses of T. guttata retinal sections and Western analyses of total retinal protein extract were performed confirming presence of select bona fide retinal proteins. Results demonstrate the utility of one-dimensional gel fractionation for mass spectrometry and will be useful for future proteomic comparison of songbird retina and brain tissues in different behavioral and pharmacological studies.