Development of endothelial-targeted CD39 as a therapy for ischaemic stroke.

BACKGROUND: Ischaemic stroke is characterized by a necrotic lesion in the brain surrounded by an area of dying cells termed the penumbra. Salvaging the penumbra either with thrombolysis or mechanical retrieval is the cornerstone of stroke management. At-risk neuronal cells release extracellular adenosine triphosphate (eATP) triggering microglial activation and causing a thromboinflammatory response culminating in endothelial activation and vascular disruption. This is further aggravated by ischaemia-reperfusion (I/R) injury that follows all reperfusion therapies. The ecto-enzyme CD39 regulates eATP by hydrolysing to adenosine which has anti-thrombotic and anti-inflammatory properties and reverses I/R injury. METHODS: We developed anti-VCAM-CD39 that targets the antithrombotic and anti-inflammatory properties of recombinant CD39 to the activated endothelium of the penumbra by binding to vascular cell adhesion molecule (VCAM)-1. Mice were subjected to 30 minutes of middle cerebral artery occlusion (MCAo) and analysed at 24h. Anti-VCAM-CD39 or control agents (saline, non-targeted CD39, or anti-VCAM-inactive CD39) were given at 3h post-MCAo. RESULTS: Anti-VCAM-CD39 treatment reduced neurological deficit; MRI confirmed significantly smaller infarcts together with an increase in cerebrovascular perfusion. Anti-VCAM-CD39 also restored blood brain barrier (BBB) integrity and reduced microglial activation. Coadministration of anti-VCAM-CD39 with thrombolytics (tPA) further reduced infarct volumes and attenuated BBB permeability with no associated increase in intracranial haemorrhage. CONCLUSION: Anti-VCAM-CD39, uniquely targeted to endothelial cells, could be a new stroke therapy even when administered 3 h post ischaemia and may further synergise with thrombolytic therapy to improve stroke outcomes.

ComBating inter-site differences in field strength: harmonizing preclinical traumatic brain injury MRI data.

Integrating datasets from multiple sites and scanners can increase statistical power for neuroimaging studies but can also introduce significant inter-site confounds. We evaluated the effectiveness of ComBat, an empirical Bayes approach, to combine longitudinal preclinical MRI data acquired at 4.7 or 9.4 T at two different sites in Australia. Male Sprague Dawley rats underwent MRI on Days 2, 9, 28, and 150 following moderate/severe traumatic brain injury (TBI) or sham injury as part of Project 1 of the NIH/NINDS-funded Centre Without Walls EpiBioS4Rx project. Diffusion-weighted and multiple-gradient-echo images were acquired, and outcomes included QSM, FA, and ADC. Acute injury measures including apnea and self-righting reflex were consistent between sites. Mixed-effect analysis of ipsilateral and contralateral corpus callosum (CC) summary values revealed a significant effect of site on FA and ADC values, which was removed following ComBat harmonization. Bland-Altman plots for each metric showed reduced variability across sites following ComBat harmonization, including for QSM, despite appearing to be largely unaffected by inter-site differences and no effect of site observed. Following harmonization, the combined inter-site data revealed significant differences in the imaging metrics consistent with previously reported outcomes. TBI resulted in significantly reduced FA and increased susceptibility in the ipsilateral CC, and significantly reduced FA in the contralateral CC compared with sham-injured rats. Additionally, TBI rats also exhibited a reversal in ipsilateral CC ADC values over time with significantly reduced ADC at Day 9, followed by increased ADC 150 days after injury. Our findings demonstrate the need for harmonizing multi-site preclinical MRI data and show that this can be successfully achieved using ComBat while preserving phenotypical changes due to TBI.

Sex biology in amyotrophic lateral sclerosis.

Although sex differences in amyotrophic lateral sclerosis (ALS) have not been studied systematically, numerous clinical and preclinical studies have shown sex to be influential in disease prognosis. Moreover, with the development of advanced imaging tools, the difference between male and female brain in structure and function and their response to neurodegeneration are more definitive. As discussed in this review, ALS patients exhibit a sex bias pertaining to the features of the disease, and their clinical, pathological, (and pathophysiological) phenotypes. Several epidemiological studies have indicated that this sex disparity stems from various aetiologies, including sex-specific brain structure and neural functioning, genetic predisposition, age, gonadal hormones, susceptibility to traumatic brain injury (TBI)/head trauma and lifestyle factors.

