Search

BACKGROUND: Neonatal hypoxia-ischemia (HI) is a major cause of perinatal brain injury and is associated with a spectrum of neuropsychiatric disorders. Although very few treatment options are currently available, doxycycline (DOXY) has been reported to be neuroprotective in neontatal HI. Our objective was to investigate the effects of DOXY on neonatal brain development in normal and HI rat pups. We hypothesized that DOXY would inhibit microglial activation but that developmentally important processes, including cytogenesis and trophic responses, would not be impaired. METHODS: To investigate the putative neurodevelopmental consequences of DOXY administration in a clinically relevant animal model of HI, we performed a time-course analysis such that postnatal rat pups received DOXY (10mg/kg) or vehicle immediately before HI (n >or= 6). We then assessed cytogenesis, proinflammatory cytokines, brain-derived neurotrophic factor (BDNF) and matrix metalloproteinases regionally and longitudinally. RESULTS: We found that DOXY significantly inhibits neuroinflammation in the frontal cortex, striatum and hippocampus; decreases interleukin-1Beta (IL-1Beta) and tumour necrosis factor-alpha (TNF-alpha); and augments BDNF following HI. In addition, DOXY-treated pups have significantly fewer 2-bromo-5-deoxyuridine (BrdU)-positive cells in the subventricular zone 6 hours post-HI. However, DOXY does not persistently affect cytogenesis in the subventricular zone or dentate gyrus up to 7 days post-HI. The BrdU-positive cells not expressing markers for mature neurons colabel with nestin, an intermediate filament protein typical of neuronal precursors. LIMITATIONS: Our study investigates 'acute' neurodevelopment over the first 7 days of life after HI injury. Further long-term investigations into adulthood are underway. CONCLUSION: Taken together, our results suggest the putative clinical potential of DOXY in the management of neonatal cerebral HI injury. [ABSTRACT FROM AUTHOR]

Minimizing central nervous system (CNS) injury from preterm birth depends upon understanding the critical pathways that underlie essential neurodevelopmental and CNS pathophysiology. Signaling by chemokine (C-X-C motif) ligand 1 (CXCL1) through its cognate receptor, CXCR2 [(C-X-C motif) receptor 2] is essential for neurodevelopment. Increased CXCR2 signaling, however, is implicated in a variety of uterine and neuropathologies, and their role in the CNS injury associated with perinatal brain injury is poorly defined. To evaluate the long-term efficacy of CXCR2 blockade in functional repair of brain injury secondary to chorioamnionitis (CHORIO), we used an established preclinical rat model of cerebral palsy. We tested the hypothesis that transient postnatal CXCR2 antagonism with SB225002 would reduce gait deficits, hypermobility, hyperactivity, and disinhibition concomitant with repair of functional and anatomical white and gray matter injury. CHORIO was induced in pregnant Sprague Dawley rats on embryonic day 18 (E18). SB225002 (3 mg/kg) was administered intraperitoneally from postnatal day 1 (P1)-P5. Rats were aged to adulthood and tested for gait, open-field behavior and cognitive and executive function deficits using a touchscreen cognitive assessment platform. Results show that transient CXCR2 blockade attenuated microstructural white matter injury after CHORIO consistent with improved anatomical connectivity, and mitigated deficits in gait coordination, posture, balance, paw placement, and stepping (p < 0.05). Animals with CHORIO were hyperactive and hypermobile with fMRI deficits in neural circuitry central to cognition. However, CXCR2 antagonism in CHORIO animals did not normalize open-field behavior, neural activity, or cognition on a touchscreen task of discrimination learning (all p > 0.05). Studies in CXCR2 knockout mice confirmed significantly impaired cognitive performance independent of CHORIO. Taken together, transient postnatal blockade of CXCR2 ameliorates aspects of the lasting neural injury after CHORIO including normalizing gait deficits and white matter injury. However, improvement in essential functional and cognitive domains are not achieved limiting the utility of this therapeutic approach for treatment of perinatal brain injury. This study emphasizes the complex, multi-faceted role of chemokines in typical neurodevelopment, circuit formation, neural network function, and injury response., (© 2024 International Society for Neurochemistry.)

Hydrocephalus is a neurological disorder that impacts approximately 85 per 100,000 individuals worldwide and is associated with motor and cognitive impairments. While many advances in surgical interventions have helped substantially improve the survival rates and quality of life of those affected, there continues to be significant gaps in our understanding of the etiology of this heterogeneous condition as well as its specific neuropsychological and functional challenges across different phases of life. To address these limitations, the Hydrocephalus Association and Rudi Schulte Research Institute organized a workshop titled, "Improving Cognitive and Psychological Outcomes in Hydrocephalus", composed of top academics in the fields of hydrocephalus, cognition, and neuropsychology, as well as individuals with hydrocephalus or their caregivers. The purpose was to review the available evidence and propose pertinent areas of further research to improve the cognitive functioning, functional status, and quality of life of individuals with hydrocephalus. These topics included cognitive and neuropsychological assessments and daily-life function of children and adults living with hydrocephalus, biomarkers of cognitive function, animal modeling of hydrocephalus, and the longitudinal impact of hydrocephalus treatment. The following paper outlines four primary areas that warrant research: (1) neuropsychological phenotypes, (2) treatment-focused research considerations, (3) translational pre-clinical tools, and (4) establishing pathways for longitudinal care. Through the efforts of this group, the goal of this manuscript is to inspire and direct scientific and clinical inquiry towards these noted research priorities to further improve the lives of individuals with hydrocephalus and their families., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable, no human data was collected. Consent for publication: N/A. Competing interests: The authors declare no competing interests., (© 2024. The Author(s).)

Prenatal alcohol exposure (PAE) is associated with long-term neurodevelopmental deficits resulting in impaired executive functioning and motor control. Intriguingly, PAE has been linked with an increased risk of transient systemic hypoxia-ischemia (TSHI), which alone results in suboptimal fetal growth and neurodevelopmental consequences. Here, using two translationally relevant preclinical models, we investigated the short-term and lasting effects of PAE and TSHI on the morphology of the medial prefrontal cortex (mPFC), a region important in executive function, and tested whether PAE interacts with TSHI to produce a distinct pattern of injury relative to either condition alone. The four experimental groups included sham (saccharin water, no TSHI), PAE (5% alcohol, no TSHI), TSHI (saccharin water, TSHI), and PAE+TSHI (5% alcohol, TSHI). Brains were extracted for Golgi-Cox staining at Postnatal Day 35 (P35) or P100 and processed for 3D Sholl analysis. The analysis of the mPFC at P35 showed no significant differences in the number of branches or dendritic length overall, although the impact of TSHI compared to alcohol was significant for both. There were no significant differences in the number of Sholl intersections overall at P35, although a sex difference was noted in PAE offspring. At P100, analysis of filament dendritic length and branching number was also significantly impacted by TSHI compared to alcohol. Interestingly, sex was also a significant factor when assessing the impact of alcohol. PAE and TSHI both had an insignificantly increased number of Sholl intersections at P100 compared to the control. The observed changes to dendritic complexity at P100 demonstrate altered neuronal morphology in the mPFC that endure into adulthood. Given the importance of the mPFC in executive functioning, these pilot data provide insight into morphological changes that may contribute to the neurobehavioral deficits observed following exposure to PAE and TSHI and highlight the need for additional investigations into this area.

The impact of the opioid epidemic on pregnant people and children is a growing public health crisis. Understanding how opioids affect the developing brain during pregnancy and postnatally remains a critical area of investigation. Biological sex plays a crucial role in all physiologic processes, with the potential for a significant impact on neonatal outcomes, including those infants with opioid exposure. Here, we aim to explore current literature on the effect of sex on neonatal outcomes following prenatal opioid exposure. Sex differences in adults with opioid use disorder have been well studied, including increased mortality among males and higher rates of psychiatric comorbidities and likelihood of relapse in females. However, such differences are not yet well understood in neonates. Emerging clinical data suggest sex-specific effects in infants with prenatal opioid exposure on the expression of genes related to feeding regulation and reward signaling pathways. Increased susceptibility to white matter injury has also been noted in female infants following prenatal opioid exposure. Understanding the impact of sex as a biological variable on neonatal outcomes following prenatal opioid exposure is paramount to improving the health and well-being of infants, children, and adults impacted by the opioid epidemic., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2024 Madurai, Jantzie and Yen.)

