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Purpose: Although up to 25% of germline variants are predicted to affect splicing, most are classified as variants of uncertain significance (VUS) because of the limited understanding of their functional consequences. Here, we investigated the impact of RNA sequencing (RNA-seq) data on the ability to resolve splicing-related VUS., Methods: Patients with VUS predicted to alter splicing identified through commercial hereditary cancer testing between October 2021 to July 2023 were included. RNA-seq was used to compare splicing patterns between patient blood samples and normal controls. VUS reclassification rates were calculated., Results: In total, 411 VUS in 52 genes predicted to affect splicing were included. RNA-seq results resolved 26.3% (108/411) of the VUS: 28 (6.8%) upgraded to pathogenic/likely pathogenic, and 80 (19.5%) downgraded to not reportable. Among the 28 upgraded, 17 (60.7%) were intronic, 9 (32.1%) were exonic missense variants, and 2 (7.1%) were exonic synonymous variants. Most of the VUS downgraded to not reportable were intronic (64/80, 80%). The remaining 16 (20%) of the downgraded VUS were synonymous variants., Conclusion: This study underscores the utility of RNA-seq to detect variants that affect splicing and could have an impact on cancer risk assessment management and treatment., Competing Interests: Youbao Sha, J. Bryce Ortiz, Sara L. Bristow, Kate Loranger, Sheetal Parmar, Adam C. ElNaggar, and Wenbo Xu are employees of Natera, Inc, with stocks or options to own stock in the company., (© 2024 The Authors.)

Background: Accurate individualized assessment of preeclampsia risk enables the identification of patients most likely to benefit from initiation of low-dose aspirin at 12 to 16 weeks of gestation when there is evidence for its effectiveness, and enables the guidance of appropriate pregnancy care pathways and surveillance., Objective: The primary objective of this study was to evaluate the performance of artificial neural network models for the prediction of preterm preeclampsia (<37 weeks' gestation) using patient characteristics available at the first antenatal visit and data from prenatal cell-free DNA screening. Secondary outcomes were prediction of early-onset preeclampsia (<34 weeks' gestation) and term preeclampsia (≥37 weeks' gestation)., Methods: This secondary analysis of a prospective, multicenter, observational prenatal cell-free DNA screening study (SMART) included singleton pregnancies with known pregnancy outcomes. Thirteen patient characteristics that are routinely collected at the first prenatal visit and 2 characteristics of cell-free DNA (total cell-free DNA and fetal fraction) were used to develop predictive models for early-onset (<34 weeks), preterm (<37 weeks), and term (≥37 weeks) preeclampsia. For the models, the "reference" classifier was a shallow logistic regression model. We also explored several feedforward (nonlinear) neural network architectures with ≥1 hidden layers, and compared their performance with the logistic regression model. We selected a simple neural network model built with 1 hidden layer and made up of 15 units., Results: Of the 17,520 participants included in the final analysis, 72 (0.4%) developed early-onset, 251 (1.4%) preterm, and 420 (2.4%) term preeclampsia. Median gestational age at cell-free DNA measurement was 12.6 weeks, and 2155 (12.3%) had their cell-free DNA measurement at ≥16 weeks' gestation. Preeclampsia was associated with higher total cell-free DNA (median, 362.3 vs 339.0 copies/mL cell-free DNA; P<.001) and lower fetal fraction (median, 7.5% vs 9.4%; P<.001). The expected, cross-validated area under the curve scores for early-onset, preterm, and term preeclampsia were 0.782, 0.801, and 0.712, respectively, for the logistic regression model, and 0.797, 0.800, and 0.713, respectively, for the neural network model. At a screen-positive rate of 15%, sensitivity for preterm preeclampsia was 58.4% (95% confidence interval, 0.569-0.599) for the logistic regression model and 59.3% (95% confidence interval, 0.578-0.608) for the neural network model. The contribution of both total cell-free DNA and fetal fraction to the prediction of term and preterm preeclampsia was negligible. For early-onset preeclampsia, removal of the total cell-free DNA and fetal fraction features from the neural network model was associated with a 6.9% decrease in sensitivity at a 15% screen-positive rate, from 54.9% (95% confidence interval, 52.9-56.9) to 48.0% (95% confidence interval, 45.0-51.0)., Conclusion: Routinely available patient characteristics and cell-free DNA markers can be used to predict preeclampsia with performance comparable to that of other patient characteristic models for the prediction of preterm preeclampsia. Logistic regression and neural network models showed similar performance., (Copyright © 2024. Published by Elsevier Inc.)

Background: This study evaluated cardiovascular health, dietary habits, physical activity, depression, and sleep quality in young university adults., Materials and Methods: A cross-sectional design was used to assess anthropometric, biochemical, and cardiovascular health behaviors. The study included 158 university students aged 18 to 30 years (65% women, 35% men, average age: 20.3 ± 2.4 years), selected through non-probabilistic sampling. Measurements included BMI, waist circumference, blood pressure, glucose, triglycerides, HDL and LDL cholesterol, and visceral fat using bioelectrical impedance. Health behaviors were evaluated via questionnaires on physical activity, fruit and vegetable consumption, smoking, ultra-processed food consumption, and sleep quality using the Pittsburgh Sleep Quality Index. The cardiovascular health index was assessed with the "Life's Essential 8" questionnaire and depression was assessed with Beck Depression Inventory. Statistical analyses included ANOVA, Fisher's F test, Student's t-test, and simple linear regression, conducted using SPSS Statistics version 25.0 (IBM Corp. Released 2017. IBM SPSS Statistics for Windows, Version 25.0. Armonk, NY: IBM Corp), with significance set at p<0.05., Results: Women showed better adherence to healthy behaviors. Higher fruit and vegetable consumption and physical activity were associated with lower visceral fat. Higher visceral fat is correlated with increased blood pressure and decreased HDL cholesterol. Smoking and frequent ultra-processed food consumption were linked to higher depression scores, which were associated with poorer sleep quality., Conclusion: Healthy lifestyle habits are crucial for physical and mental health, providing a basis for public health interventions., Competing Interests: Human subjects: Consent was obtained or waived by all participants in this study. Research Division of the School of Medicine at the Universidad Nacional Autónoma de México issued approval FM/DI/022/2021. Animal subjects: All authors have confirmed that this study did not involve animal subjects or tissue. Conflicts of interest: In compliance with the ICMJE uniform disclosure form, all authors declare the following: Payment/services info: All authors have declared that no financial support was received from any organization for the submitted work. Financial relationships: All authors have declared that they have no financial relationships at present or within the previous three years with any organizations that might have an interest in the submitted work. Other relationships: All authors have declared that there are no other relationships or activities that could appear to have influenced the submitted work., (Copyright © 2024, Maltos-Gómez et al.)

Traumatic brain injury (TBI) disrupts the blood-brain barrier (BBB), which may exacerbate neuroinflammation post-injury. Few translational studies have examined BBB dysfunction and subsequent neuroinflammation post-TBI in juveniles. We hypothesized that BBB dysfunction positively predicts microglial activation and that vulnerability to BBB dysfunction and associated neuroinflammation are dependent on age at injury. Post-natal day (PND)17 and PND35 rats ( n  = 56) received midline fluid percussion injury or sham surgery, and immunoglobulin-G (IgG) stain was quantified as a marker of extravasated blood in the brain and BBB dysfunction. We investigated BBB dysfunction and the microglial response in the hippocampus, hypothalamus, and motor cortex relative to age at injury and days post-injury (DPI; 1, 7, and 25). We measured the morphologies of ionized calcium-binding adaptor molecule 1-labeled microglia using cell body area and perimeter, microglial branch number and length, end-points/microglial cell, and number of microglia. Data were analyzed using generalized hierarchical models. In PND17 rats, TBI increased levels of IgG compared to shams. Independent of age at injury, IgG in TBI rats was higher at 1 and 7 DPI, but resolved by 25 DPI. TBI activated microglia (more cells and fewer end-points) in PND35 rats compared to respective shams. Independent of age at injury, TBI induced morphological changes indicative of microglial activation, which resolved by 25 DPI. TBI rats had fewer cells and end-points per cell at 1 and 7 DPI than 25 DPI. Independent of TBI, PND17 rats had larger, more activated microglia than PND35 rats; PND17 TBI rats had larger cell body areas and perimeters than PND35 TBI rats. Importantly, we found support in both ages that IgG quantification predicted microglial activation after TBI. The number of microglia increased with increasing IgG, whereas branch length decreased with increasing IgG, which together indicate microglial activation. Our results suggest that stabilization of the BBB after pediatric TBI may be an important therapeutic strategy to limit neuroinflammation and promote recovery., Competing Interests: No competing financial interests exist., (© Tabitha R.F. Green et al., 2024; Published by Mary Ann Liebert, Inc.)

