GFAP-immunoreactive Astrocytes Research Articles

Doxorubicin-induced neurotoxicity differently affects the hippocampal formation subregions in adult mice

Doxorubicin (DOX) is an anthracycline used to treat a wide range of tumours. Despite its effectiveness, it is associated with a long range of adverse effects, of which cognitive deficits stand out. The present study aimed to assess the neurologic adverse outcome pathways of two clinically relevant cumulative doses of DOX. Adult male CD-1 mice received biweekly intraperitoneal administrations for 3 weeks until reaching cumulative doses of 9 mg/kg (DOX9) or 18 mg/kg (DOX18). Animals were euthanized one week after the last administration, and biomarkers of oxidative stress and brain metabolism were evaluated in the whole brain. Coronal sections of fixed brains were used for specific determinations of the prefrontal cortex (PFC) and hippocampal formation (HF). In the whole brain, DOX18 tended to disrupt the antioxidant defences, affecting glutathione levels and manganese superoxide dismutase expression. Considering the regional analysis, DOX18 increased the volume of all brain areas evaluated, while GFAP-immunoreactive astrocytes decreased in the dentate gyrus (DG) and increased in the CA3 region of HF, both in a dose-dependent manner. Concerning the apoptosis pathway, whereas Bax increased in the DOX9 group, it decreased in the DOX18 group. Only in the latter group did Bcl-2 levels also decrease. While p53 only increased in the CA3 region of the DOX9 group, AIF increased in the PFC and DG of DOX18. Finally, phosphorylation of Tau decreased with the highest DOX dose in DG and CA3, while TNF-α levels increased in CA1 of DOX18. Our results indicate new pathways not yet described that could be responsible for the cognitive impairments observed in treated patients.

Impact of thyroid hormone perturbations in adult mice: brain weight and blood vessel changes, gene expression variation, and neurobehavioral outcomes

Mouse models of hyper- and hypothyroidism were used to examine the effects of thyroid hormone (TH) dyshomeostasis on the aging mammalian brain. 13–14 month-old mice were treated for 4months with either levothyroxine (hyperthyroid) or a propylthiouracil and methimazole combination (PTU/Met; hypothyroid). Hyperthyroid mice performed better on Morris Water Maze than control mice, while hypothyroid mice performed worse. Brain weight was increased in thyroxine-treated, and decreased in PTU/Met-treated animals. The brain weight change was strongly correlated with circulating and tissue T4. Quantitative measurements of microvessels were compared using digital neuropathologic methods. There was an increase in microvessel area in hyperthyroid mice. Hypothyroid mice showed a trend for elevated glial fibrillary acidic protein-immunoreactive astrocytes, indicating an increase in neuroinflammation. Gene expression alterations were associated with TH perturbation and astrocyte-expressed transcripts were particularly affected. For example, expression of Gli2 and Gli3, mediators in the Sonic Hedgehog signaling pathway, were strongly impacted by both treatments. We conclude that TH perturbations produce robust neurobehavioral, pathological, and brain gene expression changes in aging mouse models.

Changes to GFAP Immunoreactive Astrocytes in Medial Prefrontal Cortex Following Exposure to Chronic Stress and Antioxidant Supplementation in Rat Model

Introduction: Astrocytes are responsible for many essential functions of neurons in CNS. It has been recognised that chronic stress affects the morphology of astrocyte. Natural antioxidant such as honey has been used as one of the therapeutic strategies to lessen the damaging effect of chronic stress on our body. Therefore, the aim of the study is to explore the effect of natural antioxidant, Tualang honey (TH) on the morphology of astrocytes following chronic stress exposure. Methods: Thirty-two male rats were randomly divided into the 4 groups: (i) control, (ii) stress, (iii) honey, (iv) stress plus honey groups.TH was administered via oral gavage at dose of 1.0 g/kg body weight pre and post experiment. Chronic stress was exposed to animals in group (ii) and (iv) for consecutive 21 days. Anti GFAP immunohistochemistry method was employed to label astrocytes in the medial prefrontal cortex. The number of GFAP+ astrocytes and several parameters related to astrocyte processes were measured. Results: The present study showed that chronic stress reduced the GFAP immunoreactive astrocyte number and percentage of GFAP immunoreactive material. Chronic stress also caused a reduction in astrocyte process ramification as indicated by a reduction in astrocyte total number of processes, average length of processes and maximum number of intersections. However, antioxidant treatment using TH could not reverse these stress-induced changes to the astrocytes. Conclusion: These results demonstrate that chronic stress decreases the number of GFAP immunoreactive astrocyte and cause shrinking of astrocyte processes in stress-sensitive brain region, but these changes cannot be reversed by antioxidant treatment.