Importance: Prenatal opioid exposure (POE) may alter with fetal development of the immune system, which may influence long-term health and susceptibility to immune-related conditions., Objective: To compare the risk of hospitalization and emergency department presentation for immune-related conditions in children with and without POE., Design, Setting, and Participants: This retrospective, population-based cohort study used linked administrative health records of all children born in Western Australia between January 1, 2003, and December 31, 2018 (N = 401 462)., Exposure: Prenatal exposure to prescription opioids (overall and by trimester), neonatal abstinence syndrome diagnosis, and opioid indication (pain or opioid use disorder [OUD])., Main Outcomes and Measures: The main outcome was hospital admissions and emergency department presentations during which a child was diagnosed with an immune-related condition, including infections, conditions associated with an overactive immune system (eg, asthma, eczema, and allergy and anaphylaxis), and autoimmune diseases diagnosed before age 5 years or June 30, 2020. Data were analyzed between August 30, 2022, and February 27, 2023., Results: Neonates with POE (1656 [0.4%]; mean [SD] gestational age, 37.7 [2.1] weeks; 836 females [50.5%]; 820 males [49.5%]) were more likely to be born preterm, have low birth weight for gestational age, and be coexposed to cigarette smoke compared with nonexposed neonates. Perinatal opioid exposure was associated with an increased risk of perinatal infection (adjusted odds ratio [AOR], 1.62; 95% CI, 1.38-1.90) and eczema and dermatitis (AOR, 11.91; 95% CI, 9.84-14.41) compared with nonexposure. Neonatal abstinence syndrome was also associated with both conditions (AOR, 2.91 [95% CI, 2.36-3.57] and 31.11 [95% CI, 24.64-39.28], respectively). Prenatal opioid exposure was also associated with an increased risk of childhood asthma (adjusted hazard ratio [AHR], 1.44; 95% CI, 1.16-1.79), but not allergies and anaphylaxis. It was also associated with an increased risk of childhood eczema and dermatitis, but only in children with POE from opioids used to treat OUD (AHR, 1.47; 95% CI, 1.08-1.99) rather than pain. In contrast, POE from opioids used for pain was associated with an increased risk of infection (AHR, 1.44; 95% CI, 1.32-1.58), but POE to opioids used to treat OUD was not. Autoimmune conditions were rare and were not observed to be associated with POE., Conclusions and Relevance: In this cohort study, POE was associated with an increased risk of infection, eczema and dermatitis, and asthma, but not allergies and anaphylaxis or autoimmune conditions. These findings highlight the importance of further study of opioid-induced immune changes during pregnancy, the potential impact on long-term health in exposed children, and the mechanisms of opioid-induced immune dysregulation.

Objectives: Lemniscal (motor-related) and spinothalamic (neuropathic pain-related) somatosensory abnormalities affect different subsets of adults with cerebral palsy (CP). Lemniscal/motor abnormalities are associated with posterior thalamic radiation white matter disruption in individuals with CP and white matter injury. We tested the hypothesis that neuropathic pain symptoms in this population are rather associated with injury of the somatosensory (posterior group nuclei) thalamus., Methods: In this cross-sectional study, communicative adults with CP and bilateral white matter injury and neurotypical control participants volunteered to self-report pain symptoms and undergo research MRI. Posterior group thalamic nuclei volume was computed and correlated against neuropathic pain scores., Results: Participants with CP (n=6) had, on average, 24% smaller posterior group thalamic volumes (95% CI [10-39%]; corrected p=0.01) than control participants. More severe volume loss was correlated with more severe neuropathic pain scores (ρ=-0.87 [-0.99,-0.20]; p=0.02)., Discussion: Association with thalamic volume loss suggests that neuropathic pain in adults with CP may frequently be central neuropathic pain. Complementing assessments of white matter microstructure, regional brain volumes hold promise as diagnostic biomarkers for central neuropathic pain in individuals with structural brain disorders.

Background: Cutting-edge neonatal programs diagnose cerebral palsy (CP) or "high risk of CP" using validated neurobehavioral exams in combination with risk history and neuroimaging. In rat models, digital gait analyses are the gold standard adult assessment, but tools in infant rats are limited. Refinement of infant rat neurobehavioral correlates of CP will establish translational behavioral biomarkers to delineate early mechanisms of CP in both humans and rodent models of CP., Objective: To facilitate precision medicine approaches of neurodevelopmental health and integrate basic and clinical research approaches for CP, we developed and piloted a new assay of neonatal rat neurobehavior to mimic human neonate exams., Methods: Our established rat model of CP secondary to chorioamnionitis (CHORIO) that induces bilateral motor impairment reminiscent of spastic CP was used. On postnatal day 10 (P10), 5 min videos were recorded of 26 (6 sham and 20 CHORIO) animals moving freely in a cage were reviewed by an evaluator trained in the human General Movements Assessment (GMA). Non-blinded observation revealed two behaviors that differed between rat pups in each group (time spent rearing; multi-dimensional nose sweeping; and sniffing). Each video was re-coded for these criteria by an evaluator blind to group status. Differences between sham and CP groups were analyzed using a Mann-Whitney U-test or Student's t -test ( p < 0.05 level of significance)., Results: Neonatal rats with CP exhibited sensorimotor impairment and decreased spatial exploration. CP rats spent significantly less time rearing (17.85 ± 1.60 s vs. 34.8 ± 2.89 s, p = 0.007) and engaged in multi-dimensional nose sweeping and sniffing (2.2 ± 0.58 episodes vs. 5.5 ± 0.96 episodes, p = 0.03) than sham controls., Conclusions: These pilot findings of harmonized translational and precision biobehavioral assays provide an opportunity for increased expediency of clinical trials at the earliest stages of brain development.

Introduction: Novel therapeutics are emerging to mitigate damage from perinatal brain injury (PBI). Few newborns with PBI suffer from a singular etiology. Most experience cumulative insults from prenatal inflammation, genetic and epigenetic vulnerability, toxins (opioids, other drug exposures, environmental exposure), hypoxia-ischemia, and postnatal stressors such as sepsis and seizures. Accordingly, tailoring of emerging therapeutic regimens with endogenous repair or neuro-immunomodulatory agents for individuals requires a more precise understanding of ligand, receptor-, and non-receptor-mediated regulation of essential developmental hormones. Given the recent clinical focus on neurorepair for PBI, we hypothesized that there would be injury-induced changes in erythropoietin (EPO), erythropoietin receptor (EPOR), melatonin receptor (MLTR), NAD-dependent deacetylase sirtuin-1 (SIRT1) signaling, and hypoxia inducible factors (HIF1α, HIF2α). Specifically, we predicted that EPO, EPOR, MLTR1, SIRT1, HIF1α and HIF2α alterations after chorioamnionitis (CHORIO) would reflect relative changes observed in human preterm infants. Similarly, we expected unique developmental regulation after injury that would reveal potential clues to mechanisms and timing of inflammatory and oxidative injury after CHORIO that could inform future therapeutic development to treat PBI. Methods: To induce CHORIO, a laparotomy was performed on embryonic day 18 (E18) in rats with transient uterine artery occlusion plus intra-amniotic injection of lipopolysaccharide (LPS). Placentae and fetal brains were collected at 24 h. Brains were also collected on postnatal day 2 (P2), P7, and P21. EPO, EPOR, MLTR1, SIRT1, HIF1α and HIF2α levels were quantified using a clinical electrochemiluminescent biomarker platform, qPCR, and/or RNAscope. MLT levels were quantified with liquid chromatography mass spectrometry. Results: Examination of EPO, EPOR, and MLTR1 at 24 h showed that while placental levels of EPO and MLTR1 mRNA were decreased acutely after CHORIO, cerebral levels of EPO, EPOR and MLTR1 mRNA were increased compared to control. Notably, CHORIO brains at P2 were SIRT1 mRNA deficient with increased HIF1α and HIF2α despite normalized levels of EPO, EPOR and MLTR1, and in the presence of elevated serum EPO levels. Uniquely, brain levels of EPO, EPOR and MLTR1 shifted at P7 and P21, with prominent CHORIO-induced changes in mRNA expression. Reductions at P21 were concomitant with increased serum EPO levels in CHORIO rats compared to controls and variable MLT levels. Discussion: These data reveal that commensurate with robust inflammation through the maternal placental-fetal axis, CHORIO impacts EPO, MLT, SIRT1, and HIF signal transduction defined by dynamic changes in EPO, EPOR, MLTR1, SIRT1, HIF1α and HIF2α mRNA, and EPO protein. Notably, ligand-receptor mismatch, tissue compartment differential regulation, and non-receptor-mediated signaling highlight the importance, complexity and nuance of neural and immune cell development and provide essential clues to mechanisms of injury in PBI. As the placenta, immune cells, and neural cells share many common, developmentally regulated signal transduction pathways, further studies are needed to clarify the perinatal dynamics of EPO and MLT signaling and to capitalize on therapies that target endogenous neurorepair mechanisms., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Kitase, Madurai, Hamimi, Hellinger, Odukoya, Ramachandra, Muthukumar, Vasan, Sevensky, Kirk, Gall, Heck, Ozen, Orsburn, Robinson and Jantzie.)

Microglia morphological studies have been limited to the process of reviewing the most common characteristics of a group of cells to conclude the likelihood of a "pathological" milieu. We have developed an Imaris-software-based analytical pipeline to address selection and operator biases, enabling use of highly reproducible machine-learning algorithms to quantify at single-cell resolution differences between groups. We hypothesized that this analytical pipeline improved our ability to detect subtle yet important differences between groups. Thus, we studied the temporal changes in Iba1 + microglia-like cell (MCL) populations in the CA1 between P10-P11 and P18-P19 in response to intrauterine growth restriction (IUGR) at E12.5 in mice, chorioamnionitis (chorio) at E18 in rats and neonatal hypoxia-ischemia (HI) at P10 in mice. Sholl and convex hull analyses differentiate stages of maturation of Iba1 + MLCs. At P10-P11, IUGR or HI MLCs were more prominently 'ameboid', while chorio MLCs were hyper-ramified compared to sham. At P18-P19, HI MLCs remained persistently 'ameboid' to 'transitional'. Thus, we conclude that this unbiased analytical pipeline, which can be adjusted to other brain cells (i.e., astrocytes), improves sensitivity to detect previously elusive morphological changes known to promote specific inflammatory milieu and lead to worse outcomes and therapeutic responses.

