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Iron accumulation in the brain causes oxidative stress, blood-brain barrier (BBB) breakdown, and neurodegeneration. We examined the preventive effects of acetylated oligopeptides (AOP) from whey protein on iron-induced hippocampal damage compared to N-acetyl cysteine (NAC). This 5-week study used 40 male albino rats. At the start, all rats received 150 mg/kg/day of oral NAC for a week. The 40 animals were then randomly divided into four groups: Group I (control) received a normal diet; Group II (iron overload) received 60 mg/kg/day intraperitoneal iron dextran 5 days a week for 4 weeks; Group III (NAC group) received 150 mg/kg/day NAC and iron dextran; and Group IV (AOP group) received 150 mg/kg/day AOP and iron dextran. Enzyme-linked immunosorbent assay, spectrophotometry, and qRT-PCR were used to measure MMP-9, tissue inhibitor metalloproteinase-1 (TIMP-1), MDA, reduced glutathione (GSH) levels, and nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) gene expression. Histopathological and immunohistochemical detection of nestin, claudin, caspase, and GFAP was also done. MMP-9, TIMP-1, MDA, caspase, and GFAP rose in the iron overload group, while GSH, Nrf2, HO-1, nestin, and claudin decreased. The NAC and AOP administrations improved iron overload-induced biochemical and histological alterations. We found that AOP and NAC can protect the brain hippocampus from iron overload, improve BBB disruption, and provide neuroprotection with mostly no significant difference from healthy controls., (© 2024 John Wiley & Sons Ltd.)

Stress is a condition affecting different body systems. Curcumin (CUR) is a natural compound that has various pharmacological benefits. However, its poor oral bioavailability limits its therapeutic value. This study aimed to formulating curcumin loaded chitosan nanoparticles (CS.CUR.NPs) and investigate its gastroprotective and neuroprotective effects in rats subjected to cold restraint stress (CRS), in reference to conventional oral CUR preparation, and explore its underlying mechanism. Treated groups received either CUR or CS.CUR.NPs (100 mg∕kg) orally for 14 days before exposure to CRS. CRS elicited marked behavioral changes and gastric ulcer accompanied by histopathological abnormalities of the brain and stomach along with elevation of pain score. CUR and CS.CUR.NPs improved stress-induced gastric ulcer, cognitive performance, and pain sensation. Mechanistically, CRS disrupts oxidative and inflammatory status of the brain as manifested by high malondialdehyde and IL-6 and low total antioxidant capacity and IL-10, along with high C-reactive protein level. CRS decreased nuclear factor erythroid 2-related factor2 (Nrf2) and increased nuclear factor-kappa B (NF-κB) expressions. Furthermore, brain levels of unphosphorylated signal transducer and activator of transcription3 (U-STAT3) and glial fibrillary acidic protein (GFAP) were upregulated with stress. CUR and CS.CUR.NPs provided beneficial effects against harmful consequences resulting from stress with superior beneficial effects reported with CS.CUR.NPs. In conclusion, these findings shed light on the neuroprotective effect of CUR and CS.CUR.NPs against stress-induced neurobehavioral and neurochemical deficits and protection against stress-associated gastric ulcer. Moreover, we explored a potential crosslink between neuroinflammation, U-STAT3, NF-κB, and GFAP in brain dysfunction resulted from CRS., (Copyright © 2022 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Several lines of evidence have strongly implicated neuroinflammation in Parkinson's disease (PD) progression and l-dopa-induced dyskinesia. The present study investigated whether early subchronic pretreatment with the serotonin 5-HT 1A/1B receptor agonist eltoprazine plus the adenosine A 2A receptor antagonist preladenant counteracted l-dopa-induced abnormal involuntary movements (AIMs, index of dyskinesia), and neuroinflammation, in unilateral 6-hydroxydopamine(6-OHDA)-lesioned rat model of PD. The immunoreactivity of glial fibrillary acidic protein (GFAP), and the colocalization of ionized calcium binding adaptor molecule-1 (IBA-1), with interleukin (IL)-1β, tumor-necrosis-factor-α (TNF-α) and IL-10 were evaluated in the denervated caudate-putamen (CPu) and substantia nigra pars-compacta (SNc). The combined subchronic pretreatment with l-dopa plus eltoprazine and preladenant reduced AIMs induced by acute l-dopa challenge in these rats and decreased GFAP and IBA-1 immunoreactivity induced by the drug in both CPu and SNc, with reduction in IL-1β in IBA-1-positive cells in both CPu and SNc, and in TNF-α in IBA-1-positive cells in SNc. Moreover, a significant increase in IL-10 in IBA-1-positive cells was observed in SNc. Evaluation of immediate early-gene zif-268 (index of neuronal activation) after l-dopa challenge, showed an increase in its expression in denervated CPu of rats pretreated with l-dopa or l-dopa plus preladenant compared with vehicle, whereas rats pretreated with eltoprazine, with or without preladenant, had lower zif-268 expression. Finally, tyrosine hydroxylase and dopamine transporter examined to evaluate neurodegeneration, showed a significant equal decrease in all experimental groups. The present findings suggest that combination of l-dopa with eltoprazine and preladenant may be promising therapeutic strategy for delaying the onset of dyskinesia, preserving l-dopa efficacy and reducing neuroinflammation markers in nigrostriatal system of 6-OHDA-lesioned rats., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The current study investigated the role of epigenetic dysregulation of brain derived neurotrophic factor ( BDNF ) and glial fibrillary acidic protein ( GFAP ) genes and oxidative stress as possible mechanisms of autistic-like behaviors in neonatal isolation model in rats and the impact of folic acid administration on these parameters. Forty Wistar albino pups were used as follows: control, folic acid administered, isolated, and isolated folic acid treated groups. Isolated pups were separated from their mothers for 90 min daily from postnatal day (PND) 1 to 11. Pups (isolated or control) received either the vehicle or folic acid (4 mg/kg/day) orally from PND 1 to 29. Behavioral tests were done from PND 30 to 35. Oxidative stress markers and antioxidant defense in the frontal cortex homogenate were determined. DNA methylation of BDNF and GFAP genes was determined by qPCR. Histopathological examination was carried out. Neonatal isolation produced autistic-like behaviors that were associated with BDNF and GFAP hypomethylation, increased oxidative stress, increased inflammatory cell infiltration, and structural changes in the frontal cortex. Folic acid administration concurrently with isolation reduced neonatal isolation-induced autistic-like behaviors, decreased oxidative stress, regained BDNF and GFAP gene methylation, and ameliorated structural changes in the frontal cortices of isolated folic acid treated rats. Novelty: Neonatal isolation induces "autistic-like" behavior and these behaviors are reversed by folic acid supplementation. Neonatal isolation induces DNA hypomethylation of BDNF and GFAP , increased oxidative stress markers, and neuroinflammation. All of these changes were reversed by daily folic acid supplementation.

Background: Alexander disease (AxD) is a rare neurodegenerative disorder that is caused by dominant mutations in the gene encoding glial fibrillary acidic protein (GFAP), an intermediate filament that is primarily expressed by astrocytes. In AxD, mutant GFAP in combination with increased GFAP expression result in astrocyte dysfunction and the accumulation of Rosenthal fibers. A neuroinflammatory environment consisting primarily of macrophage lineage cells has been observed in AxD patients and mouse models., Methods: To examine if macrophage lineage cells could serve as a therapeutic target in AxD, GFAP knock-in mutant AxD model mice were treated with a colony-stimulating factor 1 receptor (CSF1R) inhibitor, pexidartinib. The effects of pexidartinib treatment on disease phenotypes were assessed., Results: In AxD model mice, pexidartinib administration depleted macrophages in the CNS and caused elevation of GFAP transcript and protein levels with minimal impacts on other phenotypes including body weight, stress response activation, chemokine/cytokine expression, and T cell infiltration., Conclusions: Together, these results highlight the complicated role that macrophages can play in neurological diseases and do not support the use of pexidartinib as a therapy for AxD.

Currently, no pharmacotherapy has been proven effective in treating photoreceptor degeneration in patients. Discovering readily available and safe neuroprotectants is therefore highly sought after. Here, we investigated nicotinamide mononucleotide (NMN), a precursor of nicotinamide adenine dinucleotide (NAD + ), in a retinal detachment (RD) induced photoreceptor degeneration. NMN administration after RD resulted in a significant reduction of TUNEL + photoreceptors, CD11b + macrophages, and GFAP labeled glial activation; a normalization of protein carbonyl content (PCC), and a preservation of the outer nuclear layer (ONL) thickness. NMN administration significantly increased NAD + levels, SIRT1 protein expression, and heme oxygenase-1 (HO-1) expression. Delayed NMN administration still exerted protective effects after RD. Mechanistic in vitro studies using 661W cells revealed a SIRT1/HO-1 signaling as a downstream effector of NMN-mediated protection under oxidative stress and LPS stimulation. In conclusion, NMN administration exerts neuroprotective effects on photoreceptors after RD and oxidative injury, suggesting a therapeutic avenue to treating photoreceptor degeneration.

