CHIMERA repetitive mild traumatic brain injury induces chronic behavioural and neuropathological phenotypes in wild-type and APP/PS1 mice.

BACKGROUND: The annual incidence of traumatic brain injury (TBI) in the United States is over 2.5 million, with approximately 3-5 million people living with chronic sequelae. Compared with moderate-severe TBI, the long-term effects of mild TBI (mTBI) are less understood but important to address, particularly for contact sport athletes and military personnel who have high mTBI exposure. The purpose of this study was to determine the behavioural and neuropathological phenotypes induced by the Closed-Head Impact Model of Engineered Rotational Acceleration (CHIMERA) model of mTBI in both wild-type (WT) and APP/PS1 mice up to 8 months post-injury. METHODS: Male WT and APP/PS1 littermates were randomized to sham or repetitive mild TBI (rmTBI; 2 × 0.5 J impacts 24 h apart) groups at 5.7 months of age. Animals were assessed up to 8 months post-injury for acute neurological deficits using the loss of righting reflex (LRR) and Neurological Severity Score (NSS) tasks, and chronic behavioural changes using the passive avoidance (PA), Barnes maze (BM), elevated plus maze (EPM) and rotarod (RR) tasks. Neuropathological assessments included white matter damage; grey matter inflammation; and measures of Aß levels, deposition, and aducanumab binding activity. RESULTS: The very mild CHIMERA rmTBI conditions used here produced no significant acute neurological or motor deficits in WT and APP/PS1 mice, but they profoundly inhibited extinction of fear memory specifically in APP/PS1 mice over the 8-month assessment period. Spatial learning and memory were affected by both injury and genotype. Anxiety and risk-taking behaviour were affected by injury but not genotype. CHIMERA rmTBI induced chronic white matter microgliosis, axonal injury and astrogliosis independent of genotype in the optic tract but not the corpus callosum, and it altered microgliosis in APP/PS1 amygdala and hippocampus. Finally, rmTBI did not alter long-term tau, Aß or amyloid levels, but it increased aducanumab binding activity. CONCLUSIONS: CHIMERA is a useful model to investigate the chronic consequences of rmTBI, including behavioural abnormalities consistent with features of post-traumatic stress disorder and inflammation of both white and grey matter. The presence of human Aß greatly modified extinction of fear memory after rmTBI.

An End-to-end System for Automatic Characterization of Iba1 Immunopositive Microglia in Whole Slide Imaging.

Traumatic brain injury (TBI) is one of the leading causes of death and disability worldwide. Detailed studies of the microglial response after TBI require high throughput quantification of changes in microglial count and morphology in histological sections throughout the brain. In this paper, we present a fully automated end-to-end system that is capable of assessing microglial activation in white matter regions on whole slide images of Iba1 stained sections. Our approach involves the division of the full brain slides into smaller image patches that are subsequently automatically classified into white and grey matter sections. On the patches classified as white matter, we jointly apply functional minimization methods and deep learning classification to identify Iba1-immunopositive microglia. Detected cells are then automatically traced to preserve their complex branching structure after which fractal analysis is applied to determine the activation states of the cells. The resulting system detects white matter regions with 84% accuracy, detects microglia with a performance level of 0.70 (F1 score, the harmonic mean of precision and sensitivity) and performs binary microglia morphology classification with a 70% accuracy. This automated pipeline performs these analyses at a 20-fold increase in speed when compared to a human pathologist. Moreover, we have demonstrated robustness to variations in stain intensity common for Iba1 immunostaining. A preliminary analysis was conducted that indicated that this pipeline can identify differences in microglia response due to TBI. An automated solution to microglia cell analysis can greatly increase standardized analysis of brain slides, allowing pathologists and neuroscientists to focus on characterizing the associated underlying diseases and injuries.

Age at injury and genotype modify acute inflammatory and neurofilament-light responses to mild CHIMERA traumatic brain injury in wild-type and APP/PS1 mice.

Peak incidence of traumatic brain injury (TBI) occurs in both young and old individuals, and older age at injury is associated with worse outcome and poorer recovery. Moderate-severe TBI is a reported risk factor for dementia, including Alzheimer's disease (AD), but whether mild TBI (mTBI) alters AD pathogenesis is not clear. To delineate how age at injury and predisposition to amyloid formation affect the acute response to mTBI, we used the Closed Head Impact Model of Engineered Rotational Acceleration (CHIMERA) model of TBI to induce two mild injuries in wild-type (WT) and APP/PS1 mice at either 6 or 13months of age and assessed behavioural, histological and biochemical changes up to 14days post-injury. Age at injury did not alter acute behavioural responses to mTBI, including measures of neurological status, motor performance, spatial memory, fear, or anxiety, in either strain. Young APP/PS1 mice showed a subtle and transient increase in diffuse Aß deposits after injury, whereas old APP/PS1 mice showed decreased amyloid deposits, without significant alterations in total soluble or insoluble Aß levels at either age. Age at injury and genotype showed complex responses with respect to microglial and cytokine outcomes, where post-injury neuroinflammation is increased in old WT mice but attenuated in old APP/PS1 mice. Intriguingly, silver staining confirmed axonal damage in both strains and ages, yet only young WT and APP/PS1 mice showed neurofilament-positive axonal swellings after mTBI, as this response was almost entirely attenuated in old mice. Plasma neurofilament-light levels were significantly elevated after injury only in young APP/PS1 mice. This study suggests that mild TBI has minimal effects on Aß metabolism, but that age and genotype can each modify acute outcomes related to white matter injury.