The opioid epidemic is an ongoing public health crisis, and children born following prenatal opioid exposure (POE) have increased risk of long-term cognitive and behavioral sequelae. Clinical studies have identified reduced gray matter volume and abnormal white matter microstructure in children with POE but impacts on whole-brain functional brain connectivity (FC) have not been reported. To define effects of POE on whole brain FC and white matter injury in adult animals, we performed quantitative whole-brain structural and functional MRI. We used an established rat model of POE in which we have previously reported impaired executive function in adult rats analogous to persistent neurocognitive symptoms described in humans with POE. Pregnant Sprague-Dawley rat dams received continuous methadone (12 mg/kg/day) vs. saline infusion for 28 days via osmotic mini-pumps, exposing rats to pre- and postnatal opioid until weaning. At young adult age (P60), POE and saline exposed offspring underwent in vivo MRI included diffusion tensor imaging and functional MRI (fMRI). Results indicate that fractional anisotropy (FA) was decreased in adult animals with POE [ n  = 11] compared to animals that received saline [ n  = 9] in major white matter tracts, including the corpus callosum ( p  < 0.001) and external capsule ( p  < 0.01). This change in FA was concomitant with reduced axial diffusivity in the external capsule ( p  < 0.01) and increased radial diffusivity in the corpus callosum ( p  < 0.01). fMRI analyses reveal brainwide FC was diffusely lower in POE ( p  < 10 -6 ; 10% of variance explained by group). Decreased connectivity in cortical-cortical and cortico-basal ganglia circuitry was particularly prominent with large effect sizes (Glass's Δ > 1). Taken together, these data confirm POE reduces brainwide functional connectivity as well as microstructural integrity of major white matter tracts. Altered neural circuitry, dysregulated network refinement, and diffuse network dysfunction have been implicated in executive function deficits that are common in children with POE. FC may serve as a translatable biomarker in children with POE., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2023 Chin, Kitase, Madurai, Robinson and Jantzie.)

The Hydrocephalus Association (HA) workshop, Driving Common Pathways: Extending Insights from Posthemorrhagic Hydrocephalus, was held on November 4 and 5, 2019 at Washington University in St. Louis. The workshop brought together a diverse group of basic, translational, and clinical scientists conducting research on multiple hydrocephalus etiologies with select outside researchers. The main goals of the workshop were to explore areas of potential overlap between hydrocephalus etiologies and identify drug targets that could positively impact various forms of hydrocephalus. This report details the major themes of the workshop and the research presented on three cell types that are targets for new hydrocephalus interventions: choroid plexus epithelial cells, ventricular ependymal cells, and immune cells (macrophages and microglia)., (© 2023. The Author(s).)

Immunoperinatology is an emerging field. Transdisciplinary efforts by physicians, physician-scientists, basic science researchers, and computational biologists have made substantial advancements by identifying unique immunologic signatures of specific diseases, discovering innovative preventative or treatment strategies, and establishing foundations for individualized neonatal intensive care of the most vulnerable neonates. In this review, we summarize the immunobiology and immunopathology of pregnancy, highlight omics approaches to study the maternal-fetal interface, and their contributions to pregnancy health. We examined the importance of transdisciplinary, multiomic (such as genomics, transcriptomics, proteomics, metabolomics, and immunomics) and machine-learning strategies in unraveling the mechanisms of adverse pregnancy, neonatal, and childhood outcomes and how they can guide the development of novel therapies to improve maternal and neonatal health. IMPACT: Discuss immunoperinatology research from the lens of omics and machine-learning approaches. Identify opportunities for omics-based approaches to delineate infection/inflammation-associated maternal, neonatal, and later life adverse outcomes (e.g., histologic chorioamnionitis [HCA])., (© 2022. The Author(s), under exclusive licence to the International Pediatric Research Foundation, Inc.)

Preterm birth is a principal cause of neurological disability later in life, including cognitive and behavioral deficits. Notably, cognitive impairment has greater impact on quality of life than physical disability. Survivors of preterm birth commonly have deficits of executive function. Difficulties with tasks and planning complexity correlate positively with increasing disability. To overcome these barriers for children born preterm, preclinical and clinical studies have emphasized the importance of neurorestoration. Erythropoietin (EPO) is a endogenous cytokine with multiple beneficial mechanisms of action following perinatal brain injury. While most preclinical investigations have focused on pathology and molecular mechanisms, translational studies of repair using clinically viable biobehavioral biomarkers are still lacking. Here, using an established model of encephalopathy of prematurity secondary to placental insufficiency, we tested the hypothesis that administration of EPO in the neonatal period would attenuate deficits in recognition memory and cognitive flexibility in adult rats of both sexes. We assessed cognition and executive function in two ways. First, using the classic test of novel object recognition and second, using a touchscreen platform. Touchscreen testing allows for rigorous testing of cognition and executive function in preclinical and clinical scenarios. Data show that adult rats exhibit deficits in recognition memory and cognitive flexibility following in utero placental insufficiency. Notably, neonatal treatment of EPO attenuates these deficits in adulthood and facilitates functional repair. Together, these data validate EPO neurorestoration using a clinically relevant outcome measure and support the concept that postnatal treatment following in utero injury can improve cognition and executive function through adulthood., (© 2021 Wiley Periodicals LLC.)

Multifactorial diseases are characterized by inter-individual variation in etiology, age of onset, and penetrance. These diseases tend to be relatively common and arise from the combined action of genetic and environmental factors; however, parsing the convoluted mechanisms underlying these gene-by-environment interactions presents a significant challenge to their study and management. For neurodegenerative disorders, resolving this challenge is imperative, given the enormous health and societal burdens they impose. The mechanisms by which genetic and environmental effects may act in concert to destabilize homeostasis and elevate risk has become a major research focus in the study of common disease. Emphasis is further being placed on determining the extent to which a unifying biological principle may account for the progressively diminishing capacity of a system to buffer disease phenotypes, as risk for disease increases. Data emerging from studies of common, neurodegenerative diseases are providing insights to pragmatically connect mechanisms of genetic and environmental risk that previously seemed disparate. In this review, we discuss evidence positing inflammation as a unifying biological principle of homeostatic destabilization affecting the risk, onset, and progression of neurodegenerative diseases. Specifically, we discuss how genetic variation associated with Alzheimer disease and Parkinson disease may contribute to pro-inflammatory responses, how such underlying predisposition may be exacerbated by environmental insults, and how this common theme is being leveraged in the ongoing search for effective therapeutic interventions., (© 2022. The Author(s).)

Posthemorrhagic hydrocephalus of prematurity (PHHP) remains a vexing problem for patients, their families, and the healthcare system. The complexity of the pathogenesis of PHHP also presents a unique challenge within the fields of neonatology, neurology and neurosurgery. Here we focus on pathogenesis of PHHP and its impact on the development of CSF dynamics including choroid plexus, ependymal motile cilia and glymphatic system. PHHP is contrasted with infantile hydrocephalus from other etiologies, and with other types of posthemorrhagic hydrocephalus that occur later in life. The important concept that distinguishing ventricular volume from brain health and function is highlighted. The influence of the pathogenesis of PHHP on current interventions is reviewed, with particular emphasis on how the unique pathogenesis of PHHP contributes to the high rate of failure of current existing interventions. Finally, we discuss emerging interventions. A thorough understanding of the pathogenesis of PHHP is essential to developing effective non-surgical therapeutics to prevent the transformation from severe IVH to PHHP., Competing Interests: Conflict of Interest The authors declare no conflict of interest, (Copyright © 2022 Elsevier Inc. All rights reserved.)

The use of touchscreen technology to evaluate cognitive deficits in animal models has grown tremendously over the past 20 years. The touchscreen apparatus encompasses many advantages, namely a high level of standardization and translational capability. Improvements in technology in recent years have expanded the versatility of the touchscreen platform, as it is able to test distinct cognitive modalities including working memory, attention, discrimination, and association. Importantly, touchscreen technology has allowed researchers to explore deficits in multiple pillars of cognition in a wide variety of perinatal disorders with neurological sequelae across critical developmental windows. The touchscreen platform has been used to dissect deficits in antenatal CNS injury including fetal alcohol syndrome, prenatal opioid exposure, and chorioamnionitis, to peripartum insults such as term hypoxic-ischemic encephalopathy, to early postnatal insults including infantile traumatic brain injury. Most importantly, touchscreen technology offers the sensitivity necessary to detect subtle injury and treatment-induced changes in cognition and executive function beyond those offered by more rudimentary tests of rodent cognition. Understanding the pathophysiology of these disorders in rodents is paramount to addressing these deficits in human infants and dissecting the neural circuitry essential to perinatal brain injury pathophysiology and responsiveness to novel therapeutics. Touchscreen testing provides an effective, facile, sophisticated technique to accelerate the goal of improving cognitive and behavioral outcomes of children who suffer perinatal brain injury., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Cerebral palsy (CP) is the most common cause of physical disability for children worldwide. Many infants and toddlers are not diagnosed with CP until they fail to achieve obvious motor milestones. Currently, there are no effective pharmacologic interventions available for infants and toddlers to substantially improve their trajectory of neurodevelopment. Because children with CP from preterm birth also exhibit a sustained immune system hyper-reactivity, we hypothesized that neuro-immunomodulation with a regimen of repurposed endogenous neurorestorative medications, erythropoietin (EPO) and melatonin (MLT), could improve this trajectory. Thus, we administered EPO + MLT to rats with CP during human infant-toddler equivalency to determine whether we could influence gait patterns in mature animals. After a prenatal injury on embryonic day 18 (E18) that mimics chorioamnionitis at ∼25 weeks human gestation, rat pups were born and raised with their dam. Beginning on postnatal day 15 (P15), equivalent to human infant ∼1 year, rats were randomized to receive either a regimen of EPO + MLT or vehicle (sterile saline) through P20. Gait was assessed in young adult rats at P30 using computerized digital gait analyses including videography on a treadmill. Results indicate that gait metrics of young adult rats treated with an infantile cocktail of EPO + MLT were restored compared to vehicle-treated rats (p < 0.05) and similar to sham controls. These results provide reassuring evidence that pharmacological interventions may be beneficial to infants and toddlers who are diagnosed with CP well after the traditional neonatal window of intervention., (© 2022 S. Karger AG, Basel.)