Repeated exposure to mild TBI (mTBI) has been linked to an increased risk of Alzheimer's disease (AD), chronic traumatic encephalopathy (CTE) and other neurodegenerative diseases. Some pathological features typically observed in AD have been found in postmortem brains of TBI and CTE, hence treatments tested for AD have a potential to be effective against r-mTBI outcomes. Neuroinflammation may present a possible answer due to its central role both in acute brain injury and in chronic degenerative-like disorders. Our previous studies have shown that drug nilvadipine, acting as an inhibitor of spleen tyrosine kinase (SYK), is effective at reducing inflammation, tau hyperphosphorylation and amyloid production in AD mouse models. To demonstrate the effect of nilvadipine in the absence of age-related variables, we introduced the same treatment to young r-mTBI mice. We further investigate therapeutic mechanisms of nilvadipine using its racemic properties. Both enantiomers, (+)-nilvadipine and (-)-nilvadipine, can lower SYK activity, whereas (+)-nilvadipine is also a potent L-type calcium channel blocker (CCB) and shown to be anti-hypertensive. All r-mTBI mice exhibited increased neuroinflammation and impaired cognitive performance and motor functions. Treatment with racemic nilvadipine mitigated the TBI-induced inflammatory response and significantly improved spatial memory, whereas (-)-enantiomer decreased microgliosis and improved spatial memory but failed to reduce the astroglial response to as much as the racemate. These results suggest the therapeutic potential of SYK inhibition that is enhanced when combined with the CCB effect, which indicate a therapeutic advantage of multi-action drugs for r-mTBI.

Background: Postoperative neurocognitive disorders may arise in part from adverse effects of general anaesthetics on the CNS, especially in older patients or individuals otherwise vulnerable to neurotoxicity because of systemic disease or the presence of pre-existing neuropathology. Previous studies have documented cytokine and injury biomarker responses to surgical procedures that included general anaesthesia, but it is not clear to what degree anaesthetics contribute to these responses., Methods: We performed a prospective cohort study of 59 healthy volunteers aged 40-80 yr who did not undergo surgery. Plasma markers of neurological injury and inflammation were measured immediately before and 5 h after induction of general anaesthesia with 1 minimum alveolar concentration of sevoflurane. Biomarkers included interleukin-6 (IL-6), tumour necrosis factor alpha (TNF-α), C-reactive protein (CRP), and neural injury (tau, neurofilament light [NF-L], and glial fibrillary acidic protein [GFAP])., Results: Baseline biomarkers were in the normal range, although NF-L and GFAP were elevated as a function of age. At 5 h after induction of anaesthesia, plasma tau, NF-L, and GFAP were significantly decreased relative to baseline. Plasma IL-6 was significantly increased after anaesthesia, but by a biologically insignificant degree (<1 pg ml -1 ); plasma TNF-α and CRP were unchanged., Conclusions: Sevoflurane general anaesthesia without surgery, even in older adults, did not provoke an inflammatory state or neuronal injury at a concentration that is detectable by an acute elevation of measured plasma biomarkers in the early hours after exposure., Clinical Trial Registration: NCT02275026., (Copyright © 2020 British Journal of Anaesthesia. Published by Elsevier Ltd. All rights reserved.)

Previous studies have suggested that thrombospondin-1 (TSP-1) regulates the transforming growth factor beta 1 (TGF-β1)/phosphorylated Smad2/3 (pSmad2/3) pathway. Moreover, TSP-1 is closely associated with epilepsy. However, the role of the TSP-1-regulated TGF-β1/pSmad2/3 pathway in seizures remains unclear. In this study, changes in this pathway were assessed following kainic acid (KA)-induced status epilepticus (SE) in rats. The results showed that increases in the TSP-1/TGF-β1/pSmad2/3 levels spatially and temporally matched the increases in glial fibrillary acidic protein (GFAP)/chondroitin sulfate (CS56) levels following KA administration. Inhibition of TSP-1 expression by small interfering RNA or inhibition of TGF-β1 activation with a Leu-Ser-Lys-Leu peptide significantly reduced the severity of KA-induced acute seizures. These anti-seizure effects were accompanied by decreased GFAP/CS56 expression and Smad2/3 phosphorylation. Moreover, inhibiting Smad2/3 phosphorylation with ponatinib or SIS3 also significantly reduced seizure severity, alongside reducing GFAP/CS56 immunoreactivity. These results suggest that the TSP-1-regulated TGF-β1/pSmad2/3 pathway plays a key role in KA-induced SE and astrogliosis, and that inhibiting this pathway may be a potential anti-seizure strategy.

As the elderly population grows, chronic metabolic dysfunction including obesity and diabetes are becoming increasingly common comorbidities. Hypothalamic inflammation through CNS resident microglia serves as a common pathway between developing obesity and developing systemic aging pathologies. Despite understanding aging as a life-long process involving interactions between individuals and their environment, limited studies address the dynamics of environment interactions with aging or aging therapeutics. We previously demonstrated environmental enrichment (EE) is an effective model for studying improved metabolic health and overall healthspan in mice, which acts through a brain-fat axis. Here we investigated the CSF1R inhibitor PLX5622 (PLX), which depletes microglia, and its effects on metabolic decline in aging in interaction with EE. PLX in combination with EE substantially improved metabolic outcomes in middle-aged female mice over PLX or EE alone. Chronic PLX treatment depleted 75% of microglia from the hypothalamus and reduced markers of inflammation without affecting brain-derived neurotrophic factor levels induced by EE. Adipose tissue remodeling and adipose tissue macrophage modulation were observed in response to CSF1R inhibition, which may contribute to the combined benefits seen in EE with PLX. Our study suggests benefits exist from combined drug and lifestyle interventions in aged animals.

Epilepsy is a debilitating neurological illness that affects all aspect of an individual life. Despite advancement in research there is little reduction in the incidence of this disease. Prolonged febrile seizure (PFS) has been linked to epilepsy however, the pathophysiology of this is still not clear. We therefore looked at the effect of PFS on the development of epilepsy in a pentylenetetrazole (PTZ) rat model of epilepsy. A total of 42 male Sprague-Dawley rats were used for the experiment. On post-natal day (PND) 14, PFS was induced in 14 rats. This was followed by the induction of epilepsy in the 14 PFS animal and 14 animals from the remaining 28 rats by an initial injection of PTZ at a dose of 60 mg/kg on day one followed by 35 mg/kg on alternate day until kindle. We looked at the effect of PFS on the onset and the stage of convulsion at kindle. We also observed it effect on the hippocampal glial fibrillary acidic protein (GFAP), synaptophysin and metabotropic glutamate receptor 3 (mGluR3) expression measured with immunofluorescence, LI Cor Tissue florescence and immunohistochemistry respectively. Our study showed that PFS reduced seizure threshold by decreasing the time it took animals to kindle and also increased the stage of convulsion. The hippocampal GFAP, synaptophysin and mGluR3 expressions where upregulated in PTZ rats with PFS history when compared to PTZ rats alone.These findings indicated that PFS may increase the severity of epilepsy and alter brain expression of GFAP, synaptophysin and mGluR3 proteins., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Clinical and experimental studies have demonstrated the neurotoxic and behavioural effects of cadmium. However, the exact pathophysiological mechanism(s) of cadmium neurotoxicity on the human central nervous system (CNS) is not completely understood. A rat blood-brain barrier (BBB) endothelial marker, the endothelial barrier antigen (EBA), has been identified and we have shown previously that an anti-EBA IgG1 antibody exclusively recognizes barrier-competent microvessels in the rat CNS and peripheral nervous system (PNS). Endothelial cells of peripheral tissues or brain regions possessing fenestrated microvascular endothelia do not display immunoreactivity for EBA. Here, we describe the application of sequential indirect immunofluorescence with anti-EBA, and an antibody directed against glial fibrillary acidic protein (GFAP), to evaluate the immunoreactivity patterns and morphological alterations in BBB microvessels and astrocytes, following a single, high dose of cadmium in normal, term-delivered young rats. We detected a moderate reduction in immunoreactivity and number of microvessels labelled by the anti-EBA in the forebrain, cerebellum and midbrain in cadmium-exposed rats compared with normal controls. We observed weakly GFAP-reactive astrocytes displaying cell bodies with ill-defined borders and blurred cytoplasm within the white and grey matter of cadmium-exposed brains. The astrocyte nuclei were markedly enlarged, intensely hyperchromatic and exhibited chromatin condensation with nuclear fragmentation. This study indicates for the first time that EBA is involved in, and could serve as a potentially useful marker for studying, cadmium neurotoxicity in the rat model system., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Cancer and its treatment are associated with neurotoxic side effects, including cognitive dysfunction, altered functional connectivity in the brain and structural abnormalities in white matter. There is evidence that cancer and its treatment can accelerate aging. Tau is a microtubule associated protein that contributes to microtubule stability thereby playing a key role in neuronal function. Clustering of tau is commonly observed in the aged brain and is related to cognitive decline. We hypothesized that chemotherapy-induced cognitive impairment is associated with accelerated development of tau clustering in the brain as a sign of accelerated aging. We show for the first time that treatment of adult (7-8 month-old) male C57BL/6 mice with cisplatin results in reduced cognitive function and a marked increase in the number of large endogenous tau clusters in the hippocampus when assessed 4 months later. In contrast, we detected only few small tau clusters in the hippocampus of age-matched 11-12 month-old control mice. Astrocyte GFAP expression was increased in close vicinity to the tau clusters in cisplatin-treated mice. We did not detect changes in the microglial marker Iba-1 in the brain of mice treated with cisplatin. The accelerated formation of Tau-1 clusters in cisplatin-treated mice was associated with a decrease in the levels of the post-synaptic marker PSD95 and of the presynaptic marker synaptophysin in the hippocampus. We demonstrate here for the first time that chemotherapy markedly accelerates development of signs of tauopathy and loss of synaptic integrity in the hippocampus. These findings provide a mechanistic link between chemotherapy cognitive decline and accelerated aging in cancer survivors., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: Inflammation and oxidative stress are implicated in pathogenesis of spinal cord injury (SCI). Trehalose, a nonreducing disaccharide, exhibits anti-inflammatory and antioxidant effects. The present study investigated the therapeutic efficacy of trehalose in the SCI model., Design and Setting: An experimental study was designed using 120 male Wistar rats which were randomly divided into three groups including SCI, SCI + phosphate buffer saline (vehicle) and SCI + trehalose. All rats were subjected to SCI. Immediately after SCI, vehicle and trehalose groups received intrathecal injection of buffer and trehalose, respectively., Outcome Measures: The level of tissue TNFα, IL-1β, nitric oxide, malondialdehyde, myeloperoxidase, glial fibrillary acidic protein (GFAP) as well as hindlimb function were assessed at 4 hours, 1, 3 and 7 days post-SCI., Results: Data indicated an early significant decrease in inflammatory and oxidative responses following SCI in trehalose treated group. Moreover, trehalose reduced GFAP expression as soon as 1-day post-trauma. Furthermore, trehalose treatment increased the score of hindlimb function., Conclusion: Our results indicated that treatment with trehalose reduces the development of secondary injury associated with SCI. This effect likely underlies improved neurological function.