Traumatic brain injury (TBI) can damage the hypothalamus and cause improper activation of the growth hormone (GH) axis, leading to growth hormone deficiency (GHD). GHD is one of the most prevalent endocrinopathies following TBI in adults; however, the extent to which GHD affects juveniles remains understudied. We used postnatal day 17 rats (n = 83), which model the late infantile/toddler period, and assessed body weights, GH levels, and number of hypothalamic somatostatin neurons at acute (1, 7 days post injury (DPI)) and chronic (18, 25, 43 DPI) time points. We hypothesized that diffuse TBI would alter circulating GH levels because of damage to the hypothalamus, specifically somatostatin neurons. Data were analyzed with generalized linear and mixed effects models with fixed effects interactions between the injury and time. Despite similar growth rates over time with age, TBI rats weighed less than shams at 18 DPI (postnatal day 35; P = 0.03, standardized effect size [d] = 1.24), which is around the onset of puberty. Compared to shams, GH levels were lower in the TBI group during the acute period (P = 0.196; d = 12.3) but higher in the TBI group during the chronic period (P = 0.10; d = 52.1). Although not statistically significant, TBI-induced differences in GH had large standardized effect sizes, indicating biological significance. The mean number of hypothalamic somatostatin neurons (an inhibitor of GH) positively predicted GH levels in the hypothalamus but did not predict GH levels in the somatosensory cortex. Understanding TBI-induced alterations in the GH axis may identify therapeutic targets to improve the quality of life of pediatric survivors of TBI.

Objective: To identify conditions on a reproductive carrier screening panel with the potential for carrier manifestations during pregnancy and review the implications for obstetric care., Methods: This was a retrospective cross-sectional study of consecutive samples from female patients aged 18-55 years submitted to a commercial laboratory for a 274-gene carrier screening panel (January 2020 to September 2022). A literature review was performed to identify genes on the panel with potential for pregnancy complications in carriers. Carrier expression and published recommendations for clinical management were reviewed., Results: We identified 12 genes with potential for carrier manifestations during pregnancy based on reports in the literature: nine with manifestations irrespective of the fetal genetic status ( ABCB11 , COL4A3 , COL4A4 , COL4A5 , DMD , F9 , F11 , GLA , and OTC ) and three ( CPT1A , CYP19A1 , and HADHA ) with manifestations only if the fetus is affected by the condition. Manifestations included cardiomyopathy, hemorrhage, gestational hypertensive disorders, cholestasis of pregnancy, acute fatty liver, hyperammonemic crisis, and maternal virilization. Published recommendations for carrier management were identified for 11 of the 12 genes. Of 91,637 tests performed during the study period, a pathogenic or likely pathogenic variant was identified in 2,139 (2.3%), giving a carrier frequency for any of the 12 genes of 1 in 43 (95% CI 1/41-45) 1,826 (2.0%) of the study population were identified as carriers for one of the nine genes with the potential for carrier manifestations irrespective of an affected or unaffected fetus., Conclusion: Approximately 1 in 40 female patients were identified as carriers for a condition with potential for maternal manifestations in pregnancy, including some serious or even life-threatening complications. Obstetric care professionals should be aware of the possibility of pregnancy complications among carriers and the available recommendations for management., Funding Source: This study was funded by Natera, Inc., Competing Interests: Financial Disclosure Vivienne Souter, Brittany Prigmore, Emily Becraft, Elizabeth Repass, Trevor Smart, Nina Sanapareddy, Melissa Schweitzer, J. Bryce Ortiz, and Yang Wang are employees of Natera, Inc. with stocks or options to own stocks in the company. Peter Benn is a consultant for Natera, Inc. with options to own stocks in the company. The other authors did not report any potential conflicts of interest., (Copyright © 2023 The Author(s). Published by Wolters Kluwer Health, Inc.)

Owing to a long history of anthropogenic pressures, freshwater ecosystems are among the most vulnerable to biodiversity loss 1 . Mitigation measures, including wastewater treatment and hydromorphological restoration, have aimed to improve environmental quality and foster the recovery of freshwater biodiversity 2 . Here, using 1,816 time series of freshwater invertebrate communities collected across 22 European countries between 1968 and 2020, we quantified temporal trends in taxonomic and functional diversity and their responses to environmental pressures and gradients. We observed overall increases in taxon richness (0.73% per year), functional richness (2.4% per year) and abundance (1.17% per year). However, these increases primarily occurred before the 2010s, and have since plateaued. Freshwater communities downstream of dams, urban areas and cropland were less likely to experience recovery. Communities at sites with faster rates of warming had fewer gains in taxon richness, functional richness and abundance. Although biodiversity gains in the 1990s and 2000s probably reflect the effectiveness of water-quality improvements and restoration projects, the decelerating trajectory in the 2010s suggests that the current measures offer diminishing returns. Given new and persistent pressures on freshwater ecosystems, including emerging pollutants, climate change and the spread of invasive species, we call for additional mitigation to revive the recovery of freshwater biodiversity., (© 2023. The Author(s).)

To investigate microglial mechanisms in central and peripheral inflammation after experimental traumatic brain injury (TBI), we inhibited the colony-stimulating factor-1 receptor (CSF-1R) with PLX5622 (PLX). We hypothesized that microglia depletion would attenuate central inflammation acutely with no effect on peripheral inflammation. After randomization, male mice ( n  = 105) were fed PLX or control diets (21 days) and then received midline fluid percussion injury or sham injury. Brain and blood were collected at 1, 3, or 7 days post-injury (DPI). Immune cell populations were quantified in the brain and blood by flow cytometry. Cytokines (interleukin [IL]-6, IL-1β, tumor necrosis factor-α, interferon-γ, IL-17A, and IL-10) were quantified in the blood using a multi-plex enzyme-linked immunosorbent assay. Data were analyzed using Bayesian multi-variate, multi-level models. PLX depleted microglia at all time points and reduced neutrophils in the brain at 7 DPI. PLX also depleted CD115 + monocytes, reduced myeloid cells, neutrophils, and Ly6C low monocytes in blood, and elevated IL-6. TBI induced a central and peripheral immune response. TBI elevated leukocytes, microglia, and macrophages in the brain and elevated peripheral myeloid cells, neutrophils, Ly6C int monocytes, and IL-1β in the blood. TBI lowered peripheral CD115 + and Ly6C low monocytes in the blood. TBI PLX mice had fewer leukocytes and microglia in the brain at 1 DPI, with elevated neutrophils at 7 DPI compared to TBI mice on a control diet. TBI PLX mice also had fewer peripheral myeloid cells, CD115 + , and Ly6C low monocytes in the blood at 3 DPI, but elevated Ly6C high , Ly6C int , and CD115 + monocyte populations at 7 DPI, compared to TBI mice on a control diet. TBI PLX mice had elevated proinflammatory cytokines and lower anti-inflammatory cytokines in the blood at 7 DPI compared to TBI mice on a control diet. CSF-1R inhibition reduced the immune response to TBI at 1 and 3 DPI, but elevated peripheral inflammation at 7 DPI., Competing Interests: No competing financial interests exist., (© Katherine R. Giordano et al., 2023; Published by Mary Ann Liebert, Inc.)