A pre-existing Toxoplasma gondii infection exacerbates the pathophysiological response and extent of brain damage after traumatic brain injury in mice.

Traumatic brain injury (TBI) is a key contributor to global morbidity that lacks effective treatments. Microbial infections are common in TBI patients, and their presence could modify the physiological response to TBI. It is estimated that one-third of the human population is incurably infected with the feline-borne parasite, Toxoplasma gondii, which can invade the central nervous system and result in chronic low-grade neuroinflammation, oxidative stress, and excitotoxicity-all of which are also important pathophysiological processes in TBI. Considering the large number of TBI patients that have a pre-existing T. gondii infection prior to injury, and the potential mechanistic synergies between the conditions, this study investigated how a pre-existing T. gondii infection modified TBI outcomes across acute, sub-acute and chronic recovery in male and female mice. Gene expression analysis of brain tissue found that neuroinflammation and immune cell markers were amplified in the combined T. gondii + TBI setting in both males and females as early as 2-h post-injury. Glutamatergic, neurotoxic, and oxidative stress markers were altered in a sex-specific manner in T. gondii + TBI mice. Structural MRI found that male, but not female, T. gondii + TBI mice had a significantly larger lesion size compared to their uninfected counterparts at 18-weeks post-injury. Similarly, diffusion MRI revealed that T. gondii + TBI mice had exacerbated white matter tract abnormalities, particularly in male mice. These novel findings indicate that a pre-existing T. gondii infection affects the pathophysiological aftermath of TBI in a sex-dependent manner, and may be an important modifier to consider in the care and prognostication of TBI patients.

Epilepsy phenotype and its reproducibility after lateral fluid percussion-induced traumatic brain injury in rats: Multicenter EpiBioS4Rx study project 1.

OBJECTIVE: This study was undertaken to assess reproducibility of the epilepsy outcome and phenotype in a lateral fluid percussion model of posttraumatic epilepsy (PTE) across three study sites. METHODS: A total of 525 adult male Sprague Dawley rats were randomized to lateral fluid percussion-induced brain injury (FPI) or sham operation. Of these, 264 were assigned to magnetic resonance imaging (MRI cohort, 43 sham, 221 traumatic brain injury [TBI]) and 261 to electrophysiological follow-up (EEG cohort, 41 sham, 220 TBI). A major effort was made to harmonize the rats, materials, equipment, procedures, and monitoring systems. On the 7th post-TBI month, rats were video-EEG monitored for epilepsy diagnosis. RESULTS: A total of 245 rats were video-EEG phenotyped for epilepsy on the 7th postinjury month (121 in MRI cohort, 124 in EEG cohort). In the whole cohort (n = 245), the prevalence of PTE in rats with TBI was 22%, being 27% in the MRI and 18% in the EEG cohort (p > .05). Prevalence of PTE did not differ between the three study sites (p > .05). The average seizure frequency was .317 ± .725 seizures/day at University of Eastern Finland (UEF; Finland), .085 ± .067 at Monash University (Monash; Australia), and .299 ± .266 at University of California, Los Angeles (UCLA; USA; p < .01 as compared to Monash). The average seizure duration did not differ between UEF (104 ± 48 s), Monash (90 ± 33 s), and UCLA (105 ± 473 s; p > .05). Of the 219 seizures, 53% occurred as part of a seizure cluster (≥3 seizures/24 h; p >.05 between the study sites). Of the 209 seizures, 56% occurred during lights-on period and 44% during lights-off period (p > .05 between the study sites). SIGNIFICANCE: The PTE phenotype induced by lateral FPI is reproducible in a multicenter design. Our study supports the feasibility of performing preclinical multicenter trials in PTE to increase statistical power and experimental rigor to produce clinically translatable data to combat epileptogenesis after TBI.

Treatment with the vascular endothelial growth factor-A antibody, bevacizumab, has sex-specific effects in a rat model of mild traumatic brain injury.