Traumatic brain injury (TBI) is the leading cause of mortality and morbidity in young adults and children in the industrialized countries; however, there are presently no FDA approved therapies. TBI results in oxidative stress due to the overproduction of reactive oxygen species and overwhelming of the endogenous antioxidant mechanisms. Recently, it has been reported that antioxidants including phytochemicals have a protective role against oxidative damage and inflammation after TBI. To analyze the effects of a naturally occurring antioxidant molecule, allyl isothiocyanate (AITC), on the nuclear factor erythroid 2-related factor 2 (Nrf2) and nuclear factor kappa B (NF-κB) signaling pathways in TBI, a cryogenic injury model was induced in mice. Here, we showed that AITC administered immediately after the injury significantly decreased infarct volume and blood-brain barrier (BBB) permeability. Protein levels of proinflammatory cytokines interleukin-1β (IL1β) and interleukin-6 (IL6), glial fibrillary acidic protein (GFAP) and NF-κB were decreased, while Nrf2, growth-associated protein 43 (GAP43) and neural cell adhesion molecule levels were increased with AITC when compared with vehicle control. Our results demonstrated that the antioxidant molecule AITC, when applied immediately after TBI, provided beneficial effects on inflammatory processes while improving infarct volume and BBB permeability. Increased levels of plasticity markers, as well as an antioxidant gene regulator, Nrf2, by AITC, suggest that future studies are warranted to assess the protective activities of dietary or medicinal AITC in clinical studies.

Objectives: Alcohol dependence leads to dysregulation of the neuroimmune system, but the effects of excessive alcohol consumption on key players of the neuroimmune response after episodic binge drinking in nondependence has not been readily assessed. These studies seek to determine how the neuroimmune system within the hippocampus responds to binge-like consumption prior to dependence or evidence of brain damage., Methods: C57BL/6J mice underwent the drinking in the dark (DID) paradigm to recapitulate binge consumption. Immunohistochemical techniques were employed to determine the effects of ethanol on cytokine and astrocyte responses within the hippocampus. Astrocyte activation was also assessed using qRT-PCR., Results: Our results indicated that binge-like ethanol consumption resulted in a 3.6-fold increase in the proinflammatory cytokine interleukin (IL)-1β immunoreactivity in various regions of the hippocampus. The opposite effect was seen in the anti-inflammatory cytokine IL-10. Binge-like consumption resulted in a 67% decrease in IL-10 immunoreactivity but had no effect on IL-4 or IL-6 compared with the water-drinking control group. Moreover, astrocyte activation occurred following ethanol exposure as GFAP immunoreactivity was increased over 120% in mice that experienced 3 cycles of ethanol binges. PCR analyses indicated that the mRNA increased by almost 4-fold after one cycle of DID, but this effect did not persist in abstinence., Conclusions: Altogether, these findings suggest that binge-like ethanol drinking prior to dependence causes dysregulation to the neuroimmune system. This altered neuroimmune state may have an impact on behavior but could also result in a heightened neuroimmune response that is exacerbated from further ethanol exposure or other immune-modulating events., (© 2019 S. Karger AG, Basel.)

Aim of the Study: Astrogliosis is a key contributor for many neurological disorders involving apoptosis, neuroinflammation and subsequent neuronal death. Silibinin, a polyphenol isolated from milk thistle (Silybum marianum), has been shown to suppress the astrocyte activation in various neurodegenerative disorders and also exhibit a neuroprotective role in neuroinflammation-driven oxidative damage. The present study was designed with an aim to investigate the neuroprotective effects of Silibinin against LPS induced oxido-inflammatory cascade and astrocyte activation., Materials and Methods: We have used in-silico molecular modelling techniques to study the interaction and binding affinity of silibinin with chemokine receptors associated with neuroinflammation. We have also tested silibinin against LPS induced oxido-inflammatory cascade and astrocyte activation in C6 glia cell lines., Results: In the present study, we found that treatment with silibinin significantly attenuates LPS-oxidative-nitrosative stress in C6 astrocytoma cells. We also observed the significant inhibition of induced astrocyte activity after treatment with silibinin. Moreover, molecular modelling studies have proposed a binding pose of silibinin with binding sites of p38 MAPK, CX3CR1 and P2X4 which is an important downstream cascade involved in glia cell activation and neuroinflammation., Conclusions: Overall, the findings from the current study suggests that silibinin exhibits neuroprotective activity by attenuating oxidative damage and astrocytes activation.

The primary aim of this study was to examine the effects of valproic acid (VPA) treatment on serum glial fibrillary acidic protein (GFAP) and neurofilament light chain (NF-L) levels. To achieve this aim, we obtained serum samples from: 1) 10 Yorkshire swine subjected to controlled cortical impact traumatic brain injury (CCI TBI) + polytrauma and randomized to receive either normal saline (NS) + VPA (n = 5) or NS alone (n = 5) and 2) five additional swine subjected to CCI TBI without polytrauma and treated with VPA. GFAP and NF-L levels were measured in samples obtained from baseline until 10 days post-injury using a digital immunoassay from Quanterix Corporation. We found that elevated GFAP and NF-L levels were first detected at 2 h post-injury; and peaked at 24 h and 72 h respectively. GFAP levels returned to baseline levels by Day 10, while NF-L remained elevated at Day 10. In TBI + polytrauma swine, the magnitude and duration of biomarker elevation, quantified by the area under the biomarker-concentration-versus-time curve during the first 10 days (AUC 0-10days ), was higher in the NS group, compared with the VPA group. For GFAP, the AUC 0-10days was 45,535 (IQR: 35,741-105,711) and 22,837 (IQR: 8,082-46,627) for the NS and NS+VPA groups, respectively. For NF-L, the AUC 0-10days was 43,073 (IQR: 18,739-120,794) and 4,475 (2,868-11,157) for the NS and NS+VPA groups, respectively. Twenty-four hour GFAP and NF-L levels had the strongest correlation with lesion size and time to normalization of behavior. Accordingly, we conclude that treatment with VPA results in significantly lower serum GFAP and NF-L levels. The time-point at which GFAP and NF-L levels have the strongest correlation with outcome is 24 h post-injury.

Objectives: Erythropoietin may have neuroprotective potential after ischemia of the central nervous system. Here, we conducted a study to characterize the protective effects of erythropoietin on retinal ganglion cells and gliotic reactions in an experimentally induced oligemia model., Methods: Rats were subjected to global oligemia by bilateral common carotid artery occlusion and then received either vehicle or erythropoietin via intravitreal injection after 48 h; they were euthanized one week after the injection. The densities of retinal ganglion cells and contents of glial fibrillary acidic protein (astrocytes/Müller cells) and cluster of differentiation 68 clone ED1 (microglia/macrophages), assessed by fluorescence intensity, were evaluated in frozen retinal sections by immunofluorescence and epifluorescence microscopy., Results: Retinal ganglion cells were nearly undetectable one week after oligemia compared with the sham controls; however, these cells were partially preserved in erythropoietin-treated retinas. The contents of glial fibrillary acidic protein and cluster of differentiation 68 clone ED1, markers for reactive gliosis, were significantly higher in retinas after bilateral common carotid artery occlusion than those in both sham and erythropoietin-treated retinas., Conclusions: The number of partially preserved retinal ganglion cells in the erythropoietin-treated group suggests that erythropoietin exerts a neuroprotective effect on oligemic/ischemic retinas. This effect could be related to the down-modulation of glial reactivity, usually observed in hypoxic conditions, clinically observed during glaucoma or retinal artery occlusion conditions. Therefore, glial reactivity may enhance neurodegeneration in hypoxic conditions, like normal-tension glaucoma and retinal ischemia, and erythropoietin is thus a candidate to be clinically applied after the detection of decreased retinal blood flow.

