Cortical Brain Tissue Research Articles

Mechanism of myeloid differentiation factor 2 on mediating sepsis-associated encephalopathy

To investigate the role and underlying mechanism of human myeloid differentiation protein 2 (MD2) in the process of neuronal death induced by lipopolysaccharide (LPS) by establishing an in vitro model of sepsis-associated encephalopathy (SAE) by LPS. Healthy C57BL/6J mice at 14-18 days of gestation were selected, and brain cortical tissue was taken from fetal mice. Neurons were stimulated with 0 (control), 1, 5 and 10 g/L of LPS for 24 hours. The release of lactate dehydrogenase (LDH) was detected and the death of neurons was observed. Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of inflammatory factors interleukins (IL-6, IL-1β), in order to determine the optimal dose of LPS for establishing an in vitro neuroinflammation model of SAE. The cells were divided into blank control group and LPS group. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end-labeling (TUNEL) was used to discover apoptosis. Western blotting was used to detect the expression of the relevant protein markers activated caspase-3, necroptosis-associated protein neuronal receptor-interacting serine/threonine-protein kinase 3 (RIPK3) and phosphorylated RIPK3 (p-RIPK3). Immunofluorescence chemical staining was used to detect the expressions of p-RIPK3 and microtubule-associated protein 2 (MAP2) to evaluate the type of cell death and the degree of neuronal death. Western blotting was used to detect MD2 expression. Immunofluorescence chemical staining was performed to observe the expression and distribution of p-RIPK3 and MD2 in neurons to assess whether MD2 was involved in the inflammatory response promoting neuronal death. In addition, the cells were divided into blank control group, LPS group, and MD2 interfering peptide group (LPS+TC group), and the levels of IL-6, IL-1β and LDH were detected to evaluate whether interfering with MD2 can alleviate LPS induced neuroinflammation. 10 g/L LPS induced notable neuronal death, and the release of LDH in neurons stimulated with this concentration for 24 hours was significantly higher than that in the blank control group (relative release: 1.45±0.04 vs. 1.00±0.00, P < 0.01), indicating apoptosis and necroptosis occurred in neurons, and the levels of inflammatory factors IL-6 and IL-1β were remarkable increased [IL-6 (relative level): 1.94±0.04 vs. 1.00±0.00, IL-1β (relative level): 1.53±0.09 vs. 1.00±0.00, both P < 0.01]. Compared with the blank control group, the apoptosis of cells, cleaved-caspase-3 expression, the p-RIPK3/RIPK3 ratio, and p-RIPK3 expression around neurons in the LPS group were significantly increased [cleaved-caspase-3/GAPDH: 1.55±0.10 vs. 1.00±0.00, P < 0.01; p-RIPK3/RIPK3 ratio (relative value): 1.54±0.06 vs. 1.00±0.00, P < 0.05], which suggested that typical apoptosis and necroptosis apoptosis occurred in neurons in the septic environment. Furthermore, MD2 expression was significantly increased in the LPS group compared with the blank control group (MD2/GAPDH: 1.91±0.07 vs. 1.00±0.00, P < 0.01), and MD2 expression around neurons was increased, indicating that LPS-induced MD2 upregulation may play a key role in neuroinflammation and induction of neuronal death in sepsis. In addition, compared with the LPS group, the MD2-interfering peptide could reduce the expression levels of inflammatory factors IL-6 and IL-1β [IL-6 (relative level): 1.16±0.08 vs. 1.94±0.04, IL-1β (relative level): 1.15±0.05 vs. 1.75±0.09, both P < 0.01] and decrease LDH release (relative release: 1.09±0.01 vs. 1.44±0.04, P < 0.05). LPS induced neuronal inflammatory responses via MD2, which ultimately leads to apoptosis and necroptosis. Interfering with MD2 reduces inflammation and inhibits neuronal death.

Associations of cortical SPP1 and ITGAX with cognition and common neuropathologies in older adults.

The secreted phosphoprotein 1 (SPP1) gene expressed by CD11c+ cells is known to be associated with microglia activation and neuroinflammatory diseases. As most studies rely on mouse models, we investigated these genes and proteins in the cortical brain tissue of older adults and their role in Alzheimer's disease (AD) and related disorders. We leveraged protein measurements, single-nuclei, and RNASeq data from the Religious Orders Study and Rush Memory and Aging Project (ROSMAP) of over 1200 samples for association analysis. Expression ofSPP1and its encoded protein osteopontin were associated with faster cognitive decline and greater odds of common neuropathologies. At single-cell resolution, integrin subunit alpha X (ITGAX) was highly expressed in microglia, where specific subpopulations were associated with AD and cerebral amyloid angiopathy. The study provides evidence of SPP1andITGAXassociation with cognitive decline and common neuropathologies identifying a microglial subset associated with disease.