Opioid use during pregnancy continues to rise at alarming rates with a parallel trend in the number of infants and children exposed to opioid medications each year. Prenatal opioid exposure (POE) occurs at a critical timepoint in neurodevelopment disrupting intricate pathways essential for neural-immune maturation with the potential for devastating long-term consequences. Understanding the mechanisms underlying injury associated with POE is essential to address long-term outcomes and identify diagnostic and therapeutic biomarkers in this vulnerable patient population. Using an established preclinical model of POE, we investigated changes in cerebral and peripheral inflammation and peripheral blood mononuclear cell (PBMC) activity. We hypothesized that neuroinflammation, as defined by changes in specific cerebral immune cell populations, would exist in adult rats following POE concomitant with sustained peripheral immune hyperreactivity (SPIHR). Our data demonstrated alterations in cerebral immune cells at postnatal day 60 (P60) typified by increased regulatory T cells ( p < 0.01) and neutrophils ( p < 0.05) in rats with POE compared to controls. Evaluation of serum revealed increased levels of IL-6 ( p < 0.05) and CXCL1 ( p < 0.05) at P21 in rats with POE compared to controls with no significant difference in cytokine or chemokine levels between the two groups at P60. Additionally, PBMCs isolated from rats with POE at P21 demonstrated baseline hypersecretion of IL-6 ( p < 0.01) and SPIHR with increased levels of TNF-α ( p < 0.05) and CXCL1 ( p < 0.05) following stimulation with LPS. At P60, however, there was no significant difference found in cytokine or chemokine levels secreted by PBMCs isolated from rats with POE at baseline or with LPS stimulation when compared to controls. Taken together, these data demonstrate cerebral inflammation months after prenatal opioid exposure and long after the resolution of systemic inflammation and SPIHR seen at toddler age equivalent. Chronic alterations in the cerebral immune cell populations secondary to prenatal opioid exposure may underly long-term consequences of developmental brain injury including deficits in cognition and attention. These findings may be invaluable to further investigations of precise biomarkers of injury and targeted therapeutics for this vulnerable population., Competing Interests: Conflict of interest The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Pregnancy is an inflammatory process that is carefully regulated by the placenta via immunomodulation and cell-to-cell communication of maternal and fetal tissues. Exosomes, types of extracellular vesicles, facilitate the intercellular communication and traffic biologically modifying cargo within the maternal-placental-fetal axis in normal and pathologic pregnancies. Chorioamnionitis is characterized by inflammation of chorioamniotic membranes that produces systemic maternal and fetal inflammatory responses of cytokine dysregulation and has been associated with brain injury and neurodevelopmental disorders. This review focuses on how pathologic placental exosomes propagate acute and chronic inflammation leading to brain injury. The evidence reviewed here highlights the need to investigate exosomes from pathologic pregnancies and those with known brain injury to identify new diagnostics, biomarkers, and potential therapeutic targets., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Background: Chorioamnionitis (CHORIO) is a principal risk factor for preterm birth and is the most common pathological abnormality found in the placentae of preterm infants. CHORIO has a multitude of effects on the maternal-placental-fetal axis including profound inflammation. Cumulatively, these changes trigger injury in the developing immune and central nervous systems, thereby increasing susceptibility to chronic sequelae later in life. Despite this and reports of neural-immune changes in children with cerebral palsy, the extent and chronicity of the peripheral immune and neuroinflammatory changes secondary to CHORIO has not been fully characterized., Methods: We examined the persistence and time course of peripheral immune hyper-reactivity in an established and translational model of perinatal brain injury (PBI) secondary to CHORIO. Pregnant Sprague-Dawley rats underwent laparotomy on embryonic day 18 (E18, preterm equivalent). Uterine arteries were occluded for 60 min, followed by intra-amniotic injection of lipopolysaccharide (LPS). Serum and peripheral blood mononuclear cells (PBMCs) were collected at young adult (postnatal day P60) and middle-aged equivalents (P120). Serum and PBMCs secretome chemokines and cytokines were assayed using multiplex electrochemiluminescent immunoassay. Multiparameter flow cytometry was performed to interrogate immune cell populations., Results: Serum levels of interleukin-1β (IL-1β), IL-5, IL-6, C-X-C Motif Chemokine Ligand 1 (CXCL1), tumor necrosis factor-α (TNF-α), and C-C motif chemokine ligand 2/monocyte chemoattractant protein-1 (CCL2/MCP-1) were significantly higher in CHORIO animals compared to sham controls at P60. Notably, CHORIO PBMCs were primed. Specifically, they were hyper-reactive and secreted more inflammatory mediators both at baseline and when stimulated in vitro. While serum levels of cytokines normalized by P120, PBMCs remained primed, and hyper-reactive with a robust pro-inflammatory secretome concomitant with a persistent change in multiple T cell populations in CHORIO animals., Conclusions: The data indicate that an in utero inflammatory insult leads to neural-immune changes that persist through adulthood, thereby conferring vulnerability to brain and immune system injury throughout the lifespan. This unique molecular and cellular immune signature including sustained peripheral immune hyper-reactivity (SPIHR) and immune cell priming may be a viable biomarker of altered inflammatory responses following in utero insults and advances our understanding of the neuroinflammatory cascade that leads to perinatal brain injury and later neurodevelopmental disorders, including cerebral palsy., (© 2021. The Author(s).)

Globally, approximately 11% of all infants are born preterm, prior to 37 weeks' gestation. In these high-risk neonates, encephalopathy of prematurity (EoP) is a major cause of both morbidity and mortality, especially for neonates who are born very preterm (<32 weeks gestation). EoP encompasses numerous types of preterm birth-related brain abnormalities and injuries, and can culminate in a diverse array of neurodevelopmental impairments. Of note, posthemorrhagic hydrocephalus of prematurity (PHHP) can be conceptualized as a severe manifestation of EoP. PHHP impacts the immature neonatal brain at a crucial timepoint during neurodevelopment, and can result in permanent, detrimental consequences to not only cerebrospinal fluid (CSF) dynamics, but also to white and gray matter development. In this review, the relevant literature related to the diverse mechanisms of cell death in the setting of PHHP will be thoroughly discussed. Loss of the epithelial cells of the choroid plexus, ependymal cells and their motile cilia, and cellular structures within the glymphatic system are of particular interest. Greater insights into the injuries, initiating targets, and downstream signaling pathways involved in excess cell death shed light on promising areas for therapeutic intervention. This will bolster current efforts to prevent, mitigate, and reverse the consequential brain remodeling that occurs as a result of hydrocephalus and other components of EoP.

Background: The presence of hyperphosphorylated microtubule-associated protein tau is strongly correlated with cognitive decline and neuroinflammation in Alzheimer's disease and related tauopathies. However, the role of inflammation and anti-inflammatory interventions in tauopathies is unclear. Our goal was to determine if removing anti-inflammatory interleukin-10 (IL-10) during an acute inflammatory challenge has any effect on neuronal tau pathology., Methods: We induce systemic inflammation in Il10-deficient (Il10 -/- ) versus Il10 +/+ (Non-Tg) control mice using a single intraperitoneal (i.p.) injection of lipopolysaccharide (LPS) to examine microglial activation and abnormal hyperphosphorylation of endogenous mouse tau protein. Tau phosphorylation was quantified by Western blotting and immunohistochemistry. Microglial morphology was quantified by skeleton analysis. Cytokine expression was determined by multiplex electro chemiluminescent immunoassay (MECI) from Meso Scale Discovery (MSD)., Results: Our findings show that genetic deletion of Il10 promotes enhanced neuroinflammation and tau phosphorylation. First, LPS-induced tau hyperphosphorylation was significantly increased in Il10 -/- mice compared to controls. Second, LPS-treated Il10 -/- mice showed signs of neurodegeneration. Third, LPS-treated Il10 -/- mice showed robust IL-6 upregulation and direct treatment of primary neurons with IL-6 resulted in tau hyperphosphorylation on Ser396/Ser404 site., Conclusions: These data support that loss of IL-10 activates microglia, enhances IL-6, and leads to hyperphosphorylation of tau on AD-relevant epitopes in response to acute systemic inflammation., (© 2021. The Author(s).)