Microglia play a critical role in the neuroimmune response, but little is known about the role of microglia in sleep following an inflammatory trigger. Nevertheless, decades of research have been predicated on the assumption that an inflammatory trigger increases sleep through microglial activation. We hypothesized that mice ( n = 30) with depleted microglia using PLX5622 (PLX) would sleep less following the administration of lipopolysaccharide (LPS) to induce inflammation. Brains were collected and microglial morphology was assessed using quantitative skeletal analyses and physiological parameters were recorded using non-invasive piezoelectric cages. Mice fed PLX diet had a transient increase in sleep that dissipated by week 2. Subsequently, following a first LPS injection (0.4 mg/kg), mice with depleted microglia slept more than mice on the control diet. All mice were returned to normal rodent chow to repopulate microglia in the PLX group (10 days). Nominal differences in sleep existed during the microglia repopulation period. However, following a second LPS injection, mice with repopulated microglia slept similarly to control mice during the dark period but with longer bouts during the light period. Comparing sleep after the first LPS injection to sleep after the second LPS injection, controls exhibited temporal changes in sleep patterns but no change in cumulative minutes slept, whereas cumulative sleep in mice with repopulated microglia decreased during the dark period across all days. Repopulated microglia had a reactive morphology. We conclude that microglia are necessary to regulate sleep after an immune challenge.

Nuclear medicine probes turned into the key for the identification and precise location of sentinel lymph nodes and other occult lesions (i.e., tumors) by using the systemic administration of radiotracers. Intraoperative nuclear probes are key in the surgical management of some malignancies as well as in the determination of positive surgical margins, thus reducing the extent and potential surgery morbidity. Depending on their application, nuclear probes are classified into two main categories, namely, counting and imaging. Although counting probes present a simple design, are handheld (to be moved rapidly), and provide only acoustic signals when detecting radiation, imaging probes, also known as cameras, are more hardware-complex and also able to provide images but at the cost of an increased intervention time as displacing the camera has to be done slowly. This review article begins with an introductory section to highlight the relevance of nuclear-based probes and their components as well as the main differences between ionization- (semiconductor) and scintillation-based probes. Then, the most significant performance parameters of the probe are reviewed (i.e., sensitivity, contrast, count rate capabilities, shielding, energy, and spatial resolution), as well as the different types of probes based on the target radiation nature, namely: gamma (γ), beta (β) (positron and electron), and Cherenkov. Various available intraoperative nuclear probes are finally compared in terms of performance to discuss the state-of-the-art of nuclear medicine probes. The manuscript concludes by discussing the ideal probe design and the aspects to be considered when selecting nuclear-medicine probes., (© 2022 The Authors. Medical Physics published by Wiley Periodicals LLC on behalf of American Association of Physicists in Medicine.)

Over half of fatal pediatric traumatic brain injuries are estimated to be the result of physical abuse, i.e., abusive head trauma (AHT). Although intimate partner violence (IPV) is a well-established risk for child maltreatment, little is known about IPV as an associated risk factor specifically for AHT. We performed a single-institution, retrospective review of all patients (0-17 years) diagnosed at a Level 1 pediatric trauma center with head trauma who had been referred to an in-hospital child protection team for suspicion of AHT between 2010 and 2016. Data on patient demographics, hospitalization, injury, family characteristics, sociobehavioral characteristics, physical examination, laboratory findings, imaging, discharge, and forensic determination of AHT were extracted from the institution's forensic registry. Descriptive statistics (mean, median), chi-square and Mann-Whitney U tests were used to compare patients with fatal head injuries to patients with nonfatal head injuries by clinical characteristics, family characteristics, and forensic determination. Multiple logistic regression was used to estimate adjusted odds ratios for the presence of IPV as an associated risk of AHT while controlling for other clinical and family factors. Of 804 patients with suspicion for AHT in the forensic registry, there were 240 patients with a forensic determination of AHT; 42 injuries were fatal. There were 101 families with a reported history of IPV; 64.4% of patients in families with reported IPV were <12 months of age. IPV was associated with a twofold increase in the risk of AHT (Exp( β ) = 2.3 [ p = .02]). This study confirmed IPV was an associated risk factor for AHT in a single institution cohort of pediatric patients with both fatal and nonfatal injuries. Identifying IPV along with other family factors may improve detection and surveillance of AHT in medical settings and help reduce injury, disability, and death.

The objective of this study was to determine the prevalence of sleep-wake disturbances (SWD) following pediatric traumatic brain injury (TBI), and to examine characteristics of TBI and patient demographics that might be predictive of subsequent SWD development. This single-institution retrospective study included patients diagnosed with a TBI during 2008-2019 who also had a subsequent diagnosis of an SWD. Data were collected using ICD-9/10 codes for 207 patients and included the following: age at initial TBI, gender, TBI severity, number of TBIs diagnosed prior to SWD diagnosis, type of SWD, and time from initial TBI to SWD diagnosis. Multinomial logit and negative-binomial models were fit to investigate whether the multiple types of SWD and the time to onset of SWD following TBI could be predicted by patient variables. Distributions of SWD diagnosed after TBI were similar between genders. The probability of insomnia increased with increasing patient age. The probability of 'difficulty sleeping' was highest in 7-9 year-old TBI patients. Older TBI patients had shorter time to SWD onset than younger patients. Patients with severe TBI had the shortest time to SWD onset, whereas patients with mild or moderate TBI had comparable times to SWD onset. Multiple TBI characteristics and patient demographics were predictive of a subsequent SWD diagnosis in the pediatric population. This is an important step toward increasing education among providers, parents, and patients about the risk of developing SWD following TBI.

Few translational studies have examined how age-at-injury affects the glial response to traumatic brain injury (TBI). We hypothesized that rats injured at post-natal day (PND) 17 would exhibit a greater glial response, that would persist into early adulthood, compared to rats injured at PND35. PND17 and PND35 rats ( n = 75) received a mild to moderate midline fluid percussion injury or sham surgery. In three cortical regions [peri-injury, primary somatosensory barrel field (S1BF), perirhinal], we investigated the glial response relative to age-at-injury (PND17 or PND35), time post-injury (2 hours, 1 day, 7 days, 25 days, or 43 days), and post-natal age, such that rats injured at PND17 or PND35 were compared at the same post-natal-age (e.g., PND17 + 25D post-injury = PND42; PND35 + 7D post-injury = PND42). We measured Iba1 positive microglia cells (area, perimeter) and quantified their activation status using skeletal analysis (branch length/cell, mean processes/cell, cell abundance). GFAP expression was examined using immunohistochemistry and pixel analysis. Data were analyzed using Bayesian multivariate multi-level models. Independent of age-at-injury, TBI activated microglia (shorter branches, fewer processes) in the S1BF and perirhinal cortex with more microglia in all regions compared to uninjured shams. TBI-induced microglial activation (shorter branches) was sustained in the S1BF into early adulthood (PND60). Overall, PND17 injured rats had more microglial activation in the perirhinal cortex than PND35 injured rats. Activation was not confounded by age-dependent cell size changes, and microglial cell body sizes were similar between PND17 and PND35 rats. There were no differences in astrocyte GFAP expression. Increased microglial activation in PND17 brain-injured rats suggests that TBI upregulates the glial response at discrete stages of development. Age-at-injury and aging with an injury are translationally important because experiencing a TBI at an early age may trigger an exaggerated glial response., 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 © 2022 Green, Murphy, Ortiz and Rowe.)

There is an unmet clinical need for curative therapies to treat neurodegenerative disorders. Most mainstay treatments currently on the market only alleviate specific symptoms and do not reverse disease progression. The Pituitary adenylate cyclase-activating polypeptide (PACAP), an endogenous neuropeptide hormone, has been extensively studied as a potential regenerative therapeutic. PACAP is widely distributed in the central nervous system (CNS) and exerts its neuroprotective and neurotrophic effects via the related Class B GPCRs PAC1, VPAC1, and VPAC2, at which the hormone shows roughly equal activity. Vasoactive intestinal peptide (VIP) also activates these receptors, and this close analogue of PACAP has also shown to promote neuronal survival in various animal models of acute and progressive neurodegenerative diseases. However, PACAP's poor pharmacokinetic profile (non-linear PK/PD), and more importantly its limited blood-brain barrier (BBB) permeability has hampered development of this peptide as a therapeutic. We have demonstrated that glycosylation of PACAP and related peptides promotes penetration of the BBB and improves PK properties while retaining efficacy and potency in the low nanomolar range at its target receptors. Furthermore, judicious structure-activity relationship (SAR) studies revealed key motifs that can be modulated to afford compounds with diverse selectivity profiles. Most importantly, we have demonstrated that select PACAP glycopeptide analogues ( 2LS80Mel and 2LS98Lac ) exert potent neuroprotective effects and anti-inflammatory activity in animal models of traumatic brain injury and in a mild-toxin lesion model of Parkinson's disease, highlighting glycosylation as a viable strategy for converting endogenous peptides into robust and efficacious drug candidates., Competing Interests: Conflict of Interest: CA, LS, JS, MH, TF, and RP hold patents related to the content. JS, MH, TF, and RP have equity in Teleport Pharmaceuticals, LLC, a UArizona biotech startup. This interest played no role in the design of the studies; in the collection, analyses, or interpretation of data; in the writing of the Poster, or in the decision to present the results. The remaining 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.