A proteomics analysis of 5xFAD mouse brain regions reveals the lysosome-associated protein Arl8b as a candidate biomarker for Alzheimer’s disease

BackgroundAlzheimer’s disease (AD) is characterized by the intra- and extracellular accumulation of amyloid-β (Aβ) peptides. How Aβ aggregates perturb the proteome in brains of patients and AD transgenic mouse models, remains largely unclear. State-of-the-art mass spectrometry (MS) methods can comprehensively detect proteomic alterations, providing relevant insights unobtainable with transcriptomics investigations. Analyses of the relationship between progressive Aβ aggregation and protein abundance changes in brains of 5xFAD transgenic mice have not been reported previously.MethodsWe quantified progressive Aβ aggregation in hippocampus and cortex of 5xFAD mice and controls with immunohistochemistry and membrane filter assays. Protein changes in different mouse tissues were analyzed by MS-based proteomics using label-free quantification; resulting MS data were processed using an established pipeline. Results were contrasted with existing proteomic data sets from postmortem AD patient brains. Finally, abundance changes in the candidate marker Arl8b were validated in cerebrospinal fluid (CSF) from AD patients and controls using ELISAs.ResultsExperiments revealed faster accumulation of Aβ42 peptides in hippocampus than in cortex of 5xFAD mice, with more protein abundance changes in hippocampus, indicating that Aβ42 aggregate deposition is associated with brain region-specific proteome perturbations. Generating time-resolved data sets, we defined Aβ aggregate-correlated and anticorrelated proteome changes, a fraction of which was conserved in postmortem AD patient brain tissue, suggesting that proteome changes in 5xFAD mice mimic disease-relevant changes in human AD. We detected a positive correlation between Aβ42 aggregate deposition in the hippocampus of 5xFAD mice and the abundance of the lysosome-associated small GTPase Arl8b, which accumulated together with axonal lysosomal membranes in close proximity of extracellular Aβ plaques in 5xFAD brains. Abnormal aggregation of Arl8b was observed in human AD brain tissue. Arl8b protein levels were significantly increased in CSF of AD patients.ConclusionsWe report a comprehensive biochemical and proteomic investigation of hippocampal and cortical brain tissue derived from 5xFAD transgenic mice, providing a valuable resource to the neuroscientific community. We identified Arl8b, with significant abundance changes in 5xFAD and AD patient brains. Arl8b might enable the measurement of progressive lysosome accumulation in AD patients and have clinical utility as a candidate biomarker.

Mesh-free high-resolution simulation of cerebrocortical oxygen supply with fast Fourier preconditioning.

Oxygen transfer from blood vessels to cortical brain tissue is representative of a class of problems with mixed-domain character. Large-scale efficient computation of tissue oxygen concentration is dependent on the manner in which the tubular network of blood vessels is coupled to the tissue mesh. Models which explicitly resolve the interface between the tissue and vasculature with a contiguous mesh are prohibitively expensive for very dense cerebral microvasculature. We propose a mixed-domain mesh-free technique whereby a vascular anatomical network (VAN) represented as a thin directed graph serves for convection of blood oxygen, and the surrounding extravascular tissue is represented as a Cartesian grid of 3D voxels throughout which oxygen is transported by diffusion. We split the network and tissue meshes by the Schur complement method of domain decomposition to obtain a reduced set of system equations for the tissue oxygen concentration at steady state. The use of a Cartesian grid allows the corresponding matrix equation to be solved approximately with a fast Fourier transform-based Poisson solver, which serves as an effective preconditioner for Krylov subspace iteration. The performance of this method enables the steady-state simulation of cortical oxygen perfusion for anatomically accurate vascular networks down to single micron resolution without the need for supercomputers.

Determination of Krogh Coefficient for Oxygen Consumption Measurement from Thin Slices of Rodent Cortical Tissue Using a Fick's Law Model of Diffusion.