Defining the biomechanical and biological threshold of murine mild traumatic brain injury using CHIMERA (Closed Head Impact Model of Engineered Rotational Acceleration).

CHIMERA (Closed Head Impact Model of Engineered Rotational Acceleration) is a recently described animal model of traumatic brain injury (TBI) that primarily produces diffuse axonal injury (DAI) characterized by white matter inflammation and axonal damage. CHIMERA was specifically designed to reliably generate a variety of TBI severities using precise and quantifiable biomechanical inputs in a nonsurgical user-friendly platform. The objective of this study was to define the lower limit of single impact mild TBI (mTBI) using CHIMERA by characterizing the dose-response relationship between biomechanical input and neurological, behavioral, neuropathological and biochemical outcomes. Wild-type male mice were subjected to a single CHIMERA TBI using six impact energies ranging from 0.1 to 0.7J, and post-TBI outcomes were assessed over an acute period of 14days. Here we report that single TBI using CHIMERA induces injury dose- and time-dependent changes in behavioral and neurological deficits, axonal damage, white matter tract microgliosis and astrogliosis. Impact energies of 0.4J or below produced no significant phenotype (subthreshold), 0.5J led to significant changes for one or more phenotypes (threshold), and 0.6 and 0.7J resulted in significant changes in all outcomes assessed (mTBI). We further show that linear head kinematics are the most robust predictors of duration of unconsciousness, severity of neurological deficits, white matter injury, and microgliosis following single TBI. Our data extend the validation of CHIMERA as a biofidelic animal model of DAI and establish working parameters to guide future investigations of the mechanisms underlying axonal pathology and inflammation induced by mechanical trauma.

Chronic Exposure to Androgenic-Anabolic Steroids Exacerbates Axonal Injury and Microgliosis in the CHIMERA Mouse Model of Repetitive Concussion.

Concussion is a serious health concern. Concussion in athletes is of particular interest with respect to the relationship of concussion exposure to risk of chronic traumatic encephalopathy (CTE), a neurodegenerative condition associated with altered cognitive and psychiatric functions and profound tauopathy. However, much remains to be learned about factors other than cumulative exposure that could influence concussion pathogenesis. Approximately 20% of CTE cases report a history of substance use including androgenic-anabolic steroids (AAS). How acute, chronic, or historical AAS use may affect the vulnerability of the brain to concussion is unknown. We therefore tested whether antecedent AAS exposure in young, male C57Bl/6 mice affects acute behavioral and neuropathological responses to mild traumatic brain injury (TBI) induced with the CHIMERA (Closed Head Impact Model of Engineered Rotational Acceleration) platform. Male C57Bl/6 mice received either vehicle or a cocktail of three AAS (testosterone, nandrolone and 17α-methyltestosterone) from 8-16 weeks of age. At the end of the 7th week of treatment, mice underwent two closed-head TBI or sham procedures spaced 24 h apart using CHIMERA. Post-repetitive TBI (rTBI) behavior was assessed for 7 d followed by tissue collection. AAS treatment induced the expected physiological changes including increased body weight, testicular atrophy, aggression and downregulation of brain 5-HT1B receptor expression. rTBI induced behavioral deficits, widespread axonal injury and white matter microgliosis. While AAS treatment did not worsen post-rTBI behavioral changes, AAS-treated mice exhibited significantly exacerbated axonal injury and microgliosis, indicating that AAS exposure can alter neuronal and innate immune responses to concussive TBI.

Merging pathology with biomechanics using CHIMERA (Closed-Head Impact Model of Engineered Rotational Acceleration): a novel, surgery-free model of traumatic brain injury.

BACKGROUND: Traumatic brain injury (TBI) is a major health care concern that currently lacks any effective treatment. Despite promising outcomes from many preclinical studies, clinical evaluations have failed to identify effective pharmacological therapies, suggesting that the translational potential of preclinical models may require improvement. Rodents continue to be the most widely used species for preclinical TBI research. As most human TBIs result from impact to an intact skull, closed head injury (CHI) models are highly relevant, however, traditional CHI models suffer from extensive experimental variability that may be due to poor control over biomechanical inputs. Here we describe a novel CHI model called CHIMERA (Closed-Head Impact Model of Engineered Rotational Acceleration) that fully integrates biomechanical, behavioral, and neuropathological analyses. CHIMERA is distinct from existing neurotrauma model systems in that it uses a completely non-surgical procedure to precisely deliver impacts of prescribed dynamic characteristics to a closed skull while enabling kinematic analysis of unconstrained head movement. In this study, we characterized head kinematics as well as functional, neuropathological, and biochemical outcomes up to 14d following repeated TBI (rTBI) in adult C57BL/6 mice using CHIMERA. RESULTS: Head kinematic analysis showed excellent repeatability over two closed head impacts separated at 24h. Injured mice showed significantly prolonged loss of righting reflex and displayed neurological, motor, and cognitive deficits along with anxiety-like behavior. Repeated TBI led to diffuse axonal injury with extensive microgliosis in white matter from 2-14d post-rTBI. Injured mouse brains also showed significantly increased levels of TNF-α and IL-1ß and increased endogenous tau phosphorylation. CONCLUSIONS: Repeated TBI using CHIMERA mimics many of the functional and pathological characteristics of human TBI with a reliable biomechanical response of the head. This makes CHIMERA well suited to investigate the pathophysiology of TBI and for drug development programs.