Changes in Glial Fibrillary Acidic Protein-Immunoreactive Astrocytes in the Prefrontal Cortex of the Male Rat following Chronic Khat Use.

Long-term khat consumption is associated with significant neurocognitive changes, which have been elucidated in behavioral studies. With current research showing the centrality of astrocytes and other glial cells in neuronal signaling, there is possibility that these cells are also affected by chronic khat use. There is little literature on the structural changes in the prefrontal cortex neuronal and astrocytic cytoarchitecture and morphometry in chronic khat users. The objective of this study was to describe the changes in astrocyte morphometry and structure in rats after long-term use of khat (miraa). Adult male Wistar rats, aged 2-3 months, weighing 200-300 g were randomized into four groups of 10 each (control, Group 1, Group 2, and Group 3) to correspond with those used as controls and those that received 500 mg/kg, 1000 mg/kg, and 2000 mg/kg body weight khat extracts, respectively. Fresh khat leaves were purchased from Maua market in Meru, and crude extract was prepared using lyophilization. The control rats were fed on normal diet, while the experimental groups were fed on normal diet and khat extracts using oral gavage for 6 weeks. The animals were sacrificed and their brains were removed. We performed immunohistochemical visualization of astrocytes using glial fibrillary acidic protein. Photomicrographs of the stained sections were transferred to ImageJ Fiji software to study the astrocyte density and astrocytic processes. We used Kruskal-Wallis test to correlate the four animal groups in terms of astrocyte densities. We observed an increase in the average number of astrocytes with increasing doses of khat compared to controls, with those in Group 3 (2000 mg/kg) having an exuberant reactive astrocytosis. Further, escalating khat doses resulted in increased glial fibrillary acidic protein immunoreactivity in the nuclei and astrocytic processes, gliotic changes, and increased complexity of astrocytic processes. Chronic khat use, especially at high doses, results in reactive astrocytosis and astrogliosis, which may be part of the mechanisms involved in the cognitive changes associated with its use.

Relationship between Neuronal Damage/Death and Astrogliosis in the Cerebral Motor Cortex of Gerbil Models of Mild and Severe Ischemia and Reperfusion Injury.

Neuronal loss (death) occurs selectively in vulnerable brain regions after ischemic insults. Astrogliosis is accompanied by neuronal death. It can change the molecular expression and morphology of astrocytes following ischemic insults. However, little is known about cerebral ischemia and reperfusion injury that can variously lead to damage of astrocytes according to the degree of ischemic injury, which is related to neuronal damage/death. Thus, the purpose of this study was to examine the relationship between damage to cortical neurons and astrocytes using gerbil models of mild and severe transient forebrain ischemia induced by blocking the blood supply to the forebrain for five or 15 min. Significant ischemia tFI-induced neuronal death occurred in the deep layers (layers V and VI) of the motor cortex: neuronal death occurred earlier and more severely in gerbils with severe ischemia than in gerbils with mild ischemia. Distinct astrogliosis was detected in layers V and VI. It gradually increased with time after both ischemiae. The astrogliosis was significantly higher in severe ischemia than in mild ischemia. The ischemia-induced increase of glial fibrillary acidic protein (GFAP; a maker of astrocyte) expression in severe ischemia was significantly higher than that in mild ischemia. However, GFAP-immunoreactive astrocytes were apparently damaged two days after both ischemiae. At five days after ischemiae, astrocyte endfeet around capillary endothelial cells were severely ruptured. They were more severely ruptured by severe ischemia than by mild ischemia. However, the number of astrocytes stained with S100 was significantly higher in severe ischemia than in mild ischemia. These results indicate that the degree of astrogliosis, including the disruption (loss) of astrocyte endfeet following ischemia and reperfusion in the forebrain, might depend on the severity of ischemia and that the degree of ischemia-induced neuronal damage may be associated with the degree of astrogliosis.