Mild traumatic brain injury (mTBI) involves damage to the cerebrovascular system. Vascular endothelial growth factor-A (VEGF-A) is an important modulator of vascular health and VEGF-A promotes the brain's ability to recover after more severe forms of brain injury; however, the role of VEGF-A in mTBI remains poorly understood. Bevacizumab (BEV) is a monoclonal antibody that binds to VEGF-A and neutralises its actions. To better understand the role of VEGF-A in mTBI recovery, this study examined how BEV treatment affected outcomes in rats given a mTBI. Adult Sprague-Dawley rats were assigned to sham-injury + vehicle treatment (VEH), sham-injury + BEV treatment, mTBI + VEH treatment, mTBI + BEV treatment groups. Treatment was administered intracerebroventricularly via a cannula beginning at the time of injury and continuing until the end of the study. Rats underwent behavioral testing after injury and were euthanized on day 11. In both females and males, BEV had a negative impact on cognitive function. mTBI and BEV treatment increased the expression of inflammatory markers in females. In males, BEV treatment altered markers related to hypoxia and vascular health. These novel findings of sex-specific responses to BEV and mTBI provide important insights into the role of VEGF-A in mTBI.

Orexin 2 receptor antagonism sex-dependently improves sleep/wakefulness and cognitive performance in tau transgenic mice.

BACKGROUND AND PURPOSE: Tau pathology contributes to a bidirectional relationship between sleep disruption and neurodegenerative disease. Tau transgenic rTg4510 mice model tauopathy symptoms, including sleep/wake disturbances, which manifest as marked hyperarousal. This phenotype can be prevented by early transgene suppression; however, whether hyperarousal can be rescued after onset is unknown. EXPERIMENTAL APPROACH: Three 8-week experiments were conducted with wild-type and rTg4510 mice after age of onset of hyperarousal (4.5 months): (1) Tau transgene suppression with doxycycline (200 ppm); (2) inactive phase rapid eye movement (REM) sleep enhancement with the dual orexin receptor antagonist suvorexant (50 mg·kg-1 ·day-1 ); or (3) Active phase non-NREM (NREM) and REM sleep enhancement using the selective orexin 2 (OX2 ) receptor antagonist MK-1064 (40 mg·kg-1 ·day-1 ). Sleep was assessed using polysomnography, cognition using the Barnes maze, and tau pathology using immunoblotting and/or immunohistochemistry. KEY RESULTS: Tau transgene suppression improved tauopathy and hippocampal-dependent spatial memory, but did not modify hyperarousal. Pharmacological rescue of REM sleep deficits did not improve spatial memory or tau pathology. In contrast, normalising hyperarousal by increasing both NREM and REM sleep via OX2 receptor antagonism restored spatial memory, independently of tauopathy, but only in male rTg4510 mice. OX2 receptor antagonism induced only short-lived hypnotic responses in female rTg4510 mice and did not improve spatial memory, indicating a tau- and sex-dependent disruption of OX2 receptor signalling. CONCLUSIONS AND IMPLICATIONS: Pharmacologically reducing hyperarousal corrects tau-induced sleep/wake and cognitive deficits. Tauopathy causes sex-dependent disruptions of OX2 receptor signalling/function, which may have implications for choice of hypnotic therapeutics in tauopathies.

Cardiovascular magnetic resonance imaging for sequential assessment of cardiac fibrosis in mice: technical advancements and reverse translation.

Cardiovascular magnetic resonance (CMR) imaging has become an essential technique for the assessment of cardiac function and morphology, and is now routinely used to monitor disease progression and intervention efficacy in the clinic. Cardiac fibrosis is a common characteristic of numerous cardiovascular diseases and often precedes cardiac dysfunction and heart failure. Hence, the detection of cardiac fibrosis is important for both early diagnosis and the provision of guidance for interventions/therapies. Experimental mouse models with genetically and/or surgically induced disease have been widely used to understand mechanisms underlying cardiac fibrosis and to assess new treatment strategies. Improving the appropriate applications of CMR to mouse studies of cardiac fibrosis has the potential to generate new knowledge, and more accurately examine the safety and efficacy of antifibrotic therapies. In this review, we provide 1) a brief overview of different types of cardiac fibrosis, 2) general background on magnetic resonance imaging (MRI), 3) a summary of different CMR techniques used in mice for the assessment of cardiac fibrosis including experimental and technical considerations (contrast agents and pulse sequences), and 4) provide an overview of mouse studies that have serially monitored cardiac fibrosis during disease progression and/or therapeutic interventions. Clinically established CMR protocols have advanced mouse CMR for the detection of cardiac fibrosis, and there is hope that discovery studies in mice will identify new antifibrotic therapies for patients, highlighting the value of both reverse translation and bench-to-bedside research.