To investigate the impact of experimental interventions on living biological tissue, ex vivo rodent brain slices are often used as a more controllable alternative to a live animal model. However, for meaningful results, the biological sample must be known to be healthy and viable. One of the gold-standard approaches to identifying tissue viability status is to measure the rate of tissue oxygen consumption under specific controlled conditions. Here, we work with thin (400 μm) slices of mouse cortical brain tissue which are sustained by a steady flow of oxygenated artificial cerebralspinal fluid (aCSF) at room temperature. To quantify tissue oxygen consumption (Q), we measure oxygen partial pressure (pO2) as a function of probe depth. The curvature of the obtained parabolic (or parabola-like) pO2 profiles can be used to extract Q, providing one knows the Krogh coefficient Kt, for the tissue. The oxygen trends are well described by a Fick's law diffusion-consumption model developed by Ivanova and Simeonov, and expressed in terms of ratio (Q/K), being the rate of oxygen consumption in tissue divided by the Krogh coefficient (oxygen diffusivity × oxygen solubility) for tissue. If the fluid immediately adjacent to the tissue can be assumed to be stationary (i.e., nonflowing), one may invoke conservation of oxygen flux K·(∂P/∂x) across the interface to deduce (Kt/Kf), the ratio of Krogh coefficients for tissue and fluid. Using published interpolation formulas for the effect of salt content and temperature on oxygen diffusivity and solubility for pure water, we estimate Kf, the Krogh coefficient for aCSF, and hence deduce the Kt coefficient for tissue. We distinguish experimental uncertainty from natural biological variability by using pairs of repeated profiles at the same tissue location. We report a dimensionless Krogh ratio (Kt/Kf)=0.562±0.088 (mean ± SD), corresponding to a Krogh coefficient Kt=(1.29±0.21)×10-14 mol/(m·s·Pa) for mouse cortical tissue at room temperature, but acknowledge the experimental limitation of being unable to verify that the fluid boundary layer is truly stationary. We compare our results with those reported in the literature, and comment on the challenges and ambiguities caused by the extensive use of 'biologically convenient' non-SI units for tissue Krogh coefficient.

Thiamethoxam evoked neural oxido-inflammatory stress in male rats through modulation of Nrf2/NF-kB/iNOS signaling and inflammatory cytokines: Neuroprotective effect of Silymarin

Thiamethoxam (TMX), a neonicotinoid insecticide, is a widely used insecticide with neurotoxic potential. Silymarin (SM), a milk thistle-derived flavonoid, is known with its promising biological activities. This study explored the neuroprotective effects of SM against TMX-triggered cortical injury in male rats. Animals were divided into four groups and treated daily either with SM (150 mg/kg), TMX (78.15 mg/kg), or both at the aforementioned doses for 28 days. Our results revealed marked declines in cortical SOD and CAT activities with elevations in MDA, IL-1b and TNF-α levels in TMX-treated rats. Further, TMX induced down-regulation in the gene expressions of Sod, Cat, Gpx, and Nrf-2, with up-regulation in the gene expressions of IL-1b, IL-6, iNOS, TNF-α and NF-kB. Interestingly, pre-treatment with SM provided a notable neuroprotective action against TMX-mediated cortical damage that indicates its promising antioxidant and anti-inflammatory activities. This effect may be mediated by Nrf2/NF-kB/iNOS signalling and suppression of excess free radicals and production of inflammatory cytokines. In brief, SM could be a promising therapeutic agent against TMX-mediated neural complication via its antioxidant and anti-inflammatory properties. Practical applicationsThe using of neonicotinoids as thiamethoxam is recently increased and is associated with brain damage. TMX induced excessive oxidative and inflammatory damage. Therefore, new therapeutic approaches are needed to counteract its adverse effects on the nervous system. SM, a flavonoid, is extracted from the seeds and fruits of milk thistle. Due to its potent antioxidative activity, SM have been applied to mitigate the oxidative stress as well as inflammatory disorders. Herein, we examined the potential therapeutic role of SM against TMX-induced brain oxidative stress and inflammation in rats through evaluating oxidative markers, inflammatory response, and histopathological changes in the brain cortical tissue.

Modulating Brain Activity with Invasive Brain-Computer Interface: A Narrative Review.

Brain-computer interface (BCI) can be used as a real-time bidirectional information gateway between the brain and machines. In particular, rapid progress in invasive BCI, propelled by recent developments in electrode materials, miniature and power-efficient electronics, and neural signal decoding technologies has attracted wide attention. In this review, we first introduce the concepts of neuronal signal decoding and encoding that are fundamental for information exchanges in BCI. Then, we review the history and recent advances in invasive BCI, particularly through studies using neural signals for controlling external devices on one hand, and modulating brain activity on the other hand. Specifically, regarding modulating brain activity, we focus on two types of techniques, applying electrical stimulation to cortical and deep brain tissues, respectively. Finally, we discuss the related ethical issues concerning the clinical application of this emerging technology.