Increased Prostaglandin E2 in Brainstem Respiratory Centers Is Associated With Inhibition of Breathing Movements in Fetal Sheep Exposed to Progressive Systemic Inflammation.

BackgroundPreterm newborns commonly experience apnoeas after birth and require respiratory stimulants and support. Antenatal inflammation is a common antecedent of preterm birth and inflammatory mediators, particularly prostaglandin E2 (PGE2), are associated with inhibition of vital brainstem respiratory centers. In this study, we tested the hypothesis that exposure to antenatal inflammation inhibits fetal breathing movements (FBMs) and increases inflammation and PGE2 levels in brainstem respiratory centers, cerebrospinal fluid (CSF) and blood plasma.MethodsChronically instrumented late preterm fetal sheep at 0.85 of gestation were randomly assigned to receive repeated intravenous saline (n = 8) or lipopolysaccharide (LPS) infusions (experimental day 1 = 300 ng, day 2 = 600 ng, day 3 = 1200 ng, n = 8). Fetal breathing movements were recorded throughout the experimental period. Sheep were euthanized 4 days after starting infusions for assessment of brainstem respiratory center histology.ResultsLPS infusions increased circulating and cerebrospinal fluid PGE2 levels, decreased arterial oxygen saturation, increased the partial pressure of carbon dioxide and lactate concentration, and decreased pH (p < 0.05 for all) compared to controls. LPS infusions caused transient reductions in the % of time fetuses spent breathing and the proportion of vigorous fetal breathing movements (P < 0.05 vs. control). LPS-exposure increased PGE2 expression in the RTN/pFRG (P < 0.05 vs. control) but not the pBÖTC (P < 0.07 vs. control) of the brainstem. No significant changes in gene expression were observed for PGE2 enzymes or caspase 3. LPS-exposure reduced the numbers of GFAP-immunoreactive astrocytes in the RTN/pFRG, NTS and XII of the brainstem (P < 0.05 vs. control for all) and increased microglial activation in the RTN/pFRG, preBÖTC, NTS, and XII brainstem respiratory centers (P < 0.05 vs. control for all).ConclusionChronic LPS-exposure in late preterm fetal sheep increased PGE2 levels within the brainstem, CSF and plasma, and was associated with inhibition of FBMs, astrocyte loss and microglial activation within the brainstem respiratory centers. Further studies are needed to determine whether the inflammation-induced increase in PGE2 levels plays a key role in depressing respiratory drive in the perinatal period.

Evaluation of immunoreactive for glial fibrillary acidic protein astrocytes in the periaqueductal gray matter of rats treated with monosodium glutamate.

The aim of this study was to evaluate the glial fibrillary acidic protein immunoreactive (GFAP-IR) astrocytes in the periaqueductal gray matter (PAG) of rats treated with monosodium glutamate (MSG). This study was performed on PAG sections of rats treated subcutaneously with MSG (2 g/kg b.w. (group I) and 4 g/kg b.w. (group II)) and saline (group C). Immunohistochemical reactions were performed by peroxidase-antiperoxidase method using an antibody against GFAP protein. In animals receiving MSG in dorsomedial (dm) and dorsolateral (dl) parts of PAG the majority of astrocytes had GFAP-IR bodies and few thin, poorly branched protuberances. In lateral (l) and ventrolateral (vl) parts of PAG cells with numerous, branched protuberances with GFAP expression were observed. Some astrocytes in vl PAG of group II rats were characterized by strongly stained cytoplasmic bodies and thick, multiple branched protuberances. Statistically significant differences in the number of GFAP-IR astrocytes were shown between control animals and group II in dm PAG and dl PAG. All MSG-treated animals revealed statistically significantly fewer immunonegative structures for GFAP as well as all cells in the PAG.In the PAG of rats receiving MSG, especially at high dose, there is an increase in GFAP immunoreactivity in astrocytes. The hypertrophy of GFAP-IR cells and the probable lack of their proliferation may indicate isomorphic astroglia in response to increased extracellular glutamate concentrations. However, the decrease in their number in dm PAG and dl PAG may be related to MSG cytotoxicity.