Heme oxygenase-1 (HO-1) is an evolutionarily conserved stress response enzyme and important in pregnancy maintenance, fetal and neonatal outcomes, and a variety of pathologic conditions. Here, we investigated the effects of an exposure to systemic inflammation late in gestation [embryonic day (E)15.5] on wild-type (Wt) and HO-1 heterozygous (Het, HO-1+/-) mothers, fetuses, and offspring. We show that alterations in fetal liver and spleen HO homeostasis during inflammation late in gestation can lead to a sustained dysregulation of innate immune cell populations and intracellular myeloid HO-1 expression in the spleen through young adolescence [postnatal day 25] in mice., Competing Interests: The authors have declared no competing interests exist.

Chorioamnionitis (CHORIO), placental insufficiency, and preterm birth are well-known antecedents of perinatal brain injury (PBI). Heme-oxygenase-1 (HO-1) is an important inducible enzyme in oxidative and inflammatory conditions. In the brain, HO-1 and the iron regulatory receptor, transferrin receptor-1 (TfR1), are known to be involved in iron homeostasis, oxidative stress, and cellular adaptive mechanisms. However, the role of HO pathway in the pathophysiology of PBI has not been previously studied. In this study, we set out to define the ontogeny of the HO pathway in the brain and determine if CHORIO changed its normal developmental regulation. We also aimed to determine the role of HO-1/TfR1 in CHORIO-induced neuroinflammation and peripheral inflammation in a clinically relevant rat model of PBI. We show that HO-1, HO-2, and TfR1 expression are developmentally regulated in the brain during the perinatal period. CHORIO elevates HO-1 and TfR1 mRNA expression in utero and in the early postnatal period and results in sustained increase in HO-1/TfR1 ratios in the brain. This is associated with neuroinflammatory and peripheral immune phenotype supported by a significant increase in brain mononuclear cells and peripheral blood double negative T cells suggesting a role of HO-1/TfR1 pathway dysregulation in CHORIO-induced neuroinflammation.

Substance use, specifically the use of prescription and non-prescription opioids among pregnant women, is a major public health issue and chief contributor to the opioid crisis. The prevalence of Neonatal Opioid Withdrawal Syndrome has risen 5-fold in the past decade, and is a well-recognized consequence of perinatal opioid exposure. By contrast, the long-term damage to the developing brain from opioid medications is just beginning to be recognized as a serious concern. Published data suggest that opioid exposure commencing in utero negatively affects the maturation of the neural-immune system, and trajectory of central nervous system development. Methadone induces peripheral immune hyper-reactivity, lasting structural and microstructural brain injury, and significant deficits in executive function and cognitive control in adult animals following in utero exposure. Thus, to address the cascading public health crisis stemming from the multitude of infants with in utero opioid exposure who will grow up with altered neurodevelopmental trajectories, rigorous preclinical, mechanistic studies are required. Such studies will define the long-term sequelae of prenatal opioid exposure in an effort to develop appropriate and targeted interventions. Specifically, the development of novel fluid, neuroimaging and biobehavioral biomarkers will be the most useful to aid in early identification and treatment of opioid exposed infants with the greatest risk of poor clinical outcomes. These studies will be essential to understand how in utero insults determine brain structure and function in adulthood, and what targeted interventions will be required to improve long-term outcomes in the countless children being born exposed to opioids each year., Competing Interests: None

Chronic pain is prevalent in adults with cerebral palsy. We aimed to explore associations between chronic pain and somatosensory, motor, cognitive, etiologic, and environmental factors in adults with cerebral palsy. This cross-sectional study enrolled 17 adult participants with cerebral palsy (mean age 31 years; 8 female; Gross Motor Functional Classification Status levels I-V) able to self-report and 10 neurotypical adult volunteers (mean age 34 years; 9 female). Participants reported pain characteristics, demographics, and affective factors. Physical examination included somatosensory and motor evaluation. Between-group comparisons used a ranksum test, and correlation analyses estimated effect size in terms of shared variance (ρ 2 ). Individuals with cerebral palsy reported greater pain intensity, neuropathic qualities, and nociceptive qualities than control participants. Higher pain intensity was associated with female gender (ρ 2 = 16%), anxiety/depression symptoms (ρ 2 = 10%), and lower household income (ρ 2 = 19%). It was also associated with better communicative ability (ρ 2 = 21%), spinothalamic (sharp/temperature) sensory abnormalities (ρ 2 = 33%), and a greater degree of prematurity (ρ 2 = 17%). This study highlights similarity of chronic pain associations in people with cerebral palsy with patterns seen in other populations with chronic pain. Spinothalamic sensory abnormalities suggest central pain mechanisms., (Copyright © 2020 Chin, Lenz, Ye, Campbell, Stashinko, Jantzie, Gerner, Hoon and Robinson.)

The ongoing worldwide pandemic of the novel human coronavirus SARS-CoV-2 and the ensuing disease, COVID-19, has presented enormous and unprecedented challenges for all medical specialists. However, to date, children, especially neonates, have been relatively spared from the devastating consequences of this infection. Neurologic involvement is being increasingly recognized among adults with COVID-19, who can develop sensory deficits in smell and taste, delirium, encephalopathy, headaches, strokes, and peripheral nervous system disorders. Among neonates and children, COVID-19-associated neurological manifestations have been relatively rare, yet reports involving neurologic dysfunction in this age range are increasing. As discussed in this review, pediatric neurologists and other pediatric specialists should be alert to potential neurological involvement by this virus, which might have neuroinvasive capability and carry long-term neuropsychiatric and medical consequences.

Previous reports show that moderate prenatal alcohol exposure (PAE) poses a risk factor for developing neuropathic pain following adult-onset peripheral nerve injury in male rats. Recently, evidence suggests that immune-related mechanisms underlying neuropathic pain in females are different compared to males despite the fact that both sexes develop neuropathy of similar magnitude and duration following chronic constriction injury (CCI) of the sciatic nerve. Data suggest that the actions of peripheral T cells play a greater role in mediating neuropathy in females. The goal of the current study is to identify specificity of immune cell and cytokine changes between PAE and non-PAE neuropathic females by utilizing a well-characterized rodent model of sciatic nerve damage, in an effort to unmask unique signatures of immune-related factors underlying the risk of neuropathy from PAE. Cytokines typically associated with myeloid cell actions such as interleukin (IL)-1β, tumor necrosis factor (TNF), IL-6, IL-4 and IL-10 as well as the neutrophil chemoattractant CXCL1, are examined. In addition, transcription factors and cytokines associated with various differentiated T cell subtypes are examined (anti-inflammatory FOXP3, proinflammatory IL-17A, IL-21, ROR-γt, interferon (IFN)-γ and T-bet). Lymphocyte function associated antigen 1 (LFA-1) is an adhesion molecule expressed on peripheral immune cells including T cells, and regulates T cell activation and extravasation into inflamed tissue regions. A potential therapeutic approach was explored with the goal of controlling proinflammatory responses in neuroanatomical regions critical for CCI-induced allodynia by blocking LFA-1 actions using BIRT377. The data show profound development of hindpaw allodynia in adult non-PAE control females following standard CCI, but not following minor CCI, while minor CCI generated allodynia in PAE females. The data also show substantial increases in T cell-associated proinflammatory cytokine mRNA and proteins, along with evidence of augmented myeloid/glial activation (mRNA) and induction of myeloid/glial-related proinflammatory cytokines, CCL2, IL-1β and TNF in discrete regions along the pain pathway (damaged sciatic nerve, dorsal root ganglia; DRG, and spinal cord). Interestingly, the characteristic anti-inflammatory IL-10 protein response to nerve damage is blunted in neuropathic PAE females. Moreover, T cell profiles are predominantly proinflammatory in neuropathic Sac and PAE females, augmented levels of Th17-specific proinflammatory cytokines IL-17A and IL-21, as well as the Th1-specific factor, T-bet, are observed. Similarly, the expression of RORγt, a critical transcription factor for Th17 cells, is detected in the spinal cord of neuropathic females. Blocking peripheral LFA-1 actions with intravenous (i.v.) BIRT377 reverses allodynia in Sac and PAE rats, dampens myeloid (IL-1β, TNF, CXCL1)- and T cell-associated proinflammatory factors (IL-17A and RORγt) and spinal glial activation. Moreover, i.v. BIRT377 treatment reverses the blunted IL-10 response to CCI observed only in neuropathic PAE rats and elevates FOXP3 in pain-reversed Sac rats. Unexpectedly, intrathecal BIRT377 treatment is unable to alter allodynia in either Sac or PAE neuropathic females. Together, these data provide evidence that: 1) fully differentiated proinflammatory Th17 cells recruited at the sciatic nerve, DRGs and lumbar spinal cord may interact with the local environment to shape the immune responses underlying neuropathy in female rats, and, 2) PAE primes peripheral and spinal immune responses in adult females. PAE is a risk factor in females for developing peripheral neuropathy after minor nerve injury., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