Proteomics of the dentate gyrus reveals semantic dementia specific molecular pathology

Semantic dementia (SD) is a clinical subtype of frontotemporal dementia consistent with the neuropathological diagnosis frontotemporal lobar degeneration (FTLD) TDP type C, with characteristic round TDP-43 protein inclusions in the dentate gyrus. Despite this striking clinicopathological concordance, the pathogenic mechanisms are largely unexplained forestalling the development of targeted therapeutics. To address this, we carried out laser capture microdissection of the dentate gyrus of 15 SD patients and 17 non-demented controls, and assessed relative protein abundance changes by label-free quantitative mass spectrometry. To identify SD specific proteins, we compared our results to eight other FTLD and Alzheimer’s disease (AD) proteomic datasets of cortical brain tissue, parallel with functional enrichment analyses and protein–protein interactions (PPI). Of the total 5,354 quantified proteins, 151 showed differential abundance in SD patients (adjusted P-value < 0.01). Seventy-nine proteins were considered potentially SD specific as these were not detected, or demonstrated insignificant or opposite change in FTLD/AD. Functional enrichment indicated an overrepresentation of pathways related to the immune response, metabolic processes, and cell-junction assembly. PPI analysis highlighted a cluster of interacting proteins associated with adherens junction and cadherin binding, the cadherin-catenin complex. Multiple proteins in this complex showed significant upregulation in SD, including β-catenin (CTNNB1), γ-catenin (JUP), and N-cadherin (CDH2), which were not observed in other neurodegenerative proteomic studies, and hence may resemble SD specific involvement. A trend of upregulation of all three proteins was observed by immunoblotting of whole hippocampus tissue, albeit only significant for N-cadherin. In summary, we discovered a specific increase of cell adhesion proteins in SD constituting the cadherin-catenin complex at the synaptic membrane, essential for synaptic signaling. Although further investigation and validation are warranted, we anticipate that these findings will help unravel the disease processes underlying SD.

Increased Sialylation in Alzheimer’s Disease Models

AbstractBackgroundResearch shows evidence of increased sialylation localized to pathological protein aggregates in AD autopsied brains. Sialylation is a type of glycosylation with linkage bonds of sialic acid (SA) residues on glycoproteins and glycolipids. It is unclear how changes in SA distribution in AD contributes to disease. Microglia are partially regulated by sialylation. We hypothesize decreased removal of SA residues on microglia leads to increased sialylation near amyloid‐β (Aβ) plaques, reducing microglia mediated plaque clearance.MethodA plant‐derived lectin was used to recognize α2‐6 SA bonds. Aβ plaques, microglia, and α2‐6 SA were histologically labelled in cortical brain tissue from human AD cases and 5XFAD mouse model of Aβ pathology. Stained hemibrains were imaged with confocal microscopy and quantified. To investigate the effects of reducing SA cleavage, we treated a subset of our 5XFAD and wild‐type (WT) mice with oseltamivir (Tamiflu), an inhibitor of desialylation, for five days. On day 3 and immediately following treatment, we measured cognitive and non‐cognitive behaviors using open field and novel object recognition tests.ResultQualitatively, α2‐6 SA levels displayed similar microglial‐like phenotype in 5XFAD and human AD cortical brain tissue. Two‐way ANOVA of histologically labeled hemibrains revealed 5XFAD mice had significantly increased α2‐6 SA levels compared to WT mice (p = 0.0007). High magnification image analysis supported a significant increase of α2‐6 SA area in 5XFAD mice versus WT (p&lt;0.0001), yet there was no effect of oseltamivir treatment. Behavioral data similarly demonstrated no effect of treatment. Post‐hoc analysis of open field revealed 5XFAD mice exhibited greater velocity compared to WT (p = 0.03) and 5XFAD mice lacked habituation behavior with no decrease in velocity compared to WT (p = 0.02). Investigation of Aβ pathology indicated no oseltamivir effect.ConclusionWe observed a significant increase in α2‐6 SA in 5XFAD mice similar to human AD, suggesting a plaque associated increase in sialylation. Next, we will closely examine localization of SA in the plaque microenvironment to understand the role of sialylation in microglial interactions with disease pathology. We postulate SA residues are localized to microglia interacting with Aβ plaques, potentially aggravating pathology.

TDP-43 condensates and lipid droplets regulate the reactivity of microglia and regeneration after traumatic brain injury.

Decreasing the activation of pathology-activated microglia is crucial to prevent chronic inflammation and tissue scarring. In this study, we used a stab wound injury model in zebrafish and identified an injury-induced microglial state characterized by the accumulation of lipid droplets and TAR DNA-binding protein of 43 kDa (TDP-43)+ condensates. Granulin-mediated clearance of both lipid droplets and TDP-43+ condensates was necessary and sufficient to promote the return of microglia back to the basal state and achieve scarless regeneration. Moreover, in postmortem cortical brain tissues from patients with traumatic brain injury, the extent of microglial activation correlated with the accumulation of lipid droplets and TDP-43+ condensates. Together, our results reveal a mechanism required for restoring microglia to a nonactivated state after injury, which has potential for new therapeutic applications in humans.

Endogenous PTEN-Induced Kinase 1 Regulates Dendritic Architecture and Spinogenesis.