Intimate partner violence (IPV) increases risk of traumatic brain injury (TBI). Physical assaults increase in frequency and intensity during pregnancy. The consequences of TBI during pregnancy (gravida TBI; gTBI) on offspring development is unknown, for which stress and inflammation during pregnancy worsen fetal developmental outcomes. We hypothesized that gTBI would lead to increased anxiety- and depression-related behavior, altered inflammatory responses and gut pathology, and distorted brain circuitry in mixed-sex offspring compared to mice born to control mothers. Pregnant dams received either diffuse TBI or sham injury (control) 12 days post-coitum. We found that male gTBI offspring were principal drivers of the gTBI effects on health, physiology, and behavior. For example, male, but not female, gTBI offspring weighed significantly less at weaning compared to male control offspring. At post-natal day (PND) 28, gTBI offspring had significantly weaker intralaminar connectivity onto layer 5 pre-frontal pyramidal neurons compared to control offspring. Neurological performance on anxiety-like behaviors was decreased, with only marginal differences in depressive-like behaviors, for gTBI offspring compared to control offspring. At PND42 and PND58, circulating neutrophil and monocyte populations were significantly smaller in gTBI male offspring than control male offspring. In response to a subsequent inflammatory challenge at PND75, gTBI offspring had significantly smaller circulating neutrophil populations than control offspring. Anxiety-like behaviors persisted during the immune challenge in gTBI offspring. However, spleen immune response and gut histology showed no significant differences between groups. The results compel further studies to determine the full extent of gTBI on fetal and maternal outcomes.

Past studies find that chronic stress alters inhibitory, GABAergic circuitry of neurons in distinct hippocampal subregions. Less clear is whether these effects persist weeks after chronic stress ends, and whether these effects involve changes in the total number of hippocampal GABAergic neurons or modulates the function of specific GABAergic subtypes. A transgenic mouse line (VGAT:Cre Ai9) containing an indelible marker for GABAergic neurons (tdTomato) throughout the brain was used to determine whether chronic stress alters total GABAergic neuronal number or the expression of two key GABAergic cell subtypes, calretinin expressing (CR+) and somatostatin expressing (SOM+) neurons, and whether these changes endure weeks later. Male and female mice were chronically stressed in wire mesh restrainers for 6h/d/21d (Str) or not (Con), and then allowed a 3 week rest period (Str-Rest) and compared to those without a rest period (Str-NoRest). Epifluorescent microscope images of immunohistochemistry-processed brains were quantified to estimate the total number of fluorescently-labeled hippocampal GABAergic neurons and the proportion that were CR+ or SOM+. Neither chronic stress nor sex altered the total number of GABAergic cells. In contrast, chronic stress reduced the expression of CR+ in the CA3 region of the hippocampus in both males and females, with robust reductions in the DG region of males, but not females, and these changes reversed following a rest period. Chronic stress also reduced the proportion of hippocampal SOM+ neurons and this reduction persisted even with a rest period. These results show chronic stress dynamically reduced CR expression without changing total inhibitory neuronal number and point to CR as a potential new lead to understand mechanisms by which chronic stress alters hippocampal function., (© 2020 Wiley Periodicals LLC.)

Background: Abusive head trauma (AHT) is the leading cause traumatic death in children ≤5 years of age. AHT remains seriously under-surveilled, increasing the risk of subsequent injury and death. This study assesses the clinical and social risks associated with fatal and non-fatal AHT., Methods: A single-institution, retrospective review of suspected AHT patients ≤5 years of age between 2010 and 2016 using a prospective hospital forensic registry data yielded demographic, clinical, family, psycho-social and other follow-up information. Descriptive statistics were used to look for differences between patients with AHT and accidental head trauma. Logistic regression estimated the adjusted odds ratios (AOR) for AHT. A receiver operating characteristic (ROC) curve was created to calculate model sensitivity and specificity., Results: Forensic evaluations of 783 children age ≤5 years with head trauma met the inclusion criteria; 25 were fatal with median[IQR] age 23[4.5-39.0] months. Of 758 non-fatal patients, age was 7[3.0-11.0] months; 59.5% male; 435 patients (57.4%) presented with a skull fracture, 403 (53.2%) with intracranial hemorrhage. Ultimately 242 (31.9%) were adjudicated AHT, 335(44.2%) were accidental, 181 (23.9%) were undetermined. Clinical factors increasing the risk of AHT included multiple fractures (Exp(β) = 9.9[p = 0.001]), bruising (Expβ = 5.7[p < 0.001]), subdural blood (Exp(β) = 5.3[p = 0.001]), seizures (Exp(β) = 4.9[p = 0.02]), lethargy/unresponsiveness (Exp(β) = 2.24[p = 0.02]), loss of consciousness (Exp(β) = 4.69[p = 0.001]), and unknown mechanism of injury (Exp(β) = 3.9[p = 0.001]); skull fracture reduced the risk of AHT by half (Exp(β) = 0.5[p = 0.011]). Social risks factors included prior police involvement (Exp(β) = 5.9[p = 0.001]), substance abuse (Exp(β) = 5.7[p = .001]), unknown number of adults in the home (Exp(β) = 4.1[p = 0.001]) and intimate partner violence (Exp(β) = 2.3[p = 0.02]). ROC area under the curve (AUC) = 0.90([95% CI = 0.86-0.93] p = .001) provides 73% sensitivity; 91% specificity., Conclusions: To improve surveillance of AHT, interviews should include and consider social factors including caregiver/household substance abuse, intimate partner violence, prior police involvement and household size. An unknown number of adults in home is associated with an increased risk of AHT., Study Type/level of Evidence: Prognostic, Level III., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Traumatic brain injury (TBI) survivors suffer from a range of morbidities, including post-traumatic endocrinopathies that can cause physical and mental changes in patients, greatly compromising quality of life. This study tested the hypothesis that mild and moderate diffuse TBI leads to chronic deficiencies in corticosterone (CORT) regulation following repeated exposure to restraint stress over time. Young adult male rats ( n  = 9-11/group) were subjected to mild or moderate TBI induced by midline fluid percussion injury (mFPI) or control sham surgery. At 6 and 24 h post-injury, both mild and moderate TBI resulted in elevated resting plasma CORT levels compared with uninjured shams. Independent of TBI severity, all rats had lower resting plasma CORT levels at 7, 14, 28, and 54 days post-injury compared with pre-surgery baseline CORT. Circulating levels of CORT were also evaluated under restraint stress and in response to dexamethasone (DEX), a synthetic glucocorticoid. Independent of TBI severity, restraint stress elevated CORT at 30, 60, and 90 min post-stressor initiation at all post-injury time-points. A blunted CORT response to restraint stress was observed with lower CORT levels after restraint at 28 and 54 days compared with 7 days post-injury (DPI), indicative of habituation to the stressor. A high dose of DEX lowered CORT levels at 90 min post-restraint stress initiation compared with low-dose DEX, independent of TBI severity. These results support TBI-induced CORT dysregulation at acute time-points, but additional studies that investigate the onset and progression of endocrinopathies, controlling for habituation to repeated restraint stress, are needed to inform the diagnosis and treatment of such morbidities in TBI survivors., Competing Interests: No conflicting financial interests exist., (© Rachel K. Rowe et al., 2020; Published by Mary Ann Liebert, Inc.)