Increases in GFAP immunoreactive astrocytes in the cerebellar molecular layer of young adult CBA/J mice

BackgroundCBA/J mice are standard experimental animals in auditory studies, and age-related changes in auditory pathways are well documented. However, changes in locomotion-related brain regions have not been systematically explored.ResultsWe showed an increase in immunoreactivity for glial fibrillary acidic protein (GFAP) in the cerebellar molecular layer associated with Purkinje cells in mice at 24 weeks of age but not in the younger mice. Increased GFAP immunoreactivity appeared in the form of clusters and distributed multifocally consistent with hyperplasia of astrocytes that were occasionally associated with Purkinje cell degeneration. Three out of 12 animals at 16 and 24 weeks of age exhibited pre-convulsive clinical signs. Two of these 3 animals also showed increased GFAP immunoreactivity in the cerebellum. Rotarod behavioral assessments indicated decreased performance at 24 weeks of age.ConclusionsThese results suggest minimal to mild reactive astrocytosis likely associated with Purkinje cell degeneration in the cerebellum at 24 weeks of age in CBA/J mice. These findings should be taken into consideration prior to using this mouse strain for studying neuroinflammation or aging.

Pioglitazone rescues high-fat diet-induced depression-like phenotypes and hippocampal astrocytic deficits in mice

The prevalence of diabetes is rapidly increasing worldwide and is highly associated with the incidence of depression. Pioglitazone, a Peroxisome proliferator-activated receptor gamma (PPAR-γ) agonist, is widely used for treating patients with type 2 diabetes. However, whether pioglitazone alleviates metabolic disorder-related depression and astrocytic deficits remains unclear. Here we showed that 12 weeks of high-fat diet (HFD) feeding (from 8- to 20-week-old) induced not only obesity and insulin resistance, but also depression-like behaviors in mice. Astrocytic activation, a sign closely associated with depression, was also evident in the ventral hippocampus. Four weeks of pioglitazone (10 or 20 mg/kg, daily, from 20- to 24-week-old) treatment alleviated the HFD-induced glucose-metabolic dysfunctions, upregulation of ventral hippocampal GFAP, reduction of the total process lengths and the number of branch points of the ventral hippocampal CA1 GFAP-immunoreactive astrocytes and depressive phenotypes but had no effect on anxiety-like behaviors or hippocampus-related learning and memory in mice. These findings suggest that pioglitazone could be a potential therapeutic agent for metabolic disorders and associated depression.

Catfish Epidermal Preparation Accelerates Healing of Damaged Nerve in a Sciatic Nerve Crush Injury Rat Model.

Preliminary investigations showed that preparations from Arabian Gulf catfish (Arius bilineatus, Val) epidermal gel secretion (PCEGS) exhibit potent anti-inflammatory and healing properties as shown in our previous clinical trials for the healing of non-healing diabetic foot ulcers, chronic back pain, and some other neurological disorders. Here, we report for the first time a unique preparation containing only proteins and lipids (soluble protein fraction B, SPF-FB), derived from the PCEGS accelerated the healing and recovery of sensory-motor functions of experimental sciatic nerve crush injury in rats with its unique neuroprotective and neuroregenerative properties on the spinal neurons and peripheral nerve fibers. Male rats were randomly assigned to five groups: (I) NAÏVE, (II) SHAM, (III) CRUSH treated with saline, (IV) CRUSH + SPF-FB treated with 3 mg/kg intraperitoneally (IP) and (V) CRUSH + SPF-FB treated with 6 mg/kg subcutaneously (SC) groups. The crush groups III, IV and V underwent sciatic nerve crush injury, followed by treatment daily for 14 days with saline, SPF-FB IP and SPF-FB SC. All animals were tested for the neurobehavioral parameters throughout the 6 weeks of the study. Sciatic nerve and spinal cord tissues were processed for light and electron histological examinations, stereological analysis, immunohistochemical and biochemical examinations at Week 4 and Week 6 post-injury. Administration of SPF-FB IP or SC significantly enhanced the neurobehavioral sensory and motor performance and histomorphological neuroregeneration of the sciatic nerve-injured rats. The stereological evaluation of the axon area, average axon perimeters, and myelin thickness revealed significant histomorphological evidence of neuroregeneration in the FB-treated sciatic nerve crush injured groups compared to controls at 4 and 6 weeks. SPF-FB treatment significantly prevented the increased in NeuN-immunoreactive neurons, increased GFAP immunoreactive astrocytes, and decreased GAP-43. We conclude that SPF-FB treatment lessens neurobehavioral deficits, enhances axonal regeneration following nerve injury. We conclude that SPF-FB treatment lessens neurobehavioral deficits and enhances axonal regeneration following nerve injury, as well as protects spinal neurons and enhances subcellular recovery by increasing astrocytic activity and decreasing GAP-43 expression.