Mutations in PTEN-induced kinase 1 (PINK1) contribute to autosomal recessive Parkinson's disease with cognitive and neuropsychiatric comorbidities. Disturbances in dendritic and spine architecture are hallmarks of neurodegenerative and neuropsychiatric conditions, but little is known of the impact of PINK1 on these structures. We used Pink1 -/- mice to study the role of endogenous PINK1 in regulating dendritic architecture, spine density, and spine maturation. Pink1 -/- cortical neurons of unknown sex showed decreased dendritic arborization, affecting both apical and basal arbors. Dendritic simplification in Pink1 -/- neurons was primarily driven by diminished branching with smaller effects on branch lengths. Pink1 -/- neurons showed reduced spine density with a shift in morphology to favor filopodia at the expense of mushroom spines. Electrophysiology revealed significant reductions in miniature EPSC (mEPSC) frequency in Pink1 -/- neurons, consistent with the observation of decreased spine numbers. Transfecting with human PINK1 rescued changes in dendritic architecture, in thin, stubby, and mushroom spine densities, and in mEPSC frequency. Diminished spine density was also observed in Golgi-Cox stained adult male Pink1 -/- brains. Western blot study of Pink1 -/- brains of either sex revealed reduced phosphorylation of NSFL1 cofactor p47, an indirect target of PINK1. Transfection of Pink1 -/- neurons with a phosphomimetic p47 plasmid rescued dendritic branching and thin/stubby spine density with a partial rescue of mushroom spines, implicating a role for PINK1-regulated p47 phosphorylation in dendrite and spine development. These findings suggest that PINK1-dependent synaptodendritic alterations may contribute to the risk of cognitive and/or neuropsychiatric pathologies observed in PINK1-mutated families.SIGNIFICANCE STATEMENT Loss of PINK1 function has been implicated in both familial and sporadic neurodegenerative diseases. Yet surprisingly little is known of the impact of PINK1 loss on the fine structure of neurons. Neurons receive excitatory synaptic signals along a complex network of projections that form the dendritic tree, largely at tiny protrusions called dendritic spines. We studied cortical neurons and brain tissues from mice lacking PINK1. We discovered that PINK1 deficiency causes striking simplification of dendritic architecture associated with reduced synaptic input and decreased spine density and maturation. These changes are reversed by reintroducing human PINK1 or one of its downstream mediators into PINK1-deficient mouse neurons, indicating a conserved function, whose loss may contribute to neurodegenerative processes.

Di-2-ethylhexyl phthalate affects zinc metabolism and neurogenesis in the developing rat brain

We previously observed that developmental marginal zinc deficiency affects neurogenesis. Maternal phthalate exposure could disrupt fetal zinc homeostasis by triggering an acute phase response, causing maternal liver zinc retention that limits zinc availability to the fetus. Thus, we currently investigated whether exposure to di-2-ethylhexyl phthalate (DEHP) during gestation in rats alters fetal brain neurogenesis by impairing zinc homeostasis. Dams consumed an adequate (25 μg zinc/g diet) (C) or a marginal zinc deficient (MZD) (10 μg zinc/g diet) diet, without or with DEHP (300 mg/kg BW) (C + DEHP, MZD + DEHP) from embryonic day (E) 0 to E19. To evaluate neurogenesis we measured parameters of neural progenitor cells (NPC) proliferation and differentiation. Maternal exposure to DEHP and/or zinc deficiency lowered fetal brain cortical tissue (CT) zinc concentrations. Transcription factors involved in NPC proliferation (PAX6, SOX2, EMX1), differentiation (TBR2, TBR1) and mature neurons (NeuN) were lower in MZD, MZD + DEHP and C + DEHP than in C E19 brain CT, being the lowest in the MZD + DEHP group. VGLUT1 levels, a marker of glutamatergic neurons, showed a similar pattern. Levels of a marker of GABAergic neurons, GAD65, did not vary among groups. Phosphorylated ERK1/2 levels were reduced by both MZD and DEHP, and particularly in the MZD + DEHP group. MEHP-treated human neuroblastoma IMR-32 cells and E19 brains from DEHP-treated dams showed that the zinc-regulated phosphatase PP2A can be in part responsible for DEHP-mediated ERK1/2 downregulation and impaired neurogenesis. Overall, gestational exposure to DEHP caused secondary zinc deficiency and impaired neurogenesis. These harmful effects could have long-term consequences on the adult offspring brain structure and function.