Traumatic brain injury (TBI) and Alzheimer's disease (AD) are diseases during which the fine-tuned autoregulation of the brain is lost. Despite the stark contrast in their causal mechanisms, both TBI and AD are conditions which elicit a neuroinflammatory response that is coupled with physical, cognitive, and affective symptoms. One commonly reported symptom in both TBI and AD patients is disturbed sleep. Sleep is regulated by circadian and homeostatic processes such that pathological inflammation may disrupt the chemical signaling required to maintain a healthy sleep profile. In this way, immune system activation can influence sleep physiology. Conversely, sleep disturbances can exacerbate symptoms or increase the risk of inflammatory/neurodegenerative diseases. Both TBI and AD are worsened by a chronic pro-inflammatory microenvironment which exacerbates symptoms and worsens clinical outcome. Herein, a positive feedback loop of chronic inflammation and sleep disturbances is initiated. In this review, the bidirectional relationship between sleep disturbances and inflammation is discussed, where chronic inflammation associated with TBI and AD can lead to sleep disturbances and exacerbated neuropathology. The role of microglia and cytokines in sleep disturbances associated with these diseases is highlighted. The proposed sleep and inflammation-mediated link between TBI and AD presents an opportunity for a multifaceted approach to clinical intervention., (Copyright © 2020 Green, Ortiz, Wonnacott, Williams and Rowe.)

Identifying differential responses between sexes following traumatic brain injury (TBI) can elucidate the mechanisms behind disease pathology. Peripheral and central inflammation in the pathophysiology of TBI can increase sleep in male rodents, but this remains untested in females. We hypothesized that diffuse TBI would increase inflammation and sleep in males more so than in females. Diffuse TBI was induced in C57BL/6J mice and serial blood samples were collected (baseline, 1, 5, 7 days post-injury [DPI]) to quantify peripheral immune cell populations and sleep regulatory cytokines. Brains and spleens were harvested at 7DPI to quantify central and peripheral immune cells, respectively. Mixed-effects regression models were used for data analysis. Female TBI mice had 77%-124% higher IL-6 levels than male TBI mice at 1 and 5DPI, whereas IL-1β and TNF-α levels were similar between sexes at all timepoints. Despite baseline sex differences in blood-measured Ly6C high monocytes (females had 40% more than males), TBI reduced monocytes by 67% in TBI mice at 1DPI. Male TBI mice had 31%-33% more blood-measured and 31% more spleen-measured Ly6G + neutrophils than female TBI mice at 1 and 5DPI, and 7DPI, respectively. Compared with sham, TBI increased sleep in both sexes during the first light and dark cycles. Male TBI mice slept 11%-17% more than female TBI mice, depending on the cycle. Thus, sex and TBI interactions may alter the peripheral inflammation profile and sleep patterns, which might explain discrepancies in disease progression based on sex., (© 2019 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

The influence of estrogens on modifying cognition has been extensively studied, revealing that a wide array of factors can significantly impact cognition, including, but not limited to, subject age, estrogen exposure duration, administration mode, estrogen formulation, stress history, and progestogen presence. Less known is whether long-term, extended exposure to estrogens would benefit or otherwise impact cognition. The present study examined the effects of 17β-estradiol (E2) exposure for seven months, beginning in late adulthood and continuing into middle age, using a regimen of cyclic exposure (bi-monthly subcutaneous injection of 10 μg E2), or Cyclic+Tonic exposure (bi-monthly subcutaneous injection of 10 μg E2 + Silastic capsules of E2) in ovariectomized female Fischer-344-CDF rats. Subjects were tested on a battery of learning and memory tasks. All groups learned the water radial-arm maze (WRAM) and Morris water maze tasks in a similar fashion, regardless of hormone treatment regimen. In the asymptotic phase of the WRAM, rats administered a Cyclic+Tonic E2 regimen showed enhanced performance when working memory was taxed compared to Vehicle and Cyclic E2 groups. Assessment of spatial memory on object placement and object recognition was not possible due to insufficient exploration of objects; however, the Cyclic+Tonic group showed increased total time spent exploring all objects compared to Vehicle-treated animals. Overall, these data demonstrate that long-term Cyclic+Tonic E2 exposure can result in some long-term cognitive benefits, at least in the spatial working memory domain, in a surgically menopausal rat model., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Traumatic brain injury (TBI) in children can result in long-lasting social, cognitive, and neurological impairments. In adults, TBI can lead to endocrinopathies (endocrine system disorders), but this is infrequently reported in children. Untreated endocrinopathies can elevate risks of subsequent health issues, such that early detection in pediatric TBI survivors can initiate clinical interventions. To understand the risk of endocrinopathies following pediatric TBI, we identified patients who had experienced a TBI and subsequently developed a new-onset hypothalamic regulated endocrinopathy ( n = 498). We hypothesized that pediatric patients who were diagnosed with a TBI were at higher risk of being diagnosed with a central endocrinopathy than those without a prior diagnosis of TBI. In our epidemiological assessment, we identified pediatric patients enrolled in the Arizona Health Care Cost Containment System (AHCCCS) from 2008 to 2014 who were diagnosed with one of 330 TBI International Classification of Diseases (ICD)-9 codes and subsequently diagnosed with one of 14 central endocrinopathy ICD-9 codes. Additionally, the ICD-9 code data from over 600,000 Arizona pediatric patients afforded an estimate of the incidence, prevalence, relative risk, odds ratio, and number needed to harm, regarding the development of a central endocrinopathy after sustaining a TBI in Arizona Medicaid pediatric patients. Children with a TBI diagnosis had 3.22 times the risk of a subsequent central endocrine diagnosis compared with the general population (±0.28). Pediatric AHCCCS patients with a central endocrine diagnosis had 3.2-fold higher odds of a history of a TBI diagnosis than those without an endocrine diagnosis (±0.29). Furthermore, the number of patients with a TBI diagnosis for one patient to receive a diagnosis of a central endocrine diagnosis was 151.2 (±6.12). Female subjects were more likely to present with a central endocrine diagnosis after a TBI diagnosis compared to male subjects (64.1 vs. 35.9%). These results are the first state-wide epidemiological study conducted to determine the risk of developing a hypothalamic-pituitary disorder after a TBI in the pediatric population. Our results contribute to a body of knowledge demonstrating a TBI etiology for idiopathic endocrine disorders, and thus advise physicians with regard to TBI follow-up care that includes preventive screening for endocrine disorders., (Copyright © 2020 Ortiz, Sukhina, Balkan, Harootunian, Adelson, Lewis, Oatman, Subbian, Rowe and Lifshitz.)

This study investigated the role of the dorsal hippocampus (dHPC) in the temporal entrainment of behavior, while addressing limitations of previous evidence from peak procedure experiments. Rats were first trained on a switch-timing task in which food was obtained from one of two concurrently available levers; one lever was effective after 8 s and the other after 16  s. After performance stabilized, rats underwent either bilateral NMDA lesions of the dHPC or sham lesions. After recovery, switch-timing training resumed. In a subsequent condition, the switch-timing task was modified such that food was available after either 8 or 32 s. Although dHPC lesions had subtle and complex effects on when rats stopped seeking for food at the 8-s lever (departures), it more systematically reduced the time when rats started seeking for food at the 16-s and 32-s lever (switches). No systematic effect of dHPC lesions were observed on the coefficient of quartile variation (normalized dispersion) of latencies to switch. Within the context of the pacemaker-accumulator framework of interval timing, these findings suggest that partially or wholly independent mechanisms control the initiation and termination of timed responses, and that the dHPC is primarily involved in encoding the time to start responding., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Histology and immunohistochemistry are routine methods of analysis to visualize microscopic anatomy and localize proteins within biological tissue. In neuroscience, as well as a plethora of other scientific fields, these techniques are used. Immunohistochemistry can be done on slide mounted tissue or free-floating sections. Preparing slide-mounted samples is a time intensive process. The following protocol for a technique, called the Megabrain, reduced the time taken to cryosection and mount brain tissue by up to 90% by combining multiple brains into a single frozen block. Furthermore, this technique reduced variability seen between staining rounds, in a large histochemical study. The current technique has been optimized for using rodent brain tissue in downstream immunohistochemical analyses; however, it can be applied to different scientific fields that use cryosectioning.