Electroacupuncture alleviated the depression-like behavior by regulating FGF2 and astrocytes in the hippocampus of rats with chronic unpredictable mild stress

Studies have shown that basic fibroblast growth factor (FGF2) is a neurotrophic factor associated with depression. Electroacupuncture (EA) has been shown to be an effective treatment for depression. In the current study, we observed the effects of EA on hippocampal FGF2 and astrocytes, and further investigated the mechanism underlying antidepressant effect of EA. The chronic unpredictable mild stress (CUMS) method were selected to induce depressive-like behaviors of rats. Paroxetine is a commonly used antidepressant and was used as a positive control drug in this experiment. The male adult Sprague Dawley (SD) rats were randomized to four experimental groups (normal control group, CUMS group, EA group and paroxetine group, n = 10/group). EA intervention was administered once daily for 14 days at acupuncture points Baihui (GV20) and Yintang (GV29). Rats in the paroxetine group received daily paroxetine administered intragastrical. Behavioral test, immunohistochemistry (IHC), western blot (WB) and quantitative real-time PCR (qPCR) were conducted to evaluate the intervene effect and the changes of FGF2 and astrocyte marker (glial fibrillary acidic protein, GFAP). The results showed that EA and paroxetine could improve depression-like behavior in CUMS rats, and up-regulated the expression level of FGF2 in the hippocampus, increased GFAP protein expression and the mean optical density of GFAP-immunoreactive astrocyte (GFAP-ir astrocyte). Our findings have identified that EA could ameliorate depressive-like behaviors possibly by regulating the expression of FGF2 in the hippocampus, and the mechanism might be related to the effect of FGF2 on astrocytes.

Melatonin reduces neuropathic pain behavior and glial activation through MT2 melatonin receptor modulation in a rat model of lysophosphatidylcholine-induced demyelination neuropathy

In this study, we investigated whether melatonin treatment prevents development of neuropathic pain via suppression of glial mitogen-activated protein kinases (MAPKs) activation in the cuneate nucleus (CN) in a lysophosphatidylcholine (LPC)-induced median nerve demyelination neuropathy model. Rats were fed orally with melatonin once a day at a dose of 37.5, 75, or 150 mg/kg 30 min before until 3 days after LPC treatment. Subsequently, behavioral tests were conducted on these animals, and immunohistochemistry and immunoblotting were used for qualitative and quantitative analysis of glia and MAPKs, including ERK, JNK, and p38, activation. Enzyme-linked immunosorbent assays were applied to measure pro-inflammatory cytokine responses. Furthermore, intra-CN microinjection of S26131 (MT1 receptor antagonist), 4P-PDOT (MT2 receptor antagonist), or prazosin (MT3 receptor antagonist) were performed to investigate the association between melatonin receptor subtypes and effects of melatonin on demyelination neuropathy. LPC treatment of the median nerve induced a significant increase in glial fibrillary acidic protein (GFAP; an astrocyte marker) and ED1 (an activated microglia marker) immunoreactivity in the ipsilateral CN and led to development of neuropathic pain behavior. Inspection of GFAP-immunoreactive astrocytes revealed that astrocytic hypertrophy, but not proliferation, contributed to increased GFAP immunoreactivity. Double immunofluorescence showed that both GFAP-immunoreactive astrocytes and ED1-immunoreactive microglia co-expressed p-ERK, p-JNK, and p-p38 immunoreactivity. Melatonin administration dose-dependently reduced neuropathic pain behavior, decreased glial and MAPKs activation, and diminished the release of pro-inflammatory cytokines in the ipsilateral CN after LPC treatment. Furthermore, 4P-PDOT, but not S26131 or prazosin, antagonized the therapeutic effects of melatonin. In conclusion, administration of melatonin, via its cognate MT2 receptor, inhibited activation of glial MAPKs, production of pro-inflammatory cytokines, and development of demyelination-induced neuropathic pain behavior.