Organotypic whole hemisphere brain slice models to study the effects of donor age and oxygen-glucose-deprivation on the extracellular properties of cortical and striatal tissue

BackgroundThe brain extracellular environment is involved in many critical processes associated with neurodevelopment, neural function, and repair following injury. Organization of the extracellular matrix and properties of the extracellular space vary throughout development and across different brain regions, motivating the need for platforms that provide access to multiple brain regions at different stages of development. We demonstrate the utility of organotypic whole hemisphere brain slices as a platform to probe regional and developmental changes in the brain extracellular environment. We also leverage whole hemisphere brain slices to characterize the impact of cerebral ischemia on different regions of brain tissue.ResultsWhole hemisphere brain slices taken from postnatal (P) day 10 and P17 rats retained viable, metabolically active cells through 14 days in vitro (DIV). Oxygen-glucose-deprivation (OGD), used to model a cerebral ischemic event in vivo, resulted in reduced slice metabolic activity and elevated cell death, regardless of slice age. Slices from P10 and P17 brains showed an oligodendrocyte and microglia-driven proliferative response after OGD exposure, higher than the proliferative response seen in DIV-matched normal control slices. Multiple particle tracking in oxygen-glucose-deprived brain slices revealed that oxygen-glucose-deprivation impacts the extracellular environment of brain tissue differently depending on brain age and brain region. In most instances, the extracellular space was most difficult to navigate immediately following insult, then gradually provided less hindrance to extracellular nanoparticle diffusion as time progressed. However, changes in diffusion were not universal across all brain regions and ages.ConclusionsWe demonstrate whole hemisphere brain slices from P10 and P17 rats can be cultured up to two weeks in vitro. These brain slices provide a viable platform for studying both normal physiological processes and injury associated mechanisms with control over brain age and region. Ex vivo OGD impacted cortical and striatal brain tissue differently, aligning with preexisting data generated in in vivo models. These data motivate the need to account for both brain region and age when investigating mechanisms of injury and designing potential therapies for cerebral ischemia.

Lifelong Exposure to a Low-Dose of the Glyphosate-Based Herbicide RoundUp® Causes Intestinal Damage, Gut Dysbiosis, and Behavioral Changes in Mice.

RoundUp® (RUp) is a comercial formulation containing glyphosate (N-(phosphono-methyl) glycine), and is the world’s leading wide-spectrum herbicide used in agriculture. Supporters of the broad use of glyphosate-based herbicides (GBH) claim they are innocuous to humans, since the active compound acts on the inhibition of enzymes which are absent in human cells. However, the neurotoxic effects of GBH have already been shown in many animal models. Further, these formulations were shown to disrupt the microbiome of different species. Here, we investigated the effects of a lifelong exposure to low doses of the GBH-RUp on the gut environment, including morphological and microbiome changes. We also aimed to determine whether exposure to GBH-RUp could harm the developing brain and lead to behavioral changes in adult mice. To this end, animals were exposed to GBH-RUp in drinking water from pregnancy to adulthood. GBH-RUp-exposed mice had no changes in cognitive function, but developed impaired social behavior and increased repetitive behavior. GBH-Rup-exposed mice also showed an activation of phagocytic cells (Iba-1–positive) in the cortical brain tissue. GBH-RUp exposure caused increased mucus production and the infiltration of plama cells (CD138-positive), with a reduction in phagocytic cells. Long-term exposure to GBH-RUp also induced changes in intestinal integrity, as demonstrated by the altered expression of tight junction effector proteins (ZO-1 and ZO-2) and a change in the distribution of syndecan-1 proteoglycan. The herbicide also led to changes in the gut microbiome composition, which is also crucial for the establishment of the intestinal barrier. Altogether, our findings suggest that long-term GBH-RUp exposure leads to morphological and functional changes in the gut, which correlate with behavioral changes that are similar to those observed in patients with neurodevelopmental disorders.

Altered protein expression of membrane transporters in isolated cerebral microvessels and brain cortex of a rat Alzheimer's disease model

There is growing evidence that membrane transporters expressed at the blood-brain barrier (BBB) and brain parenchymal cells play an important role in Alzheimer's disease (AD) development and progression. However, quantitative information about changes in transporter protein expression at neurovascular unit cells in AD is limited. Here, we studied the changes in the absolute protein expression of five ATP-binding cassette (ABC) and thirteen solute carrier (SLC) transporters in the isolated brain microvessels and brain cortical tissue of TgF344-AD rats compared to age-matched wild-type (WT) animals using liquid chromatography tandem mass spectrometry based quantitative targeted absolute proteomic analysis. Moreover, sex-specific alterations in transporter expression in the brain cortical tissue of this model were examined. Protein expressions of Abcg2, Abcc1 and FATP1 (encoded by Slc27a1) in the isolated brain microvessels of TgF344-AD rats were 3.1-, 2.0-, 4.3-fold higher compared to WT controls, respectively (p < 0.05). Abcc1 and 4F2hc (encoded by Slc3a2) protein expression was significantly up-regulated in the brain cortical tissue of male TgF344-AD rats compared to male WT rats (p < 0.05). The study provides novel information for the elucidation of molecular mechanisms underlying AD and valuable knowledge about the optimal use of the TgF344-AD rat model in AD drug development and drug delivery research.