The increased incidence of opioid use during pregnancy warrants investigation to reveal the impact of opioid exposure on the developing fetus. Exposure during critical periods of development could have enduring consequences for affected individuals. Particularly, evidence is mounting that developmental injury can result in immune priming, whereby subsequent immune activation elicits an exaggerated immune response. This maladaptive hypersensitivity to immune challenge perpetuates dysregulated inflammatory signaling and poor health outcomes. Utilizing an established preclinical rat model of perinatal methadone exposure, we sought to investigate the consequences of developmental opioid exposure on in vitro activation of peripheral blood mononuclear cells (PBMCs). We hypothesize that PBMCs from methadone-exposed rats would exhibit abnormal chemokine and cytokine expression at baseline, with exaggerated chemokine and cytokine production following immune stimulation compared to saline-exposed controls. On postnatal day (P) 7, pup PMBCs were isolated and cultured, pooling three pups per n . Following 3 and 24 h, the supernatant from cultured PMBCs was collected and assessed for inflammatory cytokine and chemokine expression at baseline or lipopolysaccharide (LPS) stimulation using multiplex electrochemiluminescence. Following 3 and 24 h, baseline production of proinflammatory chemokine and cytokine levels were significantly increased in methadone PBMCs ( p < 0.0001). Stimulation with LPS for 3 h resulted in increased tumor necrosis factor (TNF-α) and C-X-C motif chemokine ligand 1 (CXCL1) expression by 3.5-fold in PBMCs from methadone-exposed PBMCs compared to PBMCs from saline-exposed controls ( p < 0.0001). Peripheral blood mononuclear cell hyperreactivity was still apparent at 24 h of LPS stimulation, evidenced by significantly increased TNF-α, CXCL1, interleukin 6 (IL-6), and IL-10 production by methadone PMBCs compared to saline control PBMCs ( p < 0.0001). Together, we provide evidence of increased production of proinflammatory molecules from methadone PBMCs at baseline, in addition to sustained hyperreactivity relative to saline-exposed controls. Exaggerated peripheral immune responses exacerbate inflammatory signaling, with subsequent consequences on many organ systems throughout the body, such as the developing nervous system. Enhanced understanding of these inflammatory mechanisms will allow for appropriate therapeutic development for infants who were exposed to opioids during development. Furthermore, these data highlight the utility of this in vitro PBMC assay technique for future biomarker development to guide specific treatment for patients exposed to opioids during gestation., (Copyright © 2020 Newville, Maxwell, Kitase, Robinson and Jantzie.)

Neonatal hypoxic-ischemic encephalopathy (HIE) remains a common problem world-wide for infants born at term. The impact of HIE on long-term outcomes, especially into adulthood, is not well-described. To facilitate identification of biobehavioral biomarkers utilizing a translational platform, we sought to investigate the impact of HIE on executive function and cognitive outcomes into adulthood utilizing a murine model of HIE. HIE mice (unilateral common carotid artery occlusion to induce ischemia, followed by hypoxia with a FiO 2 of 0.08 for 45 min) and control mice were tested on discrimination and reversal touchscreen tasks (using their noses) shown to be sensitive to loss of basal ganglia or cortical function, respectively. We hypothesized that the HIE injury would result in deficits in reversal learning, revealing complex cognitive and executive functioning impairments. Following HIE, mice had a mild discrimination impairment as measured by incorrect responses but were able to learn the paradigm to similar levels as controls. During reversal, HIE mice required significantly more total trials, errors and correction trials across the paradigm. Analysis of specific stages showed that reversal impairments in HIE were driven by significant increases in all measured parameters during the late learning, striatal-mediated portion of the task. Together, these results support the concept that HIE occurring during the neonatal period results in abnormal neurodevelopment that persists into adulthood, which can impact efficient associated learning. Further, these data show that utilization of an established model of HIE coupled with touchscreen learning provides valuable information for screening therapeutic interventions that could mitigate these deficits to improve the long-term outcomes of this vulnerable population., (Copyright © 2020 Maxwell, Zimmerman, Pavlik, Newville, Carlin, Robinson, Brigman, Northington and Jantzie.)

Wu and colleagues analyzed the placental pathology from a subset of the neonates in the NEATO trial who had reports available and correlated the placental pathology findings with outcomes. This study highlights the importance of placental pathology, and its potential to bring precision medicine to critically-ill neonates. Placental pathology will likely aid stratification of neonates for clinical trials and accelerate progress for neurorepair.

The ability of small secretory microvesicles known as exosomes to influence neuronal and glial function via their microRNA (miRNA) cargo has positioned them as a novel and effective method of cell-to-cell communication. However, little is known about the role of exosome-secreted miRNAs in the regulation of glutamate receptor gene expression and their relevance for schizophrenia (SCZ) and bipolar disorder (BD). Using mature miRNA profiling and quantitative real-time PCR (qRT-PCR) in the orbitofrontal cortex (OFC) of SCZ (N = 29; 20 male and 9 female), BD (N = 26; 12 male and 14 female), and unaffected control (N = 25; 21 male and 4 female) subjects, we uncovered that miR-223, an exosome-secreted miRNA that targets glutamate receptors, was increased at the mature miRNA level in the OFC of SCZ and BD patients with positive history of psychosis at the time of death and was inversely associated with deficits in the expression of its targets glutamate ionotropic receptor NMDA-type subunit 2B (GRIN2B) and glutamate ionotropic receptor AMPA-type subunit 2 (GRIA2). Furthermore, changes in miR-223 levels in the OFC were positively and negatively correlated with inflammatory and GABAergic gene expression, respectively. Moreover, miR-223 was found to be enriched in astrocytes and secreted via exosomes, and antipsychotics were shown to control its cellular and exosomal localization in a cell-specific manner. Furthermore, addition of astrocytic exosomes in neuronal cultures resulted in a significant increase in miR-223 expression and a notable reduction in Grin2b and Gria2 mRNA levels, which was strongly inversely associated with miR-223 expression. Lastly, inhibition of astrocytic miR-223 abrogated the exosomal-mediated reduction in neuronal Grin2b expression. Taken together, our results demonstrate that the exosomal secretion of a psychosis-altered and glial-enriched miRNA that controls neuronal gene expression is regulated by antipsychotics.

The rates of opioid use disorder during pregnancy have more than quadrupled in the last decade, resulting in numerous infants suffering exposure to opioids during the perinatal period, a critical period of central nervous system (CNS) development. Despite increasing use, the characterization and definition of the molecular and cellular mechanisms of the long-term neurodevelopmental impacts of opioid exposure commencing in utero remains incomplete. Thus, in consideration of the looming public health crisis stemming from the multitude of infants with prenatal opioid exposure entering school age, we undertook an investigation of the effects of perinatal methadone exposure in a novel preclinical model. Specifically, we examined the effects of opioids on the developing brain to elucidate mechanisms of putative neural cell injury, to identify diagnostic biomarkers and to guide clinical studies of outcome and follow-up. We hypothesized that methadone would induce a pronounced inflammatory profile in both dams and their pups, and be associated with immune system dysfunction, sustained CNS injury, and altered cognition and executive function into adulthood. This investigation was conducted using a combination of cellular, molecular, biochemical, and clinically translatable biomarker, imaging and cognitive assessment platforms. Data reveal that perinatal methadone exposure increases inflammatory cytokines in the neonatal peripheral circulation, and reprograms and primes the immune system through sustained peripheral immune hyperreactivity. In the brain, perinatal methadone exposure not only increases chemokines and cytokines throughout a crucial developmental period, but also alters microglia morphology consistent with activation, and upregulates TLR4 and MyD88 mRNA. This increase in neuroinflammation coincides with reduced myelin basic protein and altered neurofilament expression, as well as reduced structural coherence and significantly decreased fractional anisotropy on diffusion tensor imaging. In addition to this microstructural brain injury, adult rats exposed to methadone in the perinatal period have significant impairment in associative learning and executive control as assessed using touchscreen technology. Collectively, these data reveal a distinct systemic and neuroinflammatory signature associated with prenatal methadone exposure, suggestive of an altered CNS microenvironment, dysregulated developmental homeostasis, complex concurrent neural injury, and imaging and cognitive findings consistent with clinical literature. Further investigation is required to define appropriate therapies targeted at the neural injury and improve the long-term outcomes for this exceedingly vulnerable patient population., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Background: Prenatal Alcohol Exposure (PAE) impacts 2% to 5% of infants born in the United States yearly. Women who consume alcohol during pregnancy have a five-fold increased rate of Chorioamnionitis (CHORIO). Both PAE and CHORIO cause microstructural injury to multiple brain regions including major white matter tracts., Objective: Utilizing two previously established animal models, we hypothesized that the combination of PAE+CHORIO would result in greater deficits in myelination and structural integrity than PAE alone., Material and Methods: Pregnant Long-Evans rats voluntarily drank 5% ethanol or saccharin until Gestational Day 19 (GD). On GD19, CHORIO was induced in one group of PAE dams by a 30 min uterine artery occlusion and injection of Lipopolysaccharide (LPS) into each amniotic sac. The remaining PAE dams and saccharin controls underwent sham surgery. Pups were born on GD22 and weaned on Postnatal Day 24 (PD). On PD28, offspring were sacrificed, and their brains examined using ex-vivo Diffusion Tensor Imaging (DTI)., Results: Compared to control, PAE alone did not affect offspring birth weights, mortality or any DTI measures. In contrast, PAE+CHORIO significantly reduced offspring survival and, in surviving pups, increased Radial Diffusivity (RD) in medial frontal cortex and decreased Fractional Anisotropy (FA) in medial and ventral frontal cortex and within capsular regions., Conclusion: The combination of moderate PAE+CHORIO results in an increased mortality, concomitant with diffuse microstructural brain injury noted in young adolescent offspring at PD28. Future studies should examine the extent to which PAE exacerbates the damage caused by CHORIO alone and whether these deficits persist into adulthood.