Stuttering associated with a pathogenic variant in the chaperone protein cyclophilin 40.

Stuttering is a common speech disorder that interrupts speech fluency and tends to cluster in families. Typically, stuttering is characterized by speech sounds, words or syllables which may be repeated or prolonged and speech that may be further interrupted by hesitations or 'blocks'. Rare variants in a small number of genes encoding lysosomal pathway proteins have been linked to stuttering. We studied a large four-generation family in which persistent stuttering was inherited in an autosomal dominant manner with disruption of the cortico-basal-ganglia-thalamo-cortical network found on imaging. Exome sequencing of three affected family members revealed the PPID c.808C>T (p.Pro270Ser) variant that segregated with stuttering in the family. We generated a Ppid p.Pro270Ser knock-in mouse model and performed ex vivo imaging to assess for brain changes. Diffusion-weighted MRI in the mouse revealed significant microstructural changes in the left corticospinal tract, as previously implicated in stuttering. Quantitative susceptibility mapping also detected changes in cortico-striatal-thalamo-cortical loop tissue composition, consistent with findings in affected family members. This is the first report to implicate a chaperone protein in the pathogenesis of stuttering. The humanized Ppid murine model recapitulates network findings observed in affected family members.

E2730, an uncompetitive γ-aminobutyric acid transporter-1 inhibitor, suppresses epileptic seizures in a rat model of chronic mesial temporal lobe epilepsy.

OBJECTIVE: More than one third of mesial temporal lobe epilepsy (MTLE) patients are resistant to current antiseizure medications (ASMs), and half experience mild-to-moderate adverse effects of ASMs. There is therefore a strong need to develop and test novel ASMs. The objective of this work is to evaluate the pharmacokinetics and neurological toxicity of E2730, a novel uncompetitive inhibitor of γ-aminobutyric acid transporter-1, and to test its seizure suppression effects in a rat model of chronic MTLE. METHODS: We first examined plasma levels and adverse neurological effects of E2730 in healthy Wistar rats. Adult male rats were implanted with osmotic pumps delivering either 10, 20, or 100 mg/kg/day of E2730 subcutaneously for 1 week. Blood sampling and behavioral assessments were performed at several timepoints. We next examined whether E2730 suppressed seizures in rats with chronic MTLE. These rats were exposed to kainic acid-induced status epilepticus, and 9 weeks later, when chronic epilepsy was established, were assigned to receive one of the three doses of E2730 or vehicle for 1 week in a randomized crossover design. Continuous video-electroencephalographic monitoring was acquired during the treatment period to evaluate epileptic seizures. RESULTS: Plasma levels following continuous infusion of E2730 showed a clear dose-related increase in concentration. The drug was well tolerated at all doses, and any sedation or neuromotor impairment was mild and transient, resolving within 48 h of treatment initiation. Remarkably, E2730 treatment in chronically epileptic rats led to seizure suppression in a dose-dependent manner, with 65% of rats becoming seizure-free at the highest dose tested. Mean seizure class did not differ between the treatment groups. SIGNIFICANCE: This study shows that continuous subcutaneous infusion of E2730 over 7 days results in a marked, dose-dependent suppression of spontaneous recurrent seizures, with minimal adverse neurological effects, in a rat model of chronic MTLE. E2730 shows strong promise as an effective new ASM to be translated into clinical trials.

Image data harmonization tools for the analysis of post-traumatic epilepsy development in preclinical multisite MRI studies.

Preclinical MRI studies have been utilized for the discovery of biomarkers that predict post-traumatic epilepsy (PTE). However, these single site studies often lack statistical power due to limited and homogeneous datasets. Therefore, multisite studies, such as the Epilepsy Bioinformatics Study for Antiepileptogenic Therapy (EpiBioS4Rx), are developed to create large, heterogeneous datasets that can lead to more statistically significant results. EpiBioS4Rx collects preclinical data internationally across sites, including the United States, Finland, and Australia. However, in doing so, there are robust normalization and harmonization processes that are required to obtain statistically significant and generalizable results. This work describes the tools and procedures used to harmonize multisite, multimodal preclinical imaging data acquired by EpiBioS4Rx. There were four main harmonization processes that were utilized, including file format harmonization, naming convention harmonization, image coordinate system harmonization, and diffusion tensor imaging (DTI) metrics harmonization. By using Python tools and bash scripts, the file formats, file names, and image coordinate systems are harmonized across all the sites. To harmonize DTI metrics, values are estimated for each voxel in an image to generate a histogram representing the whole image. Then, the Quantitative Imaging Toolkit (QIT) modules are utilized to scale the mode to a value of one and depict the subsequent harmonized histogram. The standardization of file formats, naming conventions, coordinate systems, and DTI metrics are qualitatively assessed. The histograms of the DTI metrics were generated for all the individual rodents per site. For inter-site analysis, an average of the individual scans was calculated to create a histogram that represents each site. In order to ensure the analysis can be run at the level of individual animals, the sham and TBI cohort were analyzed separately, which depicted the same harmonization factor. The results demonstrate that these processes qualitatively standardize the file formats, naming conventions, coordinate systems, and DTI metrics of the data. This assists in the ability to share data across the study, as well as disseminate tools that can help other researchers to strengthen the statistical power of their studies and analyze data more cohesively.