Repetitive Mild Closed Head Injury in Adolescent Mice Is Associated with Impaired Proteostasis, Neuroinflammation, and Tauopathy.

Repetitive mild traumatic brain injury (mTBI) in children and adolescents leads to acute and chronic neurologic sequelae and is linked to later life neurodegenerative disease. However, the biological mechanisms connecting early life mTBI to neurodegeneration remain unknown. Using an adolescent mouse repetitive closed head injury model that induces progressive cognitive impairment in males and anxiety in females in the absence of overt histopathology, we examined transcriptional and translational changes in neurons isolated from sham and injured brain in the chronic phase after injury. At 14 months, single-nuclei RNA sequencing of cortical brain tissue identified disruption of genes associated with neuronal proteostasis and evidence for disrupted ligand-receptor signaling networks in injured mice. Western blot analysis of isolated neurons showed evidence of inflammasome activation and downstream IL-1β processing, as previously demonstrated in acute CNS injury models, and accumulation of misfolded, hyperphosphorylated tau, and changes in expression of proteins suggestive of impaired translation in males but not in females. At 6 months, injured IL-1 receptor 1 (IL-1R1) KO mice, which are protected from postinjury cognitive deficits, had decreased accumulation of pro-IL-1β and misfolded tau in cortex and cerebellum, suggesting that IL-1R1 is upstream of inflammasome priming (defined as increase in pro-IL-1β) and abnormal tau phosphorylation. Together, our findings provide evidence for neuronal inflammasome activation and impaired proteostasis as key mechanisms linking repetitive mTBI in adolescence to later life neurologic dysfunction and neurodegeneration.SIGNIFICANCE STATEMENT Repetitive mild closed head injury in adolescent male mice leads to impaired proteostasis, tau phosphorylation, and inflammasome activation in neurons later in adulthood through mechanisms involving IL-1 receptor 1. The data are the first to link repetitive mild traumatic brain injury in adolescence to neurodegeneration and suggest molecular targets and pathways to prevent neurologic sequelae in the chronic period after injuries.

Cortical Dysplasia in Rats Provokes Neurovascular Alterations, GLUT1 Dysfunction, and Metabolic Disturbances That Are Sustained Post-Seizure Induction.

Focal cortical dysplasia (FCD) is associated with blood-brain barrier (BBB) dysfunction in patients with difficult-to-treat epilepsy. However, the underlying cellular and molecular factors in cortical dysplasia (CD) associated with progressive neurovascular challenges during the pro-epileptic phase, post-seizure, and during epileptogenesis remain unclear. We studied the BBB function in a rat model of congenital (in utero radiation-induced, first hit) CD and longitudinally examined the cortical brain tissues at baseline and the progressive neurovascular alterations, glucose transporter-1 (GLUT1) expression, and glucose metabolic activity at 2, 15, and 30 days following a second hit using pentylenetetrazole-induced seizure. Our study revealed through immunoblotting, immunohistochemistry, and biochemical analysis that (1) altered vascular density and prolongation of BBB albumin leakages in CD rats continued through 30 dayspost-seizure; (2) CD brain tissues showed elevated matrix metalloproteinase-9 levels at 2 days post-seizure and microglial overactivation through 30 days post-seizure; (3) BBB tight junction protein and GLUT1 levels were decreased and neuronal monocarboxylate transporter-2 (MCT2) and mammalian target of rapamycin (mTOR) levels were increased in the CD rat brain: (4) ATPase activity is elevated and a low glucose/high lactate imbalance exists in CD rats; and (5) the mTOR pathway is activated and MCT2 levels are elevated in the presence of high lactate during glucose starvation in vitro. Together, this study suggests that BBB dysfunction, including decreased GLUT1 expression and metabolic disturbance, may contribute to epileptogenesis in this CD rat model through multiple mechanisms that could be translated to FCD therapy in medically refractory epilepsy.

Protective effect of aqueous and ethanolic extracts of Lippia citriodora Kunth. on acrylamide-induced neurotoxicity.