Background: Autoantibodies to apolipoprotein A-1 (anti-ApoA-1 IgG) were shown to predict major adverse cardiovascular events and promote atherogenesis. However, their potential relationship with clinical disability and ischaemic lesion volume after acute ischaemic stroke (AIS) remains unexplored., Materials and Methods: We included n = 76 patients admitted for AIS and we investigated whether baseline serum anti-ApoA-1 IgG levels could predict (i) AIS-induced clinical disability [assessed by the modified Rankin Scale (mRS)], and (ii) AIS-related ischaemic lesion volume [assessed by Computed Tomography (CT)]. We also evaluated the possible pro-apoptotic and pro-necrotic effects of anti-ApoA-1 IgG on human astrocytoma cell line (U251) using flow cytometry., Results: High levels of anti-ApoA-1 IgG were retrieved in 15·8% (12/76) of patients. Increased baseline levels of anti-ApoA-1 IgG were independently correlated with worse mRS [β = 0·364; P = 0·002; adjusted odds ratio (OR): 1·05 (95% CI 1·01-1·09); P = 0·017] and CT-assessed ischaemic lesion volume [β = 0·333; P < 0·001; adjusted OR: 1·06 (95% CI 1·01-1·12); P = 0·048] at 3 months. No difference in baseline clinical, biochemical and radiological characteristics was observed between patients with high vs. low levels of anti-ApoA-1 IgG. Incubating human astrocytoma cells with anti-ApoA-1 IgG dose dependently induced necrosis and apoptosis of U251 cells in vitro., Conclusion: Anti-ApoA-1 IgG serum levels at AIS onset are associated with poorer clinical recovery and worse brain lesion volume 3 months after AIS. These observations could be partly explained by the deleterious effect of anti-ApoA-1 IgG on human brain cell survival in vitro and may have clinical implication in the prediction of poor outcome in AIS., (© 2016 Stichting European Society for Clinical Investigation Journal Foundation.)

Objective: This study aimed to evaluate the effect of propentofylline administration on astrocytic response following gliotoxic injury., Method: Wistar rats were injected with ethidium bromide into the cisterna pontis and treated or not with propentofylline (12.5mg/kg/day, intraperitoneal) during the experimental period. Brainstem sections were collected from 15 to 31 days after gliotoxic injection and processed for GFAP immunohistochemistry., Results and Conclusion: Results demonstrate that propentofylline decreased astrocytic activation until the 21st day, suggesting that this drug may have a role in reducing glial scar development following injury.

Retinal neurodegeneration is an early event in the pathogenesis of diabetic retinopathy (DR). Since glucagon-like peptide 1 (GLP-1) exerts neuroprotective effects in the central nervous system and the retina is ontogenically a brain-derived tissue, the aims of the current study were as follows: 1) to examine the expression and content of GLP-1 receptor (GLP-1R) in human and db/db mice retinas; 2) to determine the retinal neuroprotective effects of systemic and topical administration (eye drops) of GLP-1R agonists in db/db mice; and 3) to examine the underlying neuroprotective mechanisms. We have found abundant expression of GLP-1R in the human retina and retinas from db/db mice. Moreover, we have demonstrated that systemic administration of a GLP-1R agonist (liraglutide) prevents retinal neurodegeneration (glial activation, neural apoptosis, and electroretinographical abnormalities). This effect can be attributed to a significant reduction of extracellular glutamate and an increase of prosurvival signaling pathways. We have found a similar neuroprotective effect using topical administration of native GLP-1 and several GLP-1R agonists (liraglutide, lixisenatide, and exenatide). Notably, this neuroprotective action was observed without any reduction in blood glucose levels. These results suggest that GLP-1R activation itself prevents retinal neurodegeneration. Our results should open up a new approach in the treatment of the early stages of DR., (© 2016 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.)

Traumatic brain injury (TBI) represents a leading cause of morbidity and mortality among young individuals. Alcohol abuse is a risk factor associated with increased TBI incidence. In addition, up to 26% of TBI patients engage in alcohol consumption after TBI. Limited preclinical studies have examined the impact of post-injury alcohol exposure on TBI recovery. The aim of this study was to determine the isolated and combined effects of TBI and alcohol on cognitive, behavioral, and physical recovery, as well as on associated neuroinflammatory changes. Male Sprague-Dawley rats (∼300g) were subjected to a mild focal TBI by lateral fluid percussion (∼30PSI, ∼25ms) under isoflurane anesthesia. On day 4 after TBI, animals were exposed to either sub-chronic intermittent alcohol vapor (95% ethanol 14h on/10h off; BAL∼200mg/dL) or room air for 10days. TBI induced neurological dysfunction reflected by an increased neurological severity score (NSS) showed progressive improvement in injured animals exposed to room air (TBI/air). In contrast, TBI animals exposed to alcohol vapor (TBI/alcohol) showed impaired NSS recovery throughout the 10-day period of alcohol exposure. Open-field exploration test revealed an increased anxiety-like behavior in TBI/alcohol group compared to TBI/air group. Additionally, alcohol-exposed animals showed decreased locomotion and impaired novel object recognition. Immunofluorescence showed enhanced reactive astrocytes, microglial activation, and HMGB1 expression localized to the injured cortex of TBI/alcohol as compared to TBI/air animals. The expression of neuroinflammatory markers showed significant positive correlation with NSS. These findings indicated a close relationship between accentuated neuroinflammation and impaired neurological recovery from post-TBI alcohol exposure. The clinical implications of long-term consequences in TBI patients exposed to alcohol during recovery warrant further investigation., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: Glial and fibrotic scars inhibit neural regeneration after spinal cord injury (SCI). N-[3,4-dimethoxycinnamoyl]-anthranilic acid (tranilast) inhibits transforming growth factor β, alleviates allergic reactions, and decreases hypertrophic skin scars. We evaluated its ability to improve motor function and inhibit the spread of tissue damage in rats with SCI., Methods: Rats with SCI were divided into groups that received tranilast (30 mg/[kg · day]) by intravenous administration (group IV), tranilast (200mg/[kg · day]) by oral administration (group OR), and saline injections (control). Motor functions were assessed by determining Basso, Beattie, and Bresnahan (BBB) scores and %grip tests for 8 weeks after SCI. Histological evaluation of ionized calcium binding adaptor molecule 1 (Iba1) at 1 week after SCI and glial fibrillary acidic protein (GFAP), fibronectin, and chondroitin sulfate (CS) at week 8 was performed., Results: Motor function recovery, BBB score, and the %grip test were significantly higher in the tranilast-treated groups than in the control group. At week 1 after SCI, inflammatory-cell invasion was more severe and Iba1 expression was significantly higher in the control group. At week 8, although the number of GFAP-positive cells increased greatly from the impaction site to the proximal and distal sites in the control group, these cells were confined around a cavity in the tranilast-treated groups. GFAP distribution coincided with that of fibronectin. Anti-CS antibody level in the tranilast-treated groups was significantly lower than that in the control group., Conclusions: Tranilast inhibits inflammation in the acute phase of SCI and reduces glial and fibrotic scars and could present a new method for treating SCI., (Copyright © 2014 Elsevier B.V. All rights reserved.)

IF (intermediate filament) proteins can be cleaved by caspases to generate proapoptotic fragments as shown for desmin. These fragments can also cause filament aggregation. The hypothesis is that disease-causing mutations in IF proteins and their subsequent characteristic histopathological aggregates could involve caspases. GFAP (glial fibrillary acidic protein), a closely related IF protein expressed mainly in astrocytes, is also a putative caspase substrate. Mutations in GFAP cause AxD (Alexander disease). The overexpression of wild-type or mutant GFAP promotes cytoplasmic aggregate formation, with caspase activation and GFAP proteolysis. In this study, we report that GFAP is cleaved specifically by caspase 6 at VELD²²⁵ in its L12 linker domain in vitro. Caspase cleavage of GFAP at Asp²²⁵ produces two major cleavage products. While the C-GFAP (C-terminal GFAP) is unable to assemble into filaments, the N-GFAP (N-terminal GFAP) forms filamentous structures that are variable in width and prone to aggregation. The effect of N-GFAP is dominant, thus affecting normal filament assembly in a way that promotes filament aggregation. Transient transfection of N-GFAP into a human astrocytoma cell line induces the formation of cytoplasmic aggregates, which also disrupt the endogenous GFAP networks. In addition, we generated a neo-epitope antibody that recognizes caspase-cleaved but not the intact GFAP. Using this antibody, we demonstrate the presence of the caspase-generated GFAP fragment in transfected cells expressing a disease-causing mutant GFAP and in two mouse models of AxD. These findings suggest that caspase-mediated GFAP proteolysis may be a common event in the context of both the GFAP mutation and excess.