Acute effects of single and repeated mild traumatic brain injury on levels of neurometabolites, lipids, and mitochondrial function in male rats.

Introduction: Mild traumatic brain injuries (mTBIs) are the most common form of acquired brain injury. Symptoms of mTBI are thought to be associated with a neuropathological cascade, potentially involving the dysregulation of neurometabolites, lipids, and mitochondrial bioenergetics. Such alterations may play a role in the period of enhanced vulnerability that occurs after mTBI, such that a second mTBI will exacerbate neuropathology. However, it is unclear whether mTBI-induced alterations in neurometabolites and lipids that are involved in energy metabolism and other important cellular functions are exacerbated by repeat mTBI, and if such alterations are associated with mitochondrial dysfunction. Methods: In this experiment, using a well-established awake-closed head injury (ACHI) paradigm to model mTBI, male rats were subjected to a single injury, or five injuries delivered 1 day apart, and injuries were confirmed with a beam-walk task and a video observation protocol. Abundance of several neurometabolites was evaluated 24 h post-final injury in the ipsilateral and contralateral hippocampus using in vivo proton magnetic resonance spectroscopy (1H-MRS), and mitochondrial bioenergetics were evaluated 30 h post-final injury, or at 24 h in place of 1H-MRS, in the rostral half of the ipsilateral hippocampus. Lipidomic evaluations were conducted in the ipsilateral hippocampus and cortex. Results: We found that behavioral deficits in the beam task persisted 1- and 4 h after the final injury in rats that received repetitive mTBIs, and this was paralleled by an increase and decrease in hippocampal glutamine and glucose, respectively, whereas a single mTBI had no effect on sensorimotor and metabolic measurements. No group differences were observed in lipid levels and mitochondrial bioenergetics in the hippocampus, although some lipids were altered in the cortex after repeated mTBI. Discussion: The decrease in performance in sensorimotor tests and the presence of more neurometabolic and lipidomic abnormalities, after repeated but not singular mTBI, indicates that multiple concussions in short succession can have cumulative effects. Further preclinical research efforts are required to understand the underlying mechanisms that drive these alterations to establish biomarkers and inform treatment strategies to improve patient outcomes.

Variants in SART3 cause a spliceosomopathy characterised by failure of testis development and neuronal defects.

Squamous cell carcinoma antigen recognized by T cells 3 (SART3) is an RNA-binding protein with numerous biological functions including recycling small nuclear RNAs to the spliceosome. Here, we identify recessive variants in SART3 in nine individuals presenting with intellectual disability, global developmental delay and a subset of brain anomalies, together with gonadal dysgenesis in 46,XY individuals. Knockdown of the Drosophila orthologue of SART3 reveals a conserved role in testicular and neuronal development. Human induced pluripotent stem cells carrying patient variants in SART3 show disruption to multiple signalling pathways, upregulation of spliceosome components and demonstrate aberrant gonadal and neuronal differentiation in vitro. Collectively, these findings suggest that bi-allelic SART3 variants underlie a spliceosomopathy which we tentatively propose be termed INDYGON syndrome (Intellectual disability, Neurodevelopmental defects and Developmental delay with 46,XY GONadal dysgenesis). Our findings will enable additional diagnoses and improved outcomes for individuals born with this condition.

Nogo receptor-Fc delivered by haematopoietic cells enhances neurorepair in a multiple sclerosis model.