Objective:Acrylamide (ACR) neurotoxicity is induced by different mechanisms such as oxidative stress and apoptosis. Scientific researchs have indicated the antioxidative properties of Lippia citriodora. The protective effect of L. citriodora aqueous and ethanolic extracts on ACR-induced neurotoxicity was investigated.Materials and methods:Male Wistar rats were randomly divided into 13 groups: (1) control, (2) ACR (50 mg/kg, i.p.), (3-6) ACR+aqueous extract (12.5, 25, 50, and 100 mg/kg, i.p.), (7-10) ACR+ethanolic extract (12.5, 25, 50, and 100 mg/kg, i.p.), (11) aqueous extract (100 mg/kg), (12) ethanolic extract (100 mg/kg), and (13) ACR+Vitamin E (200 mg/kg, every other day, i.p.). After 11 days, gait score, MDA, and GSH levels in brain cortical tissue were measured. In the in vitro test, the viability of PC12 cells (using MTT test), the amount of reactive oxygen species (ROS; using DCFH-DA method), and the protein levels of Bax, Bcl2 and caspase 3 (by western blotting) were measured. Results:In the in vitro study, the IC50 for the treatment of PC 12 cells with ACR after 24 hr was 6 mM. ACR decreased cell viability, but increased ROS level, Bax/Bcl-2 ratio, and caspase-3 protein level. Pre-treatment by L. citriodora extracts (15-120 µg/ml) ameliorated the toxic effects of ACR on PC12 cells. In the in vivo experiment, ACR-induced movement disorders increased MDA but decreased GSH content. The extracts of L. citriodora improved ACR toxic effects. Conclusion:Aqueous and ethanolic extracts of L. citriodora were found to reduce ACR-induced neurotoxicity via inhibiting oxidative stress and apoptosis.

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COVER ILLUSTRATION Seeing the invisible – Intraoperative Optical Imaging (IOI) is a technique that gives us unique insights on the human brain and it's function. The method uses subtle changes in the optical properties of the exposed cortical brain tissue to detect and visualize hemodynamic changes which are connected to specific brain functions. Since IOI needs beside an undisturbed view on the cortical surface only a camera system and light within the visible range it is well suited for application during neurosurgical interventions. In our paper “Characterization of Cortical Hemodynamic Changes Following Sensory, Visual, and Speech Activation by Intraoperative Optical Imaging Utilizing Phase-Based Evaluation Methods” we used IOI to characterize the hemodynamic changes on the brain surface following different types of stimulation in patients that underwent brain tumor surgery. The graphics illustrates the several aspects of the methodology and the results of our paper. The three images in the middle are showing phase angle maps, overlaid to the white light image of the exposed brain surfaces. The different maps are representing the response towards visual (left), sensory (right), and speech activation (middle) as shown in the result section of the paper and supported by the text above the whole image composition. The grid in the front is a polar histogram, we used for quantitative analysis and visualization of the results. Overlaid to the grid is the colorbar that we used for the phase angle maps, shaped as vortex. The colors correspond with different phase angles of the polar histogram. To underline the fact that IOI can be used to visualize functional brain areas during neurosurgical interventions, we added in the back an abstract painting of a neurosurgeon, looking through the sterile draped surgical microscope. The light source references to the fact, that IOI is an optical technology and therefore based on light.

Losartan Mitigates Oxidative Stress in the Brains of Aged IL10-/- Mice

Abstract Chronic inflammation has been linked to frailty and declined cognition in older adults. Activation of the renin-angiotensin system (RAS) through the angiotensin Type1 receptor (AT1R) has been suggested as a contributory factor that links both inflammation and aging. Here we examined the impact of 4 weeks of oral Losartan treatment on IL10-/- mice brains, a mouse model of chronic inflammation and frailty. Frontal cortex, cerebellar, and hippocampal tissue of aged (100 weeks old) male IL10-/- mice were studied. Western blot techniques were employed to quantify changes in brain AT1R, nitrotyrosine (NT) as an oxidative stress marker, and Tau proteins. Our data show that aged IL-10 mice have significantly higher levels of AT1R in the cortex tissue but not in cerebellar or hippocampal tissue compared to age and sex-matched WT mice (0.63 + 0.35 vs 1.5 + 0.54, WT vs IL10, respectively, P&amp;lt;0.004). When treated with LOS, brain cortical tissue of IL10 -/- mice showed significant decreases in levels of AT1R (1.5 + 0.54 vs 0.98 + 0.50, IL10 vs LOS treated IL10, respectively, P&amp;lt;0.04), NT (0.72 + 0.12 vs 0.42 + 0.10, IL10 vs LOS treated IL10, respectively, P&amp;lt;0.009), and Tau protein (1.3 + 0.31 vs 0.15 + 0.08, IL10 vs LOS treated IL10, respectively, P&amp;lt;0.004) as compared to control IL10-/- mice. Losartan treatment had no significant effect on hippocampal AT1R or NT levels. Our results highlight the impact of Losartan, a drug commonly prescribed for the treatment of high blood pressure, on the brain-specific angiotensin system and its downstream effects on brain oxidative stress and Tau pathology.