The main target of neurotoxins is neurons because they comprise the main part of neural function, but glial cells may be indirect targets because they support the function of neurons. Among the glial cells, astrocytes in particular act as "nurse cells", regulating neuronal survival and functions. In the present study, to reveal whether a known neurotoxic substance, organophosphate dichlorvos (DDVP), affects the differentiation of astrocytes, we used an astrocyte differentiation model in rat glioma C6 cells. Morphological change and induction of GFAP expression in the differentiating C6 cells were suppressed by DDVP treatment. The known potential targets of DDVP are acetylcholine esterase (AChE), fatty acid amide hydrolase and methyl guanine methyl transferase. Among the specific inhibitors against these enzymes, the AChE inhibitor paraoxon successfully suppressed the cellular morphological changes and the induction of GFAP expression in differentiating C6 cells. These results indicate that DDVP inhibits differentiation in the C6 astrocyte-differentiation model, in which at least AChE inhibition is involved and that AChE is a potent regulator of the differentiation. Furthermore, considering that the main substrate of AChE is ACh, thus, ACh may act as regulators of astrocyte differentiation., (© 2013 Elsevier B.V. All rights reserved.)

Recent studies indicated that epigallocatechin gallate (EGCG) had neuroprotective effects on spinal cord injury (SCI).The current study was performed to determine the anti-edema effect of EGCG after SCI in rats. EGCG (100 mg/kg, i.p.) was administered to rats immediately following SCI. It was found that EGCG (100 mg/kg) could significantly reduce spinal cord water content. In addition, EGCG (100mg/kg) significantly reduced the expression of aquaporin-4(AQP4) and glial fibrillary acidic protein (GFAP) level at 24, 48 and 72h after injury, but it did not have this effect at 12 h after injury. The changes of AQP4 and GFAP protein induced by EGCG (100 mg/kg) treatment were accompanied by a reduction of spinal cord edema. Our results indicated that EGCG (100 mg/kg) could reduce spinal cord edema after SCI, which could be correlated with the down-regulation the expression of AQP4 and GFAP protein level after SCI., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Post-mortem histopathological studies report on reduced glial cell numbers in various frontolimbic areas of depressed patients implying that glial loss together with abnormal functioning could contribute to the pathophysiology of mood disorders. Astrocytes are regarded as the most abundant cell type in the brain and known for their housekeeping functions, but as recent developments suggest, they are also dynamic regulators of synaptogenesis, synaptic strength and stability and they control adult hippocampal neurogenesis. The primary aim of this review was to summarize the abundant experimental evidences demonstrating that antidepressant therapies have profound effect on astrocytes. Antidepressants modify astroglial physiology, morphology and by affecting gliogenesis they probably even regulate glial cell numbers. Antidepressants affect intracellular signaling pathways and gene expression of astrocytes, as well as the expression of receptors and the release of various trophic factors. We also assess the potential functional consequences of these changes on glutamate and glucose homeostasis and on synaptic communication between the neurons. We propose here a hypothesis that antidepressant treatment not only affects neurons, but also activates astrocytes, triggering them to carry out specific functions that result in the reactivation of cortical plasticity and can lead to the readjustment of abnormal neuronal networks. We argue here that these astrocyte specific changes are likely to contribute to the therapeutic effectiveness of the currently available antidepressant treatments and the better understanding of these cellular and molecular processes could help us to identify novel targets for the development of antidepressant drugs., (Copyright © 2012 Elsevier B.V. and ECNP. All rights reserved.)

Cannabinoid type 2 (CB(2)) agonists are neuroprotective and appear to play modulatory roles in neurodegenerative processes in Alzheimer's disease. We have studied the effect of 1-((3-benzyl-3-methyl-2,3-dihydro-1-benzofuran-6-yl) carbonyl) piperidine (MDA7)-a novel selective CB(2) agonist that lacks psychoactivity-on ameliorating the neuroinflammatory process, synaptic dysfunction, and cognitive impairment induced by bilateral microinjection of amyloid-β (Aβ)(1-40) fibrils into the hippocampal CA1 area of rats. In rats injected with Aβ(1-40) fibrils, compared with the administration of intraperitoneal saline for 14 days, treatment with 15 mg/kg of intraperitoneal MDA7 daily for 14 days (1) ameliorated the expression of CD11b (microglia marker) and glial fibrillary acidic protein (astrocyte marker), (2) decreased the secretion of interleukin-1β, (3) decreased the upsurge of CB(2) receptors, (4) promoted Aβ clearance, and (5) restored synaptic plasticity, cognition, and memory. Our findings suggest that MDA7 is an innovative therapeutic approach for the treatment of Alzheimer's disease., (Copyright © 2013 Elsevier Inc. All rights reserved.)

We evaluated the expression of glial fibrillary acidic protein (GFAP), glutamine synthetase (GS), ionized calcium binding adaptor protein-1 (Iba-1), and ferritin in rats after single or repeated lipopolysaccharide (LPS) treatment, which is known to induce endotoxin tolerance and glial activation. Male Wistar rats (200-250 g) received ip injections of LPS (100 µg/kg) or saline for 6 days: 6 saline (N = 5), 5 saline + 1 LPS (N = 6) and 6 LPS (N = 6). After the sixth injection, the rats were perfused and the brains were collected for immunohistochemistry. After a single LPS dose, the number of GFAP-positive cells increased in the hypothalamic arcuate nucleus (ARC; 1 LPS: 35.6 ± 1.4 vs control: 23.1 ± 2.5) and hippocampus (1 LPS: 165.0 ± 3.0 vs control: 137.5 ± 2.5), and interestingly, 6 LPS injections further increased GFAP expression in these regions (ARC = 52.5 ± 4.3; hippocampus = 182.2 ± 4.1). We found a higher GS expression only in the hippocampus of the 6 LPS injections group (56.6 ± 0.8 vs 46.7 ± 1.9). Ferritin-positive cells increased similarly in the hippocampus of rats treated with a single (49.2 ± 1.7 vs 28.1 ± 1.9) or repeated (47.6 ± 1.1 vs 28.1 ± 1.9) LPS dose. Single LPS enhanced Iba-1 in the paraventricular nucleus (PVN: 92.8 ± 4.1 vs 65.2 ± 2.2) and hippocampus (99.4 ± 4.4 vs 73.8 ± 2.1), but had no effect in the retrochiasmatic nucleus (RCA) and ARC. Interestingly, 6 LPS increased the Iba-1 expression in these hypothalamic and hippocampal regions (RCA: 57.8 ± 4.6 vs 36.6 ± 2.2; ARC: 62.4 ± 6.0 vs 37.0 ± 2.2; PVN: 100.7 ± 4.4 vs 65.2 ± 2.2; hippocampus: 123.0 ± 3.8 vs 73.8 ± 2.1). The results suggest that repeated LPS treatment stimulates the expression of glial activation markers, protecting neuronal activity during prolonged inflammatory challenges.

TRPV1 is a nonselective cation channel in nociceptors. TRPV1 stimulation has been shown to lead to the activation of microglia and astrocytes in the dorsal horn of the spinal cord. However, information on the effect of TRPV1 stimulation on glial activation in the trigeminal nucleus caudalis (TNC) is lacking. Here, we stimulated TRPV1 in the trigeminal afferents by a repetitive injection of 10 mmol/l capsaicin into the whisker pad for 2 days (d2 group), 4 days (d4 group), or 6 days (d6 group). As a control (c group), the vehicle was injected for 2 days. Anti-Iba1 and anti-glial fibrillary acidic protein antibodies were used to immunostain microglia and astrocytes in the TNC, respectively. The ratio of the cross-sectional area immunoreactive for Iba1 to the entire area of the TNC was increased in the d2 group compared with the c group on the injected side. Microglia were recruited to the superficial layers of the TNC. The numbers of microglia were reduced in the d4 group and the d6 group compared with the d2 group. The ratio of the cross-sectional area immunoreactive for glial fibrillary acidic protein to the entire TNC showed a significant increase in d2 group and the d4 group compared with the c group on the injected side. Behavioral analysis indicated that mechanical allodynia began to develop after 2 days of capsaicin treatment and persisted for at least 6 days after the onset of the repetitive capsaicin injection. These data indicate that TRPV1 stimulation activates the microglia and astrocytes in temporally distinct ways and that the development of mechanical allodynia is independent of such glial activation.