Nogo receptor 1 is the high affinity receptor for the potent myelin-associated inhibitory factors that make up part of the inflammatory extracellular milieu during experimental autoimmune encephalomyelitis. Signalling through the Nogo receptor 1 complex has been shown to be associated with axonal degeneration in an animal model of multiple sclerosis, and neuronal deletion of this receptor homologue, in a disease specific manner, is associated with preserving axons even in the context of neuroinflammation. The local delivery of Nogo receptor(1-310)-Fc, a therapeutic fusion protein, has been successfully applied as a treatment in animal models of spinal cord injury and glaucoma. As multiple sclerosis and experimental autoimmune encephalomyelitis exhibit large numbers of inflammatory cell infiltrates within the CNS lesions, we utilized transplantable haematopoietic stem cells as a cellular delivery method of the Nogo receptor(1-310)-Fc fusion protein. We identified CNS-infiltrating macrophages as the predominant immune-positive cell type that overexpressed myc-tagged Nogo receptor(1-310)-Fc fusion protein at the peak stage of experimental autoimmune encephalomyelitis. These differentiated phagocytes were predominant during the extensive demyelination and axonal damage, which are associated with the engulfment of the protein complex of Nogo receptor(1-310)-Fc binding to myelin ligands. Importantly, mice transplanted with haematopoietic stem cells transduced with the lentiviral vector carrying Nogo receptor(1-310)-Fc and recovered from the peak of neurological decline during experimental autoimmune encephalomyelitis, exhibiting axonal regeneration and eventual remyelination in the white matter tracts. There were no immunomodulatory effects of the transplanted, genetically modified haematopoietic stem cells on immune cell lineages of recipient female mice induced with experimental autoimmune encephalomyelitis. We propose that cellular delivery of Nogo receptor(1-310)-Fc fusion protein through genetically modified haematopoietic stem cells can modulate multifocal experimental autoimmune encephalomyelitis lesions and potentiate neurological recovery.

Persistent Changes in Mechanical Nociception in Rats With Traumatic Brain Injury Involving Polytrauma.

Traumatic brain injury (TBI) survivors often experience debilitating consequences. Due to the high impact nature of TBI, patients often experience concomitant peripheral injuries (ie, polytrauma). A common, yet often overlooked, comorbidity of TBI is chronic pain. Therefore, this study investigated how common concomitant peripheral injuries (ie, femoral fracture and muscle crush) can affect long-term behavioral and structural TBI outcomes with a particular focus on nociception. Rats were randomly assigned to 1 of 4 groups: polytrauma (POLY; ie, fracture + muscle crush + TBI), peripheral injury (PERI; ie, fracture + muscle crush + sham TBI), TBI (ie, sham fracture + sham muscle crush + TBI), and sham-injured (SHAM; ie, sham fracture + sham muscle crush + sham TBI). Rats underwent behavioral testing at 3-, 6-, and 11-weeks postinjury, and were then euthanized for postmortem magnetic resonance imaging (MRI). POLY rats had a persisting increase in pain sensitivity compared to all groups on the von Frey test. MRI revealed that POLY rats also had abnormalities in the cortical and subcortical brain structures involved in nociceptive processing. These findings have important implications and provide a foundation for future studies to determine the underlying mechanisms and potential treatment strategies for chronic pain in TBI survivors. PERSPECTIVE: Rats with TBI and concomitant peripheral trauma displayed chronic nociceptive pain and MRI images also revealed damaged brain structures/pathways that are involved in chronic pain development. This study highlights the importance of polytrauma and the affected brain regions for developing chronic pain.

Initial Severity of Injury Has Little Effect on the Temporal Profile of Long-Term Deficits in Locomotion, Anxiety, and Cognitive Function After Diffuse Traumatic Brain Injury.