Background: The need for earlier recognition of children at risk for neurobehavioral problems associated with prenatal ethanol exposure (PAE) has prompted investigations of biomarkers prognostic for altered fetal development. Here, we examined whether PAE alters the expression of angiogenesis-related proteins and cytokines in human placenta in subjects from an Ethanol, Neurodevelopment, Infant and Child Health prospective cohort., Methods: PAE was ascertained by screening questionnaires, Time-line Follow-back interviews and a panel of ethanol biomarkers at two study visits. After delivery, placental tissue samples were collected for protein analysis., Results: No significant differences in the prevalence of substance use, demographic or medical characteristics were observed between the No PAE and PAE groups. PAE was associated with significant reductions in placental expression of VEGFR2 and annexin-A4, while the levels of VEGFR1 and CCM-3 trended downward. A trend toward higher expression of the cytokines TNF-α and IL-13 was also observed in the PAE group. Receiver operating characteristic analyses of the data demonstrated a moderate-to-high degree of diagnostic accuracy for individual placental proteins. Combinations of proteins substantially increased their ability to differentiate between PAE and No PAE subjects., Conclusions: These results establish the feasibility of harvesting placental tissue for protein analyses of PAE in a prospective manner. In addition, given the importance of vascular remodeling in both placenta and developing brain, the role of angiogenic and cytokine proteins in this process warrants further investigation for their utility for predicting alterations in brain development, as well as their mechanistic role in PAE-induced pathology., (© 2019 Wiley Periodicals, Inc.)

Recently, moderate prenatal alcohol exposure (PAE) was shown to be a risk factor for peripheral neuropathy following minor nerve injury. This effect coincides with elevated spinal cord astrocyte activation and ex vivo immune cell reactivity assessed by proinflammatory cytokine interleukin (IL) -1β protein expression. Additionally, the β2-integrin adhesion molecule, lymphocyte function-associated antigen-1 (LFA-1), a factor that influences the expression of the proinflammatory/anti-inflammatory cytokine network is upregulated. Here, we examine whether PAE increases the proinflammatory immune environment at specific anatomical sites critical in the pain pathway of chronic sciatic neuropathy; the damaged sciatic nerve (SCN), the dorsal root ganglia (DRG), and the spinal cord. Additionally, we examine whether inhibiting LFA-1 or IL-1β actions in the spinal cord (intrathecal; i.t., route) could alleviate chronic neuropathic pain and reduce spinal and DRG glial activation markers, proinflammatory cytokines, and elevate anti-inflammatory cytokines. Results show that blocking the actions of spinal LFA-1 using BIRT-377 abolishes allodynia in PAE rats with sciatic neuropathy (CCI) of a 10 or 28-day duration. This effect is observed (utilizing immunohistochemistry; IHC, with microscopy analysis and protein quantification) in parallel with reduced spinal glial activation, IL-1β and TNFα expression. DRG from PAE rats with neuropathy reveal significant increases in satellite glial activation and IL-1β, while IL-10 immunoreactivity is reduced by half in PAE rats under basal and neuropathic conditions. Further, blocking spinal IL-1β with i.t. IL-1RA transiently abolishes allodynia in PAE rats, suggesting that IL-1β is in part, necessary for the susceptibility of adult-onset peripheral neuropathy caused by PAE. Chemokine mRNA analyses from SCN, DRG and spinal cord reveal that increased CCL2 occurs following CCI injury regardless of PAE and BIRT-377 treatment. These data demonstrate that PAE creates dysregulated proinflammatory IL-1β and TNFα /IL-10 responses to minor injury in the sciatic-DRG-spinal pain pathway. PAE creates a risk for developing peripheral neuropathies, and LFA-1 may be a novel therapeutic target for controlling dysregulated neuroimmune actions as a consequence of PAE.

Minimizing central nervous system (CNS) injury from preterm birth depends upon identification of the critical pathways that underlie essential neurodevelopmental and CNS pathophysiology. While chorioamnionitis (CHORIO), is a leading cause of preterm birth, the precise mechanism linking prenatal brain injury and long-term CNS injury is unknown. The chemokine (C-X-C motif) ligand 1 (CXCL1) and its cognate receptor, CXCR2, are implicated in a variety of uterine and neuropathologies, however, their role in CNS injury associated with preterm birth is poorly defined. To evaluate the putative efficacy of CXCR2 blockade in neural repair secondary to CHORIO, we tested the hypothesis that transient postnatal CXCR2 antagonism would reduce neutrophil activation and mitigate cerebral microstructural injury in rats. To this end, a laparotomy was performed on embryonic day 18 (E18) in Sprague Dawley rats, with uterine arteries transiently occluded for 60 min, and lipopolysaccharide (LPS, 4 μg/sac) injected into each amniotic sac. SB225002, a CXCR2 antagonist (3 mg/kg), was administered intraperitoneally from postnatal day 1 (P1)-P5. Brains were collected on P7 and P21 and analyzed with western blot, immunohistochemistry and ex vivo diffusion tensor imaging (DTI). Results demonstrate that transient CXCR2 blockade reduced cerebral neutrophil activation (myeloperoxidase expression/MPO) and mitigated connexin43 expression, indicative of reduced neuroinflammation at P7 ( p < 0.05 for all). CXCR2 blockade also reduced alpha II-spectrin calpain-mediated cleavage, improved pNF/NF ratio, and minimized Iba1 and GFAP expression consistent with improved neuronal and axonal health and reduced gliosis at P21. Importantly, DTI revealed diffuse white matter injury and decreased microstructural integrity following CHORIO as indicated by lower fractional anisotropy (FA) and elevated radial diffusivity (RD) in major white matter tracts ( p < 0.05). Early postnatal CXCR2 blockade also reduced microstructural abnormalities in white matter and hippocampus at P21 ( p < 0.05). Together, these data indicate that transient postnatal blockade of CXCR2 ameliorates perinatal abnormalities in inflammatory signaling, and facilitates neural repair following CHORIO. Further characterization of neuroinflammatory signaling, specifically via CXCL1/CXCR2 through the placental-fetal-brain axis, may clarify stratification of brain injury following preterm birth, and improve use of targeted interventions in this highly vulnerable patient population.

Preterm birth is an important cause of perinatal brain injury (PBI). Neurological injury in extremely preterm infants often begins in utero with chorioamnionitis (CHORIO) or inflammation/infection of the placenta and concomitant placental insufficiency. Studies in humans have shown dysregulated inflammatory signaling throughout the placental-fetal brain axis and altered peripheral immune responses in children born preterm with cerebral palsy (CP). We hypothesized that peripheral immune responses would be altered in our well-established rat model of CP. Specifically, we proposed that isolated peripheral blood mononuclear cells (PBMCs) would be hyperresponsive to a second hit of inflammation throughout an extended postnatal time course. Pregnant Sprague-Dawley dams underwent a laparotomy on embryonic day 18 (E18) with occlusion of the uterine arteries (for 60 min) followed by intra-amniotic injection of lipopolysaccharide (LPS, 4 μg/sac) to induce injury in utero. Shams underwent laparotomy only, with equivalent duration of anesthesia. Laparotomies were then closed, and the rat pups were born at E22. PBMCs were isolated from pups on postnatal day 7 (P7) and P21, and subsequently stimulated in vitro with LPS for 3 or 24 h. A secreted inflammatory profile analysis of conditioned media was performed using multiplex electrochemiluminescent immunoassays, and the composition of inflammatory cells was assayed with flow cytometry (FC). Results indicate that CHORIO PBMCs challenged with LPS are hyperreactive and secrete significantly more tumor necrosis factor α (TNFα) and C-X-C chemokine ligand 1 at P7. FC confirmed increased intracellular TNFα in CHORIO pups at P7 following LPS stimulation, in addition to increased numbers of CD11b/c immunopositive myeloid cells. Notably, TNFα secretion was sustained until P21, with increased interleukin 6, concomitant with increased expression of integrin β1, suggesting both sustained peripheral immune hyperreactivity and a heightened activation state. Taken together, these data indicate that in utero injury primes the immune system and augments enhanced inflammatory signaling. The insidious effects of primed peripheral immune cells may compound PBI secondary to CHORIO and/or placental insufficiency, and thereby render the brain susceptible to future chronic neurological disease. Further understanding of inflammatory mechanisms in PBI may yield clinically important biomarkers and facilitate individualized repair strategies and treatments., (© 2019 S. Karger AG, Basel.)