Previous studies reported that astrocyte response to amyloid-β (Aβ) before obvious neuronal damage could be detected in Alzheimer's disease (AD). It is suggested that astrocytes play a key role in AD pathologies. In this study, we investigated the effects of Aβ(42) oligomer on the proliferation and activation of astrocytes by in vitro experiments. The results showed that Aβ(42) oligomers could convert astrocytes to responsive astrocytes. It was revealed by MTT and ELISA assays that the viability of astrocytes gradually decreased, and the release of brain-derived neurotrophic factor increased with elevated Aβ(42) concentration and prolonged duration. Reverse-transcription polymerase chain reaction (RT-PCR) assay indicated that Aβ(42) oligomers increased the expression of glial fibers acid protein and interleukin-1β in a dose-dependent but not time-dependent manner. It was showed that A8, a mouse monoclonal antibody, was able to protect the cultured astrocytes against the toxicity of Aβ(42) oligomers. The result demonstrated that A8 could inhibit Aβ(42) oligomers toxic effects on astrocytes and that, alone, A8 could promote the proliferation of astrocytes in certain time. The present study laid a theoretical foundation for further understanding the effects of Aβ(42) on astrocytes and, hence, is conducive to the theoretical understanding and clinical therapies of AD progression.

Objective: The impact of present disease-modifying treatments (DMTs) in multiple sclerosis (MS) on nerve injury and reactive astrogliosis is still unclear. Therefore, we studied the effect of natalizumab treatment on the release of 2 brain-specific tissue damage markers into cerebrospinal fluid (CSF) in MS patients., Methods: CSF samples from 92 patients with relapsing forms of MS were collected in a prospective manner prior to natalizumab treatment and after 6 or 12 months. In 86 cases, natalizumab was used as second-line DMT due to breakthrough of disease activity. The levels of neurofilament light (NFL) and glial fibrillary acidic protein (GFAP) were determined using highly sensitive in-house developed enzyme-linked immunosorbent assays., Results: Natalizumab treatment led to a 3-fold reduction of NFL levels, from a mean value of 1,300 (standard deviation [SD], 2,200) to 400 (SD, 270) ng/l (p < 0.001). The later value was not significantly different from that found in healthy control subjects (350 ng/l; SD, 170; n = 28). Subgroup analysis revealed a consistent effect on NFL release, regardless of previous DMT or whether patients had relapses or were in remission within 3 months prior to natalizumab treatment. No differences between pre- and post-treatment levels of GFAP were detected., Interpretation: Our data demonstrate that natalizumab treatment reduces the accumulation of nerve injury in relapsing forms of MS. It is anticipated that highly effective anti-inflammatory treatment can reduce axonal loss, thereby preventing development of permanent neurological disability., (Copyright © 2010 American Neurological Association.)

The environmental heavy metal toxicant, lead (Pb) has been shown to be more harmful to the central nervous system (CNS) of children than to adults, given that Pb exposure affects the neural system during development. Because growth factors and cytokines play very important roles in development of the CNS, we have examined the impact of Pb exposure on the expression of cytokines during CNS development. Cytokine expression was studied in post-natal-day 21 (pnd21) mice by microarray, real-time RT-PCR, Luminex, and ELISA methodologies. BALB/c mouse pups were exposed to Pb through the dam's drinking water (0.1 mM Pb acetate), from gestation-day 8 (gd8) to pnd21. Two cytokines, interleukin-6 (IL-6) and transforming growth factor-β1 (TGF-β1), displayed significantly changed transcript levels in the presence of Pb. IL-6 and TGF-β1 both have signal transduction cascades that can cooperatively turn on the gene for the astrocyte marker glial-fibrillary acidic protein (GFAP). Microarray results indicated that Pb exposure significantly increased expression of GFAP. Pb also modulated IL-6, TGF-β1, and IL-18 protein expression in select brain regions. The deleterious effects of Pb on learning and long-term memory are posited to result from excessive astrocyte growth and/or activation with concomitant interference with neural connections. Differential neural expression of cytokines in brain regions needs to be further investigated to mechanistically associate Pb and neuroinflammation with behavioral and cognitive changes., (Copyright © 2010 Wiley Periodicals, Inc.)

Use of methamphetamine is increasingly a significant factor for the spread of human immunodeficiency virus type 1, for in certain populations, there is a convergence of methamphetamine abuse with human immunodeficiency virus type 1 infection. Methamphetamine and human immunodeficiency virus type 1 are both individually neuropathogenic, and the neuropathology caused by these two agents occurs in overlapping brain regions. However, the biological interaction of methamphetamine with lentiviruses remains unknown. Here, we investigate the effects of simultaneous exposure of these two agents on disease progression using the feline immunodeficiency virus model. The study models the bingeing methamphetamine user with sequential and repeated episodes of use, which were interrupted by periods of abstinence. Methamphetamine exposure significantly accelerated and enhanced the severity of the feline immunodeficiency virus model-induced central nervous system functional pathology, as measured in delays in brainstem auditory evoked potentials. Reciprocally, feline immunodeficiency virus enhanced the severity of the methamphetamine-induced effects on brain monoamine neurotransmitter and dopamine transporter levels. The results of this study indicate that a dual potentiation occurred. That is, methamphetamine enhanced feline immunodeficiency virus model-induced central nervous system disease and feline immunodeficiency virus model enhanced the toxic effects of methamphetamine, heralding a significant concern for those individuals that are exposed to both agents.

Purpose: Early life status epilepticus (SE) could enhance the vulnerability of the immature brain to a second SE in adulthood (two-hit seizure model). Naloxone has been proved to possess inflammation inhibitory effects in nervous system. This study was designed to evaluate the dose-dependent protective effects of naloxone in kainic acid (KA)-induced two-hit seizure model., Methods: After KA-induced SE at postnatal day 15 (P15), Sprague-Dawley rats were infused with either saline or different doses (1.92, 3.84, 5.76, and 7.68 mg/kg) of naloxone continuously for 12 h. De novo synthesis of cytokines (interleukin-1 beta [IL-1 beta], S100B) was assessed by real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) at 12 h after P15 SE. Glial activation states were analyzed by western blotting of glial markers (glial fibrillary acidic protein [GFAP], S100B, Iba1) both at 12 h after P15 SE and at P45. After a second SE at P45, cognitive deteriorations were evaluated by Morris water tests and neuron injuries were evaluated by TdT-mediated dUTP nick end labeling (TUNEL) assays., Results: Naloxone reduced IL-1 beta synthesis and microglial activation most potently at a dose of 3.84 mg/kg. Attenuation of S100B synthesis and astrocyte activation were achieved most dramatically by naloxone at a dose of 5.76 mg/kg, which is equal to the most powerful dose in ameliorating cognitive injuries and neuron apoptosis after second SE., Conclusions: Naloxone treatment immediately after early life SE could dose-dependently reduce cytokine production, glial activation, and further lower the vulnerability of immature brains to a second hit in adulthood.

Reactive astrocytosis seems to be strongly implicated in the development and maintenance of inflammatory and neurodegenerative disorders. We design a new toxic model treatment with 3-nitropropionic acid (3-NP), a mitochondrial complex II irreversible inhibitor, to induce in rats Huntington's disease (HD) like syndrome, characterized by hindlimb dystonia, involuntary choreiform movements and reduced global activity. In an attempt to find out whether molecular and morphological changes in the neuro-glial network could be involved in the pathogenesis of this disease, we developed a protocol of subchronic intra-peritoneal 3-NP intoxication. Moreover we set up specific, highly discriminative, behavioral tests to detect very early mild motor disabilities in 3-NP treated rats. This treatment did not cause severe cell death. However, in the Caudate-Putamen (CPu) of all 3-NP treated animals we found a massive astrogliosis, revealed by increased GFAP levels, paralleled by changes of the glial glutamate transporter GLAST distribution. To these glial changes we detected a transcriptional upregulation of c-fos and Sub-P in the striatal medium spiny neurons (MSN). We propose that this model of 3-NP intoxication along with the designed set of behavioral analyses allow to unmask in a very early phase the motor deficits and the underlying morpho-molecular changes associated to the onset of motor disabilities in the HD-like syndrome. Therefore this model unveil the key role played by the different components of the tripartite synapse in the pathogenesis of the HD, a putative non-cell-autonomous disease., (2009 Elsevier Ltd. All rights reserved.)

Background: In addition to caudal subnucleus caudalis (Vc) of the spinal trigeminal complex, recent studies indicate that the subnuclei interpolaris/caudalis (Vi/Vc) transition zone plays a unique role in processing deep orofacial nociceptive input. Studies also suggest that glia and inflammatory cytokines contribute to the development of persistent pain. By systematically comparing the effects of microinjection of the antiinflammatory cytokine interleukin (IL)-10 and two glial inhibitors, fluorocitrate and minocycline, we tested the hypothesis that there was a differential involvement of Vi/Vc and caudal Vc structures in deep and cutaneous orofacial pain., Results: Deep or cutaneous inflammatory hyperalgesia, assessed with von Frey filaments, was induced in rats by injecting complete Freund's adjuvant (CFA) into the masseter muscle or skin overlying the masseter, respectively. A unilateral injection of CFA into the masseter or skin induced ipsilateral hyperalgesia that started at 30 min, peaked at 1 d and lasted for 1-2 weeks. Secondary hyperalgesia on the contralateral site also developed in masseter-, but not skin-inflamed rats. Focal microinjection of IL-10 (0.006-1 ng), fluorocitrate (1 microg), and minocycline (0.1-1 microg) into the ventral Vi/Vc significantly attenuated masseter hyperalgesia bilaterally but without an effect on hyperalgesia after cutaneous inflammation. Injection of the same doses of these agents into the caudal Vc attenuated ipsilateral hyperalgesia after masseter and skin inflammation, but had no effect on contralateral hyperalgesia after masseter inflammation. Injection of CFA into the masseter produced significant increases in N-methyl-D-aspartate (NMDA) receptor NR1 serine 896 phosphorylation and glial fibrillary acidic protein (GFAP) levels, a marker of reactive astrocytes, in Vi/Vc and caudal Vc. In contrast, cutaneous inflammation only produced similar increases in the Vc., Conclusion: These results support the hypothesis that the Vi/Vc transition zone is involved in deep orofacial injury and suggest that glial inhibition and interruption of the cytokine cascade after inflammation may provide pain relief.