The brain is capable of improving from a chronically stressed state. The hippocampus in particular appears to "recover" from chronic stress-induced morphological and functional deficits following a post-stress rest period of several weeks. We previously found that hippocampal brain-derived neurotrophic factor (BDNF) was necessary for spatial ability to improve following a post-stress rest period. The following studies are the first to investigate the involvement of BDNF and its TrkB receptor on the recovery process following the end of chronic stress, as it pertains to hippocampal dendritic retraction and spatial memory deficits. In the first study, hippocampal BDNF was downregulated via RNA interference and then hippocampal CA3 and CA1 dendritic complexity were evaluated following chronic stress and a post-stress rest period in male Sprague-Dawley rats. Downregulating hippocampal BDNF prevented the enhancement of CA3 apical dendritic complexity following the rest period. Moreover, chronic stress and downregulated BDNF in the post-stress rest group led to regionally specific enhancements in CA1 dendritic complexity. In the second study, we tested whether the TrkB receptor was involved by administering daily systemic injections of ANA-12, a TrkB receptor antagonist, during the three-week post-stress rest period. ANA-12 prevented the improvement in spatial ability and CA3 apical dendritic complexity following the post-stress rest period. These data demonstrate that hippocampal BDNF acting via its TrkB receptor is necessary during the post-stress rest period in order to improve the impaired hippocampal structural and cognitive outcomes that occur in response to chronic stress., (Copyright © 2018 IBRO. Published by Elsevier Ltd. All rights reserved.)

Synthetic cathinones, frequently referred to as "bath salts", have significant abuse potential, and recent evidence suggests that these novel psychoactive substances can also produce cognitive deficits as well as cytotoxic effects. However, most of these latter findings have been obtained either using high concentrations in vitro or following non-contingent high dose administration in vivo. The present study utilized a model of long-term voluntary binge-like self-administration to determine potential detrimental effects of synthetic cathinones on cognitive function and their known underlying neural circuits, collectively referred to as neurocognitive dysfunction. Male Sprague-Dawley rats were allowed to self-administer the cocaine-like synthetic cathinone 3,4-methylenedioxypyrovalerone (MDPV, 0.03 mg/kg/infusion i.v.) in 96-hr sessions, or saline as a control. A total of five 96-hr sessions were conducted, each separated by 3 days of abstinence in the home cage. Three weeks following the last 96-hr session, animals underwent assessment of cognitive function using spatial object recognition (SOR) and novel object recognition (NOR) tasks, after which brains were harvested and assessed for neurodegeneration using FluoroJade C (FJC). Compared to animals self-administering saline, animals self-administering MDPV demonstrated (1) robust drug intake that escalated over time, (2) deficits in NOR but not SOR, and (3) neurodegeneration in the perirhinal and entorhinal cortices. These results indicate that repeated binge-like intake of MDPV can induce neurocognitive dysfunction. In addition, utilization of rodent models of extended binge-like intake may provide insight into potential mechanisms and/or approaches to prevent or reverse the detrimental effects of abused substances on cognitive and neurobiological functioning. This article is part of the Special Issue entitled 'Designer Drugs and Legal Highs.', (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Chronic stress results in functional and structural changes to the brain and especially the hippocampus. Decades of research have provided insights into the mechanisms by which chronic stress impairs hippocampal-mediated cognition and the corresponding reduction of hippocampal CA3 apical dendritic complexity. Yet, when chronic stress ends and time passes, which we refer to as a "post-stress rest period," hippocampal-mediated spatial memory deficits begin to improve and CA3 apical dendritic arbors increase in complexity. The processes by which the hippocampus improves from a chronically stressed state are not simply the reversal of the mechanisms that produced spatial memory deficits and CA3 apical dendritic retraction. This review will discuss our current understanding of how a chronically stressed hippocampus improves after a post-stress rest period. Untangling the mechanisms that allow for this post-stress plasticity is a critical next step in understanding how to promote resilience in the face of stressors., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Chronic stress leads to a dysregulated inhibitory tone that could impact hippocampal-dependent spatial learning and memory. The present study examined whether spatial memory deficits resulting from chronic stress could be overcome by antagonizing the GABA A receptor, a prominent inhibitory receptor of GABA in the hippocampus. Young adult male Sprague-Dawley rats were chronically stressed (STR, wire mesh restraint, 6h/d/21d) or placed in a no-stress control group (CON). When chronic restraint ended, rats were tested on a 2-trial object placement (OP) task at a delay (3h) that would result in chance performance without intervention and then on novel object recognition (NOR) and the elevated plus maze (EPM) to assess non-spatial memory and anxiety profile. In CON rats, Bicuculline (BIC, 0, 0.25, 0.5mg/kg), a GABA A antagonist, injected 30min prior to training led to facilitated OP performance with 0.25 and 0.5mg/kg doses. In contrast, STR rats required BIC at the highest dose (0.5mg/kg) to improve OP performance. While overall object exploration was decreased by chronic stress, motivation or anxiety profile were unlikely to explain these results. These findings reveal two different dose response functions for BIC in control and chronically stressed rats, with the dose response function of BIC being shifted to the right for chronically stressed rats compared to controls in order to improve spatial memory. While the literature demonstrates that chronic stress disrupts hippocampal inhibitory tone, the current study reveals that a single injection to antagonize the GABA A receptor can restore hippocampal-dependent spatial memory in chronically stressed subjects., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The current understanding of how chronic stress impacts hippocampal dendritic arbor complexity and the subsequent relationship to hippocampal-dependent spatial memory is reviewed. A surge in reports investigating hippocampal dendritic morphology is occurring, but with wide variations in methodological detail being reported. Consequently, this review systematically outlines the basic neuroanatomy of relevant hippocampal features to help clarify how chronic stress or glucocorticoids impact hippocampal dendritic complexity and how these changes occur in parallel with spatial cognition. Chronic stress often leads to hippocampal CA3 apical dendritic retraction first with other hippocampal regions (CA3 basal dendrites, CA1, dentate gyrus, DG) showing dendritic retraction when chronic stress is sufficiently robust or long lasting. The stress-induced reduction in hippocampal CA3 apical dendritic arbor complexity often coincides with impaired hippocampal function, such as spatial learning and memory. Yet, when chronic stress ends and a post-stress recovery period ensues, the atrophied dendritic arbors and poor spatial abilities often improve. However, this process differs from a simple reversal of chronic stress-induced deficits. Recent reports suggest that this return to baseline-like functioning is uniquely different from non-stressed controls, emphasizing the need for further studies to enhance our understanding of how a history of stress subsequently alters an organism's spatial abilities. To provide a consistent framework for future studies, this review concludes with an outline for a quick and easy reference on points to consider when planning chronic stress studies with the goal of measuring hippocampal dendritic complexity and spatial ability., (Copyright © 2016 Elsevier Inc. All rights reserved.)

In the pathophysiology of traumatic brain injury (TBI), the amygdala remains understudied, despite involvement in processing emotional and stressful stimuli associated with anxiety disorders, such as post-traumatic stress disorder (PTSD). Because the basolateral amygdala (BLA) integrates inputs from sensory and other limbic structures coordinating emotional learning and memory, injury-induced changes in circuitry may contribute to psychiatric sequelae of TBI. This study quantified temporal changes in dendritic complexity of BLA neurons after experimental diffuse TBI, modeled by midline fluid percussion injury. At post-injury days (PIDs) 1, 7, and 28, brain tissue from sham and brain-injured adult, male rats was processed for Golgi, glial fibrillary acidic protein (GFAP), or silver stain and analyzed to quantify BLA dendritic branch intersections, activated astrocytes, and regional neuropathology, respectively. Compared to sham, brain-injured rats at all PIDs showed enhanced dendritic branch intersections in both pyramidal and stellate BLA neuronal types, as evidenced by Sholl analysis. GFAP staining in the BLA was significantly increased at PID1 and 7 in comparison to sham. However, the BLA was relatively spared from neuropathology, demonstrated by an absence of argyrophilic accumulation over time, in contrast to other brain regions. These data suggest an early and persistent enhancement of dendritic complexity within the BLA after a single diffuse TBI. Increased dendritic complexity would alter information processing into and through the amygdala, contributing to emotional symptoms post-TBI, including PTSD., Competing Interests: Author Disclosure Statement No competing financial interests exist.