Characterization of Vimentin-Immunoreactive Astrocytes in the Human Brain.

Astrocytes are commonly identified by their expression of the intermediate filament protein glial fibrillary acidic protein (GFAP). GFAP-immunoreactive (GFAP-IR) astrocytes exhibit regional heterogeneity in density and morphology in the mouse brain as well as morphological diversity in the human cortex. However, regional variations in astrocyte distribution and morphology remain to be assessed comprehensively. This was the overarching objective of this postmortem study, which mainly exploited the immunolabeling of vimentin (VIM), an intermediate filament protein expressed by astrocytes and endothelial cells which presents the advantage of more extensively labeling cell structures. We compared the densities of vimentin-immunoreactive (VIM-IR) and GFAP-IR astrocytes in various brain regions (prefrontal and primary visual cortex, caudate nucleus, mediodorsal thalamus) from male individuals having died suddenly in the absence of neurological or psychiatric conditions. The morphometric properties of VIM-IR in these brain regions were also assessed. We found that VIM-IR astrocytes generally express the canonical astrocytic markers Aldh1L1 and GFAP but that VIM-IR astrocytes are less abundant than GFAP-IR astrocytes in all human brain regions, particularly in the thalamus, where VIM-IR cells were nearly absent. About 20% of all VIM-IR astrocytes presented a twin cell morphology, a phenomenon rarely observed for GFAP-IR astrocytes. Furthermore VIM-IR astrocytes in the striatum were often seen to extend numerous parallel processes which seemed to give rise to large VIM-IR fiber bundles projecting over long distances. Moreover, morphometric analyses revealed that VIM-IR astrocytes were more complex than their mouse counterparts in functionally homologous brain regions, as has been previously reported for GFAP-IR astrocytes. Lastly, the density of GFAP-IR astrocytes in gray and white matter were inversely correlated with vascular density, but for VIM-IR astrocytes this was only the case in gray matter, suggesting that gliovascular interactions may especially influence the regional heterogeneity of GFAP-IR astrocytes. Taken together, these findings reveal special features displayed uniquely by human VIM-IR astrocytes and illustrate that astrocytes display important region- and marker-specific differences in the healthy human brain.

Slc6a3-dependent expression of a CAPS-associated Nlrp3 allele results in progressive behavioral abnormalities and neuroinflammation in aging mice

BackgroundAn association between neuroinflammation and age-related neurologic disorders has been established but the molecular mechanisms and cell types involved have not been thoroughly characterized. Activity of the proinflammatory NLRP3 inflammasome is implicated in Alzheimer’s and Parkinson’s disease and our recent studies in patients suggest that dopaminergic neurons within the degenerating mesencephalon express NLRP3 throughout the progression of PD. Here, we directly test the impact of enhanced inflammasome activity in mesencephalic neurons by characterizing motor function, tissue integrity, and neuroinflammation in aging mice harboring hyperactivating mutations within the endogenous murine Nlrp3 locus, enabled only in cells expressing the dopaminergic neuron-specific Slc6a3 promoter.MethodsWe compared mice harboring inducible alleles encoding the cryopyrin-associated periodic syndrome activating mutations Nlrp3A350V and Nlrp3L351P inserted into the endogenous mouse Nlrp3 locus. Tissue specific expression was driven by breeding these animals with mice expressing Cre recombinase under the control of the dopaminergic neuron-specific Slc6a3 promoter. The experimental mice, designed to express hyperactive NLRP3 only when the endogenous mouse Nlrp3 promotor is active in dopaminergic neurons, were analyzed throughout 18 months of aging using longitudinal motor function assessments. Biochemical and histologic analyses of mesencephalic tissues were conducted in 1- and 18-month-old animals.ResultsWe observed progressive and significant deficits in motor function in animals expressing Nlrp3L351P, compared with animals expressing Nlrp3WT and Nlrp3A350V. Age-dependent neuroinflammatory changes in the mesencephalon were noted in all animals. Analysis of GFAP-immunoreactive astrocytes in the substantia nigra revealed a significant increase in astrocyte number in animals expressing Nlrp3L351P compared with Nlrp3WT and Nlrp3A350V. Further analysis of Nlrp3L351P striatal tissues indicated genotype specific gliosis, elevated Il1b expression, and both morphologic and gene expression indicators of proinflammatory A1 astrocytes.ConclusionsDopaminergic neurons have the potential to accumulate NLRP3 inflammasome activators with age, including reactive oxygen species, dopamine metabolites, and misfolded proteins. Results indicate the Nlrp3 locus is active in dopaminergic neurons in aging mice, and that the hyperactive Nlrp3L351P allele can drive neuroinflammatory changes in association with progressive behavioral deficits. Findings suggest neuronal NLRP3 inflammasome activity may contribute to neuroinflammation observed during normal aging and the progression of neurologic disorders.