Soluble forms of amyloid-beta (Abeta) have been considered responsible for cognitive dysfunction prior to senile plaque formation in Alzheimer's disease (AD). As its mechanism is not well understood, we examined the effects of repeated i.c.v. infusion of soluble Alphabeta(25-35) on peptidergic system and glial cells in the pathogenesis of AD. The present study aims to investigate the protective effects of memantine on Abeta(25-35)-induced changes in peptidergic and glial systems. Infusion of Alphabeta(25-35) decreased the level of immunoreactive somatostatin (SS) and substance P (SP) in the hippocampus prior to neuronal loss or caspase activation, which is correlated with the loss of spine density and activation of inducible nitric-oxide synthase (iNOS). Biochemical experiment with peptide-degrading enzymes, prolyl oligopeptidase (POP) and endopeptidase 24.15 (EP 24.15) activities demonstrated a concomitant increase with the activation of glial marker proteins, glial fibrillary acidic protein (GFAP) and CD11b in the Abeta-treated hippocampus. Double immunostaining experiments of EP 24.15 and GFAP/CD11b antibodies clearly demonstrated the co-localization of neuro peptidases with astrocytes and microglia. Treatment with memantine, a non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist significantly attenuated Abeta(25-35)-induced changes of neuropeptides, their metabolizing enzymes, glial marker proteins, and activation of iNOS. Taken together, the data implies that memantine exerts its protective effects by modulating the neuropeptide system as a consequence of suppressing the glial cells and oxidative stress in AD model rat brain regions.

This study was to clarify whether acrylamide (ACR) will induce apoptosis and astrogliosis in an astrocytic cell line in vitro. Different time- and dose-dependent cytotoxic studies were conducted upon neuronal (SH-SY5Y) and glial cell lines (U-1240 MG) under exposure to ACR up to 72h. We showed that SH-SY5Y cells were more sensitive in cytotoxic assays than U-1240 MG cells, and significantly decreased cell viability was observed at concentrations higher than 1mM with increased lactate dehydrogenase leakage observed only at 5 and 10mM in U-1240 MG cells. The ACR-induced apoptotic responses and phosphorylation of p53 protein at Ser15 for U-1240 MG cells were identified at 48h. The increase of glial fibrillary acidic protein (GFAP) as a chemical-induced astrogliotic response was found to be associated with different ACR concentrations and exposure times, particularly at >or=48h of >or=2mM. In addition, immunocytochemical staining at 36h of 5 and 10mM treatments had significantly higher density of GFAP than the control. Thus, ACR-induced effects can be seen in neuronal and astrocytic cells. These results suggest that ACR exposure may lead to apoptotic and astrogliotic effects in human astrocytoma cells in vitro in a time- and dose-dependent manner.

Tocopherols and tocotrienols constitute the vitamin E family. Although alpha-tocotrienol is the most neuroprotective form of vitamin E proved to be effective against stroke, alpha-tocopherol is the most abundant in nature and is used most often for disease prevention/treatment. A recent metaanalysis of human studies suggested that alpha-tocopherol supplementation increases all-cause mortality. Therefore, we investigated the effects of alpha-tocopherol ( approximately 44 mg/kg body weight; equivalent to 2,600 mg/human/day) on the central nervous system (CNS) of stroke-prone spontaneously hypertensive rats (SHRSP). SHRSP treated with high dose alpha-tocopherol had significantly higher blood pressure than untreated controls fed a basal diet that contained approximately 4 mg tocopherols/kg body weight, but neither group experienced a change in degree of lipid peroxidation in serum or CNS tissue. Biochemical/immunohistochemical analyses demonstrated that expressions of phosphorylated neurofilament H protein, glial fibrillary acidic protein and cathepsin D in the CNS tissue were significantly enhanced in alpha-tocopherol-supplemented rats, whereas expressions of SOD2 and Bcl-xL were diminished in response to alpha-tocopherol supplementation. Similarly, the frequency of cathepsin D-positive cells, corresponding mostly to microglial cells, was significantly increased in alpha-tocopherol-supplemented rats. Alpha-tocopherol supplementation also increased the number of lysosomes and lipofuscin granules in perikarya of both hippocampal pyramidal and Purkinje cells. Furthermore, alpha-tocopherol supplementation increased the frequency of glial filaments and lipofuscin granules in astrocytes and lysosomes in microglial cells that were frequently occupied with phagocytosed inclusion structures. The present results are the first to suggest that a very high dose of alpha-tocopherol supplementation increases blood pressure in SHRSP rats and influences the CNS tissue in a manner that seems adverse., (2008 Wiley-Liss, Inc.)

Associated with neuronal death, profound synaptic changes occur in the spinal cord during the apoptotic process triggered after axotomy in neonatal rats. With respect to this, the major histocompatibility complex of class I (MHC class I) has recently emerged as a new mechanism related to synaptic stripping and plasticity. The present study investigated the impact of upregulating MHC class I expression by treatment with beta interferon (beta INF) on motoneuron survival, synaptic plasticity and astrogliosis after neonatal sciatic nerve injury. P2 rats were subjected to unilateral axotomy followed by three days of beta INF treatment. The results were analyzed by counting Nissl stained motoneurons, immunohistochemistry (anti-synaptophysin, MHC class I, GFAP and Iba-1) and transmission electron microscopy. INF treatment induced an increased expression of MHC class I, which resulted in a stronger synaptic elimination process in the spinal cord, as seen by the synaptophysin labeling. GFAP and Iba-1 upregulation were not significantly altered by the INF treatment, displaying the same degree of enhanced reactivity as compared to the placebo group. The ultrastructural analysis showed that, apart from the overall reduction of inputs in the neuropil, no statistical differences were present when comparing the INF and placebo treated animals. Also, neuronal survival was not altered by cytokine administration. The present results provide evidence that MHC class I upregulation after neonatal injury does not change the fate of lesioned motoneurons. In this way, the lack of neurotrophic support may cause broader synaptic loss, which superposes the more subtle effects of the upregulation of MHC class I.

Estrogens are considered neurotrophic for dopamine neurons. Parkinson's disease is more frequent in males than in females, and more prevalent in females with short reproductive life. Estrogens are neuroprotective against neurotoxic agents for dopamine neurons in vivo and in vitro. Here, we have investigated the role of estrogens in wild-type (WT) and parkin null mice (PK-/-). WT mice present sexual dimorphisms in neuroprotective mechanisms (Bcl-2/Bax, chaperones, and GSH), but some of these inter-sex differences disappear in PK-/-. Tyrosine hydroxylase (TH) protein and TH+ cells decreased earlier and more severely in female than in male PK-/- mice. Neuronal cultures from midbrain of WT and PK-/- mice were treated with estradiol from 10 min to 48 h. Short-term treatments activated the mitogen-activated protein kinase pathway of WT and PK-/- neurons and the phosphatidylinositol 3'-kinase/AKT/glycogen synthase kinase-3 pathway of WT but not of PK-/- cultures. Long-term treatments with estradiol increased the number of TH+ neurons, the TH expression, and the extension of neurites, and decreased the level of apoptosis, the expression of glial fibrillary acidic protein, and the number of microglial cells in WT but not in PK-/- cultures. The levels of estrogen receptor-alpha were elevated in midbrain cultures and in the striatum of adult PK-/- male mice, suggesting that suppression of parkin changes the estrogen receptor-alpha turnover. From our data, it appears that parkin participates in the cellular estrogen response which could be of interest in the management of parkin-related Parkinson's disease patients.