Chronic restraint stress alters hippocampal-dependent spatial learning and memory in a sex-dependent manner, impairing spatial performance in male rats and leaving intact or facilitating performance in female rats. Moreover, these stress-induced spatial memory deficits improve following post-stress recovery in males. The current study examined whether restraint administered in an unpredictable manner would eliminate these sex differences and impact a post-stress period on spatial ability and limbic glutamic acid decarboxylase (GAD65) expression. Male (n=30) and female (n=30) adult Sprague-Dawley rats were assigned to non-stressed control (Con), chronic stress (Str-Imm), or chronic stress given a post-stress recovery period (Str-Rec). Stressed rats were unpredictably restrained for 21 days using daily non-repeated combinations of physical context, duration, and time of day. Then, all rats were tested on the radial arm water maze (RAWM) for 2 days and given one retention trial on the third day, with brains removed 30min later to assess GAD65 mRNA. In Str-Imm males, deficits occurred on day 1 of RAWM acquisition, an impairment that was not evident in the Str-Rec group. In contrast, females did not show significant outcomes following chronic stress or post-stress recovery. In males, amygdalar GAD65 expression negatively correlated with RAWM performance on day 1. In females, hippocampal CA1 GAD65 positively correlated with RAWM performance on day 1. These results demonstrate that GABAergic function may contribute to the sex differences observed following chronic stress. Furthermore, unpredictable restraint and a recovery period failed to eliminate the sex differences on spatial learning and memory., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Chronic restraint stress impairs hippocampal-mediated spatial learning and memory, which improves following a post-stress recovery period. Here, we investigated whether brain-derived neurotrophic factor (BDNF), a protein important for hippocampal function, would alter the recovery from chronic stress-induced spatial memory deficits. Adult male Sprague-Dawley rats were infused into the dorsal hippocampal cornu ammonis (CA)3 region with an adeno-associated viral vector containing the sequence for a short hairpin RNA (shRNA) directed against BDNF or a scrambled sequence (Scr). Rats were then chronically restrained (wire mesh, 6 h/day for 21 days) and assessed for spatial learning and memory using a radial arm water maze (RAWM) either immediately after stressor cessation (Str-Imm) or following a 21-day post-stress recovery period (Str-Rec). All groups learned the RAWM task similarly, but differed on the memory retention trials. Rats in the Str-Imm group, regardless of adeno-associated viral contents, committed more errors in the spatial reference memory domain on the single retention trial during day 3 than did the non-stressed controls. Importantly, the typical improvement in spatial memory following the recovery from chronic stress was blocked with the shRNA against BDNF, as Str-Rec-shRNA performed worse on the RAWM compared with the non-stressed controls or Str-Rec-Scr. The stress effects were specific for the reference memory domain, but knockdown of hippocampal BDNF in unstressed controls briefly disrupted spatial working memory as measured by repeated entry errors on day 2 of training. These results demonstrated that hippocampal BDNF was necessary for the recovery from stress-induced hippocampal-dependent spatial memory deficits in the reference memory domain., (© 2014 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Chronic stress or glucocorticoid exposure simplifies hippocampal Cornu Ammonis region 3 (CA3) apical dendritic arbors in male rats. In contrast to males, chronic stress either reduces CA3 basal branching or exerts no observable morphological effects in gonadally intact female rats. Under conditions that females display stress-induced CA3 dendritic retraction, such as that following ovariectomy, chronic exposure to 17β-estradiol or cholesterol can negate these changes. Whether glucocorticoids produce CA3 dendritic retraction in ovariectomized females and whether neuroprotection from 17β-estradiol or cholesterol is sex-specific remains unknown. The current study examined the effects of chronic glucocorticoid exposure, in conjunction with 17β-estradiol or cholesterol administration, on hippocampal CA3 dendritic complexity. Adult male and female Sprague-Dawley rats were gonadectomized and implanted with 25% 17β-estradiol in cholesterol, 100% cholesterol, or blank Silastic capsules. Rats were then assigned to either a 21-day corticosterone (CORT) drink (400μg/ml CORT, 2.4% ethanol in tap water) or tap water (Tap, 2.4% ethanol in tap water) treatment. Brains were processed for Golgi staining, and hippocampal CA3 dendritic architecture was quantified. Results showed 21-day CORT administration reduced hippocampal CA3 apical dendritic branch points, CA3 apical dendritic length, body weight gain, and adrenal weights compared to male and female control counterparts. Furthermore, male and female rats implanted with Silastic capsules containing cholesterol or 25% 17β-estradiol in cholesterol were protected from CORT-induced CA3 apical dendritic branch reduction. No effects were observed in the CA3 basal dendritic arbors. The present results demonstrate that CORT produces hippocampal CA3 dendritic retraction in gonadectomized male and female rats and that cholesterol and 25% 17β-estradiol in cholesterol prevent this dendritic simplification., (Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.)

Chronic stress results in reversible spatial learning impairments in the Morris water maze that correspond with hippocampal CA3 dendritic retraction in male rats. Whether chronic stress impacts different types of memory domains, and whether these can similarly recover, is unknown. This study assessed the effects of chronic stress with and without a post-stress delay to evaluate learning and memory deficits within two memory domains, reference and working memory, in the radial arm water maze (RAWM). Three groups of 5-month-old male Sprague-Dawley rats were either not stressed [control (CON)], or restrained (6 h/day for 21 days) and then tested on the RAWM either on the next day [stress immediate (STR-IMM)] or following a 21-day delay [stress delay (STR-DEL)]. Although the groups learned the RAWM task similarly, groups differed in their 24-h retention trial assessment. Specifically, the STR-IMM group made more errors within both the spatial reference and working memory domains, and these deficits corresponded with a reduction in apical branch points and length of hippocampal CA3 dendrites. In contrast, the STR-DEL group showed significantly fewer errors in both the reference and working memory domains than the STR-IMM group. Moreover, the STR-DEL group showed better RAWM performance in the reference memory domain than did the CON group, and this corresponded with restored CA3 dendritic complexity, revealing long-term enhancing actions of chronic stress. These results indicate that chronic stress-induced spatial working and reference memory impairments, and CA3 dendritic retraction, are reversible, with chronic stress having lasting effects that can benefit spatial reference memory, but with these lasting beneficial effects being independent of CA3 dendritic complexity., (© 2011 The Authors. European Journal of Neuroscience © 2011 Federation of European Neuroscience Societies and Blackwell Publishing Ltd.)

Background: The electronic Schizophrenia Treatment Adherence Registry (e-STAR) is a prospective, observational study of patients with schizophrenia designed to evaluate long-term treatment outcomes in routine clinical practice., Methods: Parameters were assessed at baseline and at 3 month intervals for 2 years in patients initiated on risperidone long-acting injection (RLAI) (n=1345) or a new oral antipsychotic (AP) (n=277; 35.7% and 36.5% on risperidone and olanzapine, respectively) in Spain. Hospitalization prior to therapy was assessed by a retrospective chart review., Results: At 24 months, treatment retention (81.8% for RLAI versus 63.4% for oral APs, p<0.0001) and reduction in Clinical Global Impression Severity scores (-1.14 for RLAI versus -0.94 for APs, p=0.0165) were significantly higher with RLAI. Compared to the pre-switch period, RLAI patients had greater reductions in the number (reduction of 0.37 stays per patient versus 0.2, p<0.05) and days (18.74 versus 13.02, p<0.01) of hospitalizations at 24 months than oral AP patients., Conclusions: This 2 year, prospective, observational study showed that, compared to oral antipsychotics, RLAI was associated with better treatment retention, greater improvement in clinical symptoms and functioning, and greater reduction in hospital stays and days in hospital in patients with schizophrenia. Improved treatment adherence, increased efficacy and reduced hospitalization with RLAI offer the opportunity of substantial therapeutic improvement in schizophrenia.

Chronic obstructive pulmonary disease (COPD) is one of the main causes of death in Spain and elsewhere in the world, with an estimated 18,000 and 2.75 million deaths annually. Mortality is predicted to increase in the next few years due to smoking and the aging population. Multiple studies confirm that COPD is underreported as a cause of death on death certificates, due to the difficulty of determining the final cause of death in these patients. The main causes of mortality in COPD range from lung cancer and cardiovascular disease in patients with mild COPD to respiratory failure in the most advanced stages. Fortunately, in the latest updates, guidelines for the management and treatment of the disease identify reduction of mortality as one of the main clinical objectives to be achieved in these patients., (Copyright © 2009 Sociedad Española de Neumología y Cirugía Torácica. Published by Elsevier Espana. All rights reserved.)