Toll-like receptor (TLR)3 activation during the neonatal period produces responses linked to the origins of neuropsychiatric disorders. Although there is sexual dimorphism in neuropsychiatric disorders, it is unknown if brain responses to TLR3 activation are sex-specific. We hypothesized that poly I:C in a post-natal day (P)8 model induces a sexually dimorphic inflammatory responses. C57BL6 mice received intraperitoneal injection of poly I:C (10 mg/kg) or vehicle [normal saline (NS)] at P8. Pups were killed at 6 or 14 h for caspase 3 and 8 activity assays, NFkB ELISA, IRF3, AP1, and GFAP western blotting and cytokines/chemokines gene expression real time qRT-PCR (4-6/group). A second group of pups were killed at 24 h (P9) or 7 days (P15) after poly I:C to assess astrocytic (GFAP) and microglia (Iba1) activation in the hippocampus, thalamus and cortex using immunohistochemistry, and gene and protein expression of cytokines/chemokines using real time RT-PCR and MSD, respectively (4-6/group). Non-parametric analysis was applied. Six hours after poly I:C, caspase-3 and -8 activities in cytosolic fractions were 1.6 and 2.8-fold higher in poly I:C-treated than in NS-treated female mice, respectively, while gene expressions of pro-inflammatory cytokines were upregulated in both sexes. After poly I:C, IRF3 nuclear translocation occurred earlier (6 h) in female mice and later (14 h) in male mice. At 14 h after poly I:C, only male mice also had increased nuclear NFκB levels (88%, p < 0.001) and GFAP expression coinciding with persistent IL-6 and FAS gene upregulation (110 and 77%, respectively; p < 0.001) and IL-10 gene downregulation (-42%, p < 0.05). At 24 h after poly I:C, IL-1β, CXCL-10, TNF-α, and MCP-1 were similarly increased in both sexes but at 7 days after exposure, CXCL-10 and INFγ were increased and IL-10 was decreased only in female mice. Accordingly, microglial activation persisted at 7 days after poly I:C in the hippocampus, thalamus and cortex of female mice. This preliminary study suggests that TLR3 activation may produce in the developing neonatal mouse brain a sexually dimorphic response with early activation of caspase-dependent pathways in female mice, activation of inflammatory cascades in both sexes, which then persists in female mice. Further well-powered studies are essential to confirm these sex-specific findings.

Aim: The majority of preclinical studies investigating aberrant glial-neuroimmune actions underlying neuropathic pain have focused on male rodent models. Recently, studies have shown peripheral immune cells play a more prominent role than glial cells in mediating pathological pain in females. Here, we compared the onset and duration of allodynia in males and females, and the anti-allodynic action of a potentially novel therapeutic drug (BIRT377) that not only antagonizes the action of lymphocyte function-associated antigen-1 (LFA-1) to reduce cell migration in the periphery, but may also directly alter the cellular inflammatory bias., Methods: Male and female mice were subjected to peripheral nerve injury chronic constriction injury (CCI) applying two methods, using either 4-0 or 5-0 chromic gut suture material, to examine potential sex differences in the onset, magnitude and duration of allodynia. Hindpaw sensitivity before and after CCI and application of intravenous BIRT377 was assessed. Peripheral and spinal tissues were analyzed for protein (multiplex electrochemiluminescence technology) and mRNA expression (quantitative real-time PCR). The phenotype of peripheral T cells was determined using flow cytometry., Results: Sex differences in proinflammatory CCL2 and IL-1β and the anti-inflammatory IL-10 were observed from a set of cytokines analyzed. A profound proinflammatory T cell (Th17) response in the periphery and spinal cord was also observed in neuropathic females. BIRT377 reversed pain, reduced IL-1β and TNF, and increased IL-10 and transforming growth factor (TGF)-β1, also an anti-inflammatory cytokine, in both sexes. However, female-derived T cell cytokines are transcriptionally regulated by BIRT377, as demonstrated by reducing proinflammatory IL-17A production with concurrent increases in IL-10, TGF-β1 and the anti-inflammatory regulatory T cell-related factor, FOXP3., Conclusion: This study supports that divergent peripheral immune and neuroimmune responses during neuropathy exists between males and females. Moreover, the modulatory actions of BIRT377 on T cells during neuropathy are predominantly specific to females. These data highlight the necessity of including both sexes for studying drug efficacy and mechanisms of action in preclinical studies and clinical trials., Competing Interests: Conflicts of interest All authors declared that there are no conflicts of interest.

Posthemorrhagic hydrocephalus of prematurity (PHHP) remains a global challenge. Early preterm infants (<32 weeks gestation), particularly those exposed to chorioamnionitis (CAM), are prone to intraventricular hemorrhage (IVH) and PHHP. We established an age-appropriate, preclinical model of PHHP with progressive macrocephaly and ventriculomegaly to test whether non-surgical neonatal treatment could modulate PHHP. We combined prenatal CAM and postnatal day 1 (P1, equivalent to 30 weeks human gestation) IVH in rats, and administered systemic erythropoietin (EPO) plus melatonin (MLT), or vehicle, from P2 to P10. CAM-IVH rats developed progressive macrocephaly through P21. Macrocephaly was accompanied by ventriculomegaly at P5 (histology), and P21 ( ex vivo MRI). CAM-IVH rats showed impaired performance of cliff aversion, a neonatal neurodevelopmental test. Neonatal EPO+MLT treatment prevented macrocephaly and cliff aversion impairment, and significantly reduced ventriculomegaly. EPO+MLT treatment prevented matted or missing ependymal motile cilia observed in vehicle-treated CAM-IVH rats. EPO+MLT treatment also normalized ependymal yes-associated protein (YAP) mRNA levels, and reduced ependymal GFAP-immunolabeling. Vehicle-treated CAM-IVH rats exhibited loss of microstructural integrity on diffusion tensor imaging, which was normalized in EPO+MLT-treated CAM-IVH rats. In summary, combined prenatal systemic inflammation plus early postnatal IVH caused progressive macrocephaly, ventriculomegaly and delayed development of cliff aversion reminiscent of PHHP. Neonatal systemic EPO+MLT treatment prevented multiple hallmarks of PHHP, consistent with a clinically viable, non-surgical treatment strategy.

Survivors of infant traumatic brain injury (TBI) are prone to chronic neurological deficits that impose lifelong individual and societal burdens. Translation of novel interventions to clinical trials is hampered in part by the lack of truly representative preclinical tests of cognition and corresponding biomarkers of functional outcomes. To address this gap, the ability of a high-dose, extended, post-injury regimen of erythropoietin (EPO, 3000U/kg/dose × 6d) to prevent chronic cognitive and imaging deficits was tested in a postnatal day 12 (P12) controlled-cortical impact (CCI) model in rats, using touchscreen operant chambers and regional analysis of diffusion tensor imaging (DTI). Results indicate that EPO prevents functional injury and MRI injury after infant TBI. Specifically, subacute DTI at P30 revealed widespread microstructural damage that is prevented by EPO. Assessment of visual discrimination on a touchscreen operant chamber platform demonstrated that all groups can perform visual discrimination. However, CCI rats treated with vehicle failed to pass reversal learning, and perseverated, in contrast to sham and CCI-EPO rats. Chronic DTI at P90 showed EPO treatment prevented contralateral white matter and ipsilateral lateral prefrontal cortex damage. This DTI improvement correlated with cognitive performance. Taken together, extended EPO treatment restores executive function and prevents microstructural brain abnormalities in adult rats with cognitive deficits in a translational preclinical model of infant TBI. Sophisticated testing with touchscreen operant chambers and regional DTI analyses may expedite translation and effective yield of interventions from preclinical studies to clinical trials. Collectively, these data support the use of EPO in clinical trials for human infants with TBI.

Cerebral palsy (CP) is the leading cause of motor impairment for children worldwide and results from perinatal brain injury (PBI). To test novel therapeutics to mitigate deficits from PBI, we developed a rat model of extreme preterm birth (<28 weeks of gestation) that mimics dual intrauterine injury from placental underperfusion and chorioamnionitis. We hypothesized that a sustained postnatal treatment regimen that combines the endogenous neuroreparative agents erythropoietin (EPO) and melatonin (MLT) would mitigate molecular, sensorimotor, and cognitive abnormalities in adults rats following prenatal injury. On embryonic day 18 (E18), a laparotomy was performed in pregnant Sprague-Dawley rats. Uterine artery occlusion was performed for 60 min to induce placental insufficiency via transient systemic hypoxia-ischemia, followed by intra-amniotic injections of lipopolysaccharide, and laparotomy closure. On postnatal day 1 (P1), approximately equivalent to 30 weeks of gestation, injured rats were randomized to an extended EPO + MLT treatment regimen, or vehicle (sterile saline) from P1 to P10. Behavioral assays were performed along an extended developmental time course ( n  = 6-29). Open field testing shows injured rats exhibit hypermobility and disinhibition and that combined neonatal EPO + MLT treatment repairs disinhibition in injured rats, while EPO alone does not. Furthermore, EPO + MLT normalizes hindlimb deficits, including reduced paw area and paw pressure at peak stance, and elevated percent shared stance after prenatal injury. Injured rats had fewer social interactions than shams, and EPO + MLT normalized social drive. Touchscreen operant chamber testing of visual discrimination and reversal shows that EPO + MLT at least partially normalizes theses complex cognitive tasks. Together, these data indicate EPO + MLT can potentially repair multiple sensorimotor, cognitive, and behavioral realms following PBI, using highly translatable and sophisticated developmental testing platforms.