Traumatic brain injury (TBI) is associated with persistent impairments in multiple domains, including cognitive and neuropsychiatric function. Previous literature has suggested that the risk of such impairments may differ as a function of the initial severity of injury, with moderate-severe TBI (msTBI) associated with more severe cognitive dysfunction and mild TBI (mTBI) associated with a higher risk of developing an anxiety disorder. Despite this, relatively few pre-clinical studies have investigated the time course of behavioral change after different severities of injury. The current study compared the temporal profile of functional deficits incorporating locomotion, cognition, and anxiety up to 12 months post-injury after an mTBI, repeated mild TBI (rmTBI), and single msTBI in an experimental model of diffuse TBI. Injury appeared to alter the effect of aging on locomotor activity, with both msTBI and rmTBI rats showing a decrease in locomotion at 12 months relative to their earlier performance on the task, an effect not observed in shams or after a single mTBI. Further, mTBI seemed to be associated with decreased anxiety over time, as measured by increased time spent in the open arm of the elevated plus maze from 3 to 12 months post-injury. No significant findings were observed on spatial memory or volumetric magnetic resonance imaging. Future studies will need to use a more comprehensive behavioral battery, capable of capturing subtle alterations in function, and longer time points, following rats into old age, in order to more fully assess the evolution of persistent behavioral deficits in key domains after different severities of TBI, as well as their accompanying neuroimaging changes. Given the prevalence and significance of such deficits post-TBI for a person's quality of life, as well as the elevated risk of neurodegenerative disease post-injury, such investigations may play a critical role in identifying optimal windows of therapeutic intervention post-injury.

Serum neurofilament light as a biomarker of vulnerability to a second mild traumatic brain injury.

A second mild traumatic brain injury (mTBI) sustained prior to neuropathological recovery can lead to exacerbated effects. Without objective indicators of this neuropathology, individuals may return to activities at risk of mTBI when their brain is still vulnerable. With axonal injury recognized as a neuropathological hallmark of mTBI, we hypothesized that serum levels of neurofilament light (NfL), a highly sensitive biomarker of axonal injury, may be predictive of vulnerability to worse outcomes in the event of a second mTBI. Given this hypothesis is difficult to test clinically, we used a two-hit model of mTBI in rats and staggered inter-injury intervals by 1-, 3-, 7-, or 14-days. Repeat-mTBI rats were dichotomized into NfLhigh (NfL>median at the time of re-injury) and NfLlow (NfL

Platelet-targeted thrombolysis for treatment of acute ischemic stroke.

Thrombolysis with tissue-type plasminogen activator (tPA) remains the main treatment for acute ischemic stroke. Nevertheless, tPA intervention is limited by a short therapeutic window, low recanalization rates, and a risk of intracranial hemorrhage (ICH), highlighting the clinical demand for improved thrombolytic drugs. We examined a novel thrombolytic agent termed "SCE5-scuPA," comprising a single-chain urokinase plasminogen activator (scuPA) fused with a single-chain antibody (SCE5) that targets the activated glycoprotein IIb/IIIa platelet receptor, for its effects in experimental stroke. SCE5-scuPA was first tested in a whole blood clot degradation assay to show the benefit of platelet-targeted thrombolysis. The tail bleeding time, blood clearance, and biodistribution were then determined to inform the use of SCE5-scuPA in mouse models of photothrombotic stroke and middle cerebral artery occlusion against tenecteplase. The impacts of SCE5-scuPA on motor function, ICH, blood-brain barrier (BBB) integrity, and immunosuppression were evaluated. Infarct size was measured by computed tomography imaging and magnetic resonance imaging. SCE5-scuPA enhanced clot degradation ex vivo compared with its nonplatelet-targeting control. The maximal SCE5-scuPA dose that maintained hemostasis and a rapid blood clearance was determined. SCE5-scuPA administration both before and 2 hours after photothrombotic stroke reduced the infarct volume. SCE5-scuPA also improved neurologic deficit, decreased intracerebral blood deposits, preserved the BBB, and alleviated immunosuppression poststroke. In middle cerebral artery occlusion, SCE5-scuPA did not worsen stroke outcomes or cause ICH, and it protected the BBB. Our findings support the ongoing development of platelet-targeted thrombolysis with SCE5-scuPA as a novel emergency treatment for acute ischemic stroke with a promising safety profile.

A companion to the preclinical common data elements and case report forms for in vivo rodent neuroimaging: A report of the TASK3-WG3 Neuroimaging Working Group of the ILAE/AES Joint Translational Task Force.

The International League Against Epilepsy/American Epilepsy Society (ILAE/AES) Joint Translational Task Force established the TASK3 working groups to create common data elements (CDEs) for various aspects of preclinical epilepsy research studies, which could help improve the standardization of experimental designs. In this article, we discuss CDEs for neuroimaging data that are collected in rodent models of epilepsy, with a focus on adult rats and mice. We provide detailed CDE tables and case report forms (CRFs), and with this companion manuscript, we discuss the methodologies for several imaging modalities and the parameters that can be collected.