T cell antigen recognition involves the formation of a structured interface between antigen-presenting and T cells that facilitates the specific transmission of activating and desensitizing stimuli. The molecular machinery that organizes the signaling molecules and controls their disposition in response to activation remains poorly understood. We show here that in T cells Discs large (Dlg1), a PDZ domain-containing protein, is recruited upon activation to cortical actin and forms complexes with early participants in T cell activation. Transient overexpression of Dlg1 attenuates basal and Vav1-induced NFAT reporter activation. Reduction of Dlg1 expression by RNA interference enhances both CD3- and superantigen-mediated NFAT activation. Attenuation of antigen receptor signaling appears to be a complex, highly orchestrated event that involves the mutual segregation of important elements of the early signaling complex., (Copyright The Rockerfeller University Press)

The toxicity of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) has been demonstrated in the seabream Sparus aurata specimens. Liver presented hepatocytic alterations, with an increase of lipid droplets and glycogen granules. Ultrastructural modifications of hepatocytes included RER fractionation, glycogen augmentation, as well as a rise in the number of lipid droplets, vacuoles and secondary lysosomes. In the gills, secondary lamellar epithelium showed hyperplasia, hypertrophy and lamellar fusion on the edge of the filaments. At the end of the exposure period (1 pg1(-1) TCDD for 20 days), some organelles in epithelial cells of the secondary lamellae and the tubular system of the chloride cells appeared altered. In the liver of TCDD-exposed specimens, immunoreactive cytochrome P-450 1A was concentrated close to the cytoplasmic and nuclear membranes, and positive granules were also evident throughout cytoplasm of the hepatocytes. Significant cytochrome P-450 staining was especially evident in endothelium of the hepatic vascular system. At the beginning of the exposure (1 pg 1(-1) TCDD, for 5 and 10 days), cytochrome P-450 immunostaining was observed in the cytoplasm of scarce hepatic cells and after 20 days of treatment, specific immunostained cytoplasmic granules were detected in most hepatocytes. In gills of TCDD-treated specimens, pillar-endothelial cells showed a cytochrome P-450 1A immunostaining concentrated close to the base of gill filaments and dispersed through the gill lamellae. There was also significant cytochrome staining of the endothelium of the branchial vascular system. However, no cytochrome immunoreactivity was observed in epithelial-respiratory cells.

In this paper we investigate by means of immunohistochemistry, the tissue distribution of constitutive cytochrome P4501A (CYPIA), from hatching until 30 days posthatching in developing Siberian sturgeon, Acipenser baeri. For this purpose, a polyclonal (BN-1) antiserum developed against a conservative sequence of piscine CYP1A and a monoclonal (C10-7) antiserum directed against cod CYP1A were used on paraffin-embedded samples. From hatching onwards, distinct CYP1A immunoreactivity was distinctly observed in the following tissues and cells: envelope of oil droplets, matrix and syncytium of the yolk-sac, sinusoids, biliary epithelial cells and hepatocytes. In the digestive tract, buccopharyngeal, oesophageal, gastric and intestinal epithelia, as well as the cytoplasm and brush border of enterocytes were CYP1A-positive. Interestingly, gastric glands and melanin-plug present within lumen of the digestive system were strongly immunoreactive. Kidney (epithelia of renal tubules), gills (pillar and endothelial cells), skin (epithelial cells), muscle fibres of heart and eye (retina) were positive. In brain, we observed a strong CYP1A staining in the developing telencephalon and especially in olfactory system, as well as in those nerve fibres running ventrally toward the posterior brain. A strong CYP1A staining was observed in vascular endothelia of all organs/tissues, especially in the liver. In general, the intensity of CYP1A immunostaining increased during larval development, suggesting besides its known metabolic function (endogenous and/or exogenous), a possible participation of this heme-protein in control of cell division, regulation of growth and differentiation.

CYP1A is a major inducible enzyme in the metabolism of xenobiotic substrates. In this paper we investigate by means of immunohistochemistry, the tissue distribution of constitutive cytochrome P4501A (CYP1A) during the period of endogenous nutrition (from hatching until day 4) in developing gilthead seabream, Sparus aurata larvae. For this purpose, a polyclonal antiserum (BN-1, Biosense Laboratories) directed against conserved piscine CYP1A sequences was used on paraffin-embedded sections from seabream larvae. From hatching onward, CYP1A immunoreactivity was observed in the following tissues and cells: syncytial, oil-globule envelopes and matrix of the yolk-sac, kidney (epithelia of renal tubules), cardiac muscle cells, skin epidermal cells, troncal musculature, enterocytes of different intestinal regions, goblet cells of the bucco-pharyngeal region, gill epithelial cells and the endothelia of the vascular system of various tissues (especially from liver and brain). Moreover, eye (retina), olfactory epithelium and some positive nerve fibers located in the proximity of the olfactory bulbs and running ventrally toward the posterior brain were strongly CYP1A immunoreactive. In general, the intensity of immunostaining increased with larval development.

The mouse lymphoma (L5178Y) cell mutant M10 is defective in rejoining DNA double-strand breaks and is hypersensitive to ionizing radiation. The introduction of human chromosome 5 into M10 cells by microcell mediated chromosome transfer complemented the ionizing-radiation hypersensitivity defect of this cell line. The presence of chromosome 5 in the microcell hybrids was shown using PCR with chromosome-specific primers and fluorescence in situ hybridization. From this data we conclude that the gene that corrects the radiation hypersensitivity of M10 cells is located on chromosome 5 and tentatively assigned to the 5q14 to 5pter region. We designate this gene XRCC4L.

Objective: To assess the effectiveness of verapamil, hydrocortisone sodium succinate, and phosphatidylcholine in the prevention of experimental adhesions., Design: Randomized trial., Material: 80 rats., Interventions: Laparotomy and intraperitoneal irrigation with saline 40 degrees C, then verapamil hydrochloride 1 mg/kg intravenously 15 min before, during, and after irrigation; or hydrocortisone sodium succinate 50 mg/kg intravenously half an hour before irrigation; or phosphatidylcholine 5.5 mg/kg orally eight days before and seven days after irrigation plus 0.5 mg/ml in the irrigation fluid; or no further intervention., Main Outcome Measures: Development of adhesions two weeks after irrigation, and completeness of wound healing., Results: Adhesions developed in 13 of 19 control animals; 7 of 20 that were given verapamil; 6 of 20 that were given hydrocortisone; and 3 of 20 given phosphatidylcholine., Conclusion: Adhesions that developed in rats after laparotomy and intraperitoneal irrigation with saline at 40 degrees C can be significantly reduced by phosphatidylcholine.

We investigated the unstimulated and stimulated migratory activities of lavaged alveolar macrophages (AMs) in vitro over the course of alveolar clearance of three different mass lung burdens of microspheres. Our intent was to uncover potentially important relationships between the migratory behaviors of the AM and the retention kinetics of particles. Groups of adult, male Fischer-344 rats were intratracheally instilled with approximately 86 micrograms (low burden, LB), approximately 1 mg (medium burden, MB), or approximately 3.7 mg (high burden, HB) of polystyrene microspheres (2.13 microns diameter), or with carrier vehicle (phosphate buffered saline, PBS) alone. The lung retention kinetics of the particles were determined over an approximately 170 day period. On days 14, approximately 57, and approximately 85, lavaged AMs were assessed for their abilities to migrate through 5-microns pore membranes in response to inactivated rat serum (unstimulated condition) and yeast-activated rat serum (stimulated condition). The retention characteristics of the three burdens could be satisfactorily described by two-component, negative exponential equations. The kinetics of retention of the LB and MB were similar, although some evidence indicated the MB slightly retarded the lung clearance process. Deposition of the HB resulted in more marked prolongations of both the early, more rapid, and the slower, longer term components of alveolar clearance. The unstimulated migration indices of AMs from the particle-instilled lungs were generally not significantly different from those of AMs from PBS-instilled lungs except for a significant increase in the migration indices of LB AMs at the last assay time. The stimulated migration indices of MB and HB AMs were significantly decreased on assay days 14 and approximately 57. On day approximately 85, however, the migration indices of LB, MB, and HB AMs were all increased above the PBS AMs. Comparisons of the frequency distributions of particles in the unstimulated and stimulated AM that migrated to those in corresponding parent AM populations consistently indicated a preferential migration of particle-free AMs and of AMs with lesser loads of microspheres. The overall results of this study suggest that the unstimulated and stimulated migratory activities of particle-laden AMs are depressed in vitro. Our results also suggest that the migratory activities of generally particle-free AMs may be enhanced over a prolonged period of time following the deposition of particles in the lung.