Towards clinical management of traumatic brain injury: a review of models and mechanisms from a biomechanical perspective.

Traumatic brain injury (TBI) is a major worldwide healthcare problem. Despite promising outcomes from many preclinical studies, the failure of several clinical studies to identify effective therapeutic and pharmacological approaches for TBI suggests that methods to improve the translational potential of preclinical studies are highly desirable. Rodent models of TBI are increasingly in demand for preclinical research, particularly for closed head injury (CHI), which mimics the most common type of TBI observed clinically. Although seemingly simple to establish, CHI models are particularly prone to experimental variability. Promisingly, bioengineering-oriented research has advanced our understanding of the nature of the mechanical forces and resulting head and brain motion during TBI. However, many neuroscience-oriented laboratories lack guidance with respect to fundamental biomechanical principles of TBI. Here, we review key historical and current literature that is relevant to the investigation of TBI from clinical, physiological and biomechanical perspectives, and comment on how the current challenges associated with rodent TBI models, particularly those involving CHI, could be improved.

The liver X receptor agonist GW3965 improves recovery from mild repetitive traumatic brain injury in mice partly through apolipoprotein E.

Traumatic brain injury (TBI) increases Alzheimer's disease (AD) risk and leads to the deposition of neurofibrillary tangles and amyloid deposits similar to those found in AD. Agonists of Liver X receptors (LXRs), which regulate the expression of many genes involved in lipid homeostasis and inflammation, improve cognition and reduce neuropathology in AD mice. One pathway by which LXR agonists exert their beneficial effects is through ATP-binding cassette transporter A1 (ABCA1)-mediated lipid transport onto apolipoprotein E (apoE). To test the therapeutic utility of this pathway for TBI, we subjected male wild-type (WT) and apoE-/- mice to mild repetitive traumatic brain injury (mrTBI) followed by treatment with vehicle or the LXR agonist GW3965 at 15 mg/kg/day. GW3965 treatment restored impaired novel object recognition memory in WT but not apoE-/- mice. GW3965 did not significantly enhance the spontaneous recovery of motor deficits observed in all groups. Total soluble Aß(40) and Aß(42) levels were significantly elevated in WT and apoE-/- mice after injury, a response that was suppressed by GW3965 in both genotypes. WT mice showed mild but significant axonal damage at 2 d post-mrTBI, which was suppressed by GW3965. In contrast, apoE-/- mice showed severe axonal damage from 2 to 14 d after mrTBI that was unresponsive to GW3965. Because our mrTBI model does not produce significant inflammation, the beneficial effects of GW3965 we observed are unlikely to be related to reduced inflammation. Rather, our results suggest that both apoE-dependent and apoE-independent pathways contribute to the ability of GW3965 to promote recovery from mrTBI.

Network actions of pentobarbital in the rat mesopontine tegmentum on sensory inflow through the spinothalamic tract.

The recent discovery of a barbiturate-sensitive "general anesthesia switch" mechanism localized in the rat brain stem mesopontine tegmental anesthesia area (MPTA) has challenged the current view of the nonspecific actions of general anesthetic agents in the CNS. In this study we provide electrophysiological evidence that the antinociception, which accompanies the behavioral state resembling general anesthesia following pentobarbital (PB) microinjections into the MPTA of awake rats, could be accompanied by the attenuation of sensory transmission through the spinothalamic tract (STT). Following bilateral microinjections of PB into the MPTA spontaneous firing rate (SFR), antidromic firing index (FI), and sciatic (Sc) as well as sural (Su) nerve-evoked responses (ER) of identified lumbar STT neurons in the isoflurane-anesthetized rat were quantified using extracellular recording techniques. Microinjections of PB into the MPTA significantly suppressed the SFR (47%), magnitudes of Sc- (26%) and Su-ER (36%), and FI (41%) of STT neurons. Microinjections of PB-free vehicle control did not alter any of the above-cited electrophysiological parameters. The results from this study suggest that antinociception, which occurs during the anesthesia-like state following PB microinjections into the MPTA, may be due, in part, to (in)direct inhibition of STT neurons via switching mechanism(s) located in the MPTA. This study provides a provenance for investigating electrophysiologically the actions on STT neurons of other current agents used clinically to maintain the state of general anesthesia.