There is growing evidence indicating that reactive nitrogen species (RNS) and reactive oxygen species (ROS) are a major contributor to the pathogenesis and progression of Parkinson's disease. Here we investigated whether edaravone (free radical scavenger), minocycline (inducible nitric oxide synthase, iNOS inhibitor), 7-nitroindazole (neuronal NOS, nNOS inhibitor), fluvastatin (endothelial NOS, eNOS activator) and pitavastatin (eNOS activator) can protect against MPTP neurotoxicity in mice under the same condition. The present study showed that 7-nitroindazole could protect dose-dependently against the striatal dopamine depletions in mice 5 days after MPTP treatment. In contrast, edaravone, minocycline, fluvastatin and pitavastatin did not show the neuroprotective effect on MPTP-induced striatal dopamine depletion. Our immunohistochemical study showed that TH (tyrosine hydroxylase) and DAT (dopamine transporter) immunoreactivity was decreased significantly in the striatum and substantia nigra 5 days after MPTP treatment. The administration of 7-nitroindazole showed a protective effect against the severe reductions in levels of TH and DAT immunoreactivity in the striatum and substantia nigra 5 days after MPTP treatment. Furthermore, our Western blot analyses study showed the remarkable loss of TH protein levels in the striatum 5 days after MPTP treatment. In contrast, 7-nitroindazole prevented a significant loss in TH protein levels in the striatum 5 days after MPTP treatment. On the other hand, GFAP (glial fibrillary acidic protein) immunoreactivity increased significantly in the striatum and substantia nigra, 5 days after MPTP treatment. 7-Nitroindazole ameliorated severe increases in number of GFAP immunoreactive astrocytes in the striatum and substantia nigra 5 days after MPTP treatment. Furthermore, our Western blot analyses study showed the increase of GFAP protein levels in the striatum 5 days after MPTP treatment. 7-Nitroindazole prevented a significant increase in the GFAP protein levels in the striatum 5 days after MPTP treatment. The present results indicate that 7-nitroindazole can protect dose-dependently against the striatal dopamine depletions in mice 5 days after MPTP treatment. In contrast, edaravone, minocycline, fluvastatin and pitavastatin did not show the neuroprotective effect on MPTP-induced striatal dopamine depletions. These findings demonstrate that the overexpression of nNOS may play a major role in the neurotoxic processes of MPTP, as compared to the production of ROS, the overexpression of iNOS and the modulation of eNOS. Thus, our findings provide strong evidence for neuroprotective properties of nNOS inhibitor in this animal model of Parkinson's disease.

Background: It is well known that long-term morphine administration results in tolerance, which limits the clinical use of this drug in pain management., Methods: Male Wistar rats were randomly assigned to receive one of four different infusions: morphine [15 microg/h, intrathecal (i.t.)], saline, MK-801 (5 microg/h, i.t.) plus morphine (15 microg/h, i.t.), or MK-801 (5 microg/h, i.t.) alone., Results: Morphine infusion induced a maximal antinociceptive effect on day 1 and tolerance on day 3, and the maximal anti-receptive tolerance was observed on day 5. Co-infusing MK-801 with morphine attenuated morphine's anti-receptive tolerance. Two-dimensional gel electrophoretic analysis of spinal proteins revealed that eight protein spots were up-regulated in morphine-tolerant rats, and that they were significantly inhibited by MK-801 co-infusion. Among the up-regulated proteins, glial fibrillary acid protein (GFAP), a glial-specific maker, was identified by mass spectrometry. This finding was also confirmed by Western blot analysis., Conclusion: Using proteomic analysis, we identified eight GFAP protein spots that were up-regulated in the dorsal horn of morphine-tolerant rat spinal cords. This up-regulation was partly inhibited by N-methyl-D-aspartate receptor antagonist MK-801 co-infusion, which suggests that GFAP protein can be considered to be a pathogenesis marker of morphine tolerance.

Manganese (Mn) is a cofactor for some metalloprotein enzymes, including Mn-superoxide dismutase (Mn-SOD), a mitochondrial enzyme predominantly localized in neurons, and glutamine synthetase (GS), which is selectively expressed in astroglial cells. The detoxifying effects of GS and Mn-SOD in the brain, involve catabolizing glutamate and scavenging superoxide anions, respectively. Mn intoxication is characterized by impaired function of the basal ganglia. However, it is unclear whether regional central nervous system expression of manganoproteins is also affected. Here, we use immunocytochemistry in the adult rat brain, to examine whether Mn overload selectively affects the expression of GS, Mn-SOD, Cu/Zn-SOD, another component of the SOD family, and glial fibrillary acid protein (GFAP), a specific marker of astrocytes. After chronic Mn overload in drinking water for 13 weeks, we found that the number and immunostaining intensity of GS- and Mn-SOD-positive cells was significantly decreased in the striatum and globus pallidus, but not in the cerebral frontal cortex. In addition, we found that GS enzymatic activity was decreased in the strio-pallidal regions but not in the cerebral cortex of Mn-treated animals. In contrast, Cu/Zn-SOD- and GFAP-immunoreactivity was unchanged in both the cerebral cortex and basal ganglia of Mn-treated rats. Thus, we conclude that in response to chronic Mn overload, a down-regulation of some manganoproteins occurs in neurons and astrocytes of the striatum and globus pallidus, probably reflecting the vulnerability of these regions to Mn toxicity.

We investigated the involvement of gamma-aminobutyric acid(A) (GABA(A)) receptors in the neuroprotective effect of gamma-glutamylethylamide (theanine), a component of Japanese green tea, following a 4-h middle cerebral artery (MCA) occlusion in mice. Theanine (1 mg/kg) reduced the size of the cerebral infarct and alterations of NeuN, GFAP, and Iba 1 expression levels at 24 h after MCA occlusion. This neuroprotective effect of theanine was prevented by bicuculline (GABA(A)-receptor antagonist, 10 mg/kg) but not 3-mercaptopropionic acid (glutamate decarboxylase inhibitor). These results suggest that the neuroprotective effect of theanine is mediated, at least in part, by GABA(A) receptors.

It has previously been shown that small peptide molecules derived from the apolipoprotein E (ApoE) receptor binding region are anti-inflammatory in nature and can improve outcome following head injury. The present study evaluated the preclinical efficacy of COG1410, a small molecule ApoE-mimetic peptide (1410 daltons), following cortical contusion injury (CCI). Animals were prepared with a unilateral CCI of the sensorimotor cortex (SMC) or sham procedure. Thirty mins post-CCI the animals received i.v. infusions of 0.8 mg/kg COG1410, 0.4 mg/kg COG1410, or vehicle. Starting on day 2, the animals were tested on a battery of behavioral measures to assess sensorimotor (vibrissae-forelimb placing and forelimb use-asymmetry), and motor (tapered balance beam) performance. Administration of the 0.8 mg/kg dose of COG1410 significantly improved recovery on the vibrissae-forelimb and limb asymmetry tests. However, no facilitation was observed on the tapered beam. The low dose (0.4 mg/kg) of COG1410 did not show any significant differences compared to vehicle. Lesion analysis revealed that the 0.8 mg/kg dose of COG1410 significantly reduced the size of the injury cavity compared to the 0.4 mg/kg dose and vehicle. The 0.8 mg/kg dose also reduced the number of glial fibrillary acid protein (GFAP+) reactive cells in the injured cortex. These results suggest that a single dose of COG1410 facilitates behavioral recovery and provides neuroprotection in a dose and task-dependent manner. Thus, the continued clinical development of ApoE based therapeutics is warranted and could represent a novel strategy for the treatment of traumatic brain injuries.

Alcohol exposure during pregnancy may cause fetal alcohol syndrome (FAS), characterized by impaired cognitive functions. Neurogenesis occurs in the adult hippocampus and is functionally associated with learning, memory, and mood disorders. However, whether early postnatal exposure to alcohol impairs neurogenesis and through which mechanisms it occurs is poorly understood. Here, we report that a single episode of alcohol exposure in postnatal day 7 (P7) decreases neurogenesis in the adult hippocampus. Furthermore, we demonstrate a co-localization of glial fibrillar acidic protein, nestin, and vimentin with activated caspase-3 12 h after ethanol treatment. Finally, we show that the number of primary neurospheres derived from the hippocampi of alcohol-exposed mice is reduced compared to controls. These findings suggest that alcohol exposure in postnatal mice reduces the pool of neural stem/progenitor cells in the DG, and subsequently results in a decrease of adult neurogenesis. This may explain certain aspects of impaired hippocampal functions in FAS.

Glial scar formation around implanted silicon neural probes compromises their ability to facilitate long-term recordings. One approach to modulate the tissue reaction around implanted probes in the brain is to develop probe coatings that locally release anti-inflammatory drugs. In this study, we developed a nitrocellulose-based coating for the local delivery of the anti-inflammatory drug dexamethasone (DEX). Silicon neural probes with and without nitrocellulose-DEX coatings were implanted into rat brains, and inflammatory response was evaluated 1 week and 4 weeks post implantation. DEX coatings significantly reduced the reactivity of microglia and macrophages 1 week post implantation as evidenced by ED1 immunostaining. CS56 staining demonstrated that DEX treatment significantly reduced chondroitin sulfate proteoglycan (CSPG) expression 1 week post implantation. Both at 1-week and at 4-week time points, GFAP staining for reactive astrocytes and neurofilament (NF) staining revealed that local DEX treatment significantly attenuated astroglial response and reduced neuronal loss in the vicinity of the probes. Weak ED1, neurocan, and NG2-positive signal was detected 4 weeks post implantation for both coated and uncoated probes, suggesting a stabilization of the inflammatory response over time in this implant model. In conclusion, this study demonstrates that the nitrocellulose-DEX coating can effectively attenuate the inflammatory response to the implanted neural probes, and reduce neuronal loss in the vicinity of the coated probes. Thus anti-inflammatory probe coatings may represent a promising approach to attenuate astroglial scar and reduce neural loss around implanted neural probes.