Astrocyte Changes in the Prefrontal Cortex From Aged Non-suicidal Depressed Patients.

Glia alterations in the anterior cingulate cortex (ACC) and dorsolateral prefrontal cortex (DLPFC) have been postulated to play an important role in the pathophysiology of psychiatric disorders. Astroglia is the most abundant type of glial cells in the central nervous system. The expression levels of astrocyte markers (glial fibrillary acidic protein (GFAP), synemin-α, synemin-β, vimentin, nestin) in isolated gray matter from postmortem ACC and DLPFC were determined to investigate the possible involvement of astrocytes in depression. Donors were aged non-suicidal subjects with bipolar disorder (BPD) or major depressive disorder (MDD), and matched controls. GFAP mRNA levels were significantly increased in the ACC of BPD patients. However, GFAP immunohistochemistry showed that the area fraction of GFAP immunoreactive astrocytes was decreased in the ACC of BPD patients, while there were no changes in the cell density and integrated optical density (IOD), indicating that there might be a reduction of GFAP-positive astrocyte processes and remodeling of the astrocyte network in BPD. Furthermore, in controls, DLPFC GFAP mRNA levels were significantly lower with a time of death at daytime (08:01–20:00 h) compared to nighttime (20:01–08:00 h). In depression, such a diurnal pattern was not present. These findings in BPD and MDD subjects warrant further studies given the crucial roles of astrocytes in the central nervous system.

P: 22 Deficit of Short Working Memory in Rat With Thioacetamide-Induced Progressive Acute Hepatic Encephalopathy Involving Serotonin Innervation and Astroglia Dysfunctions

BACKGROUND: Hepatic encephalopathy (HE) is defined as the whole neurological and neuropsychiatric disorders arising from both acute and chronic liver failures. Patients with chronic HE exhibited an impairment of learning and memory abilities as well as attention. The aim of the present study is to evaluate the progression of short working memory alteration in acute HE and to delineate the glial and the neuronal alterations which may underlie such cognitive impairment. METHODS: The study was performed in Sprague-Dawley rats with acute liver failure induced by thioacetamide (TAA) at a dose of 300 mg/kg i.p. Different stages of acute HE was defined as: 12 h, 24 h and 36 h following administration of TAA. Working memory was assessed by the T-Maze test via the percentage of alternation behavior, as well, an immunohistochemical analysis of GFAP in the hippocampus and serotonin (5-HT) within the dorsal Raphe nucleus (DRN). RESULTS: Our data showed a progressive loss of the alternation behavior, which was accompanied by a time dependent and region-specific changes of GFAP-immunoreactive astrocytes within the hippocampus together with a reduced 5-HT immunoreactivity within the DRN. CONCLUSIONS: Our data revealed for the first time, a progressive loss of short memory function in acute HE, resulting from acute liver dysfunction which may involve a possible gliopathy as well as a 5-HTergic dysfunctions.

Astrogliosis Associated With Behavioral Abnormality in a Non-anaphylactic Mouse Model of Cow's Milk Allergy.

Etiology of neuropsychiatric disorders is complex, involving multiple factors that can affect the type and severity of symptoms. Although precise causes are far from being identified, allergy or other forms of hypersensitivity to dietary ingredients have been implicated in triggering or worsening of behavioral and emotional symptoms, especially in patients suffering from depression, anxiety, attention-deficit hyperactivity, and/or autism. Among such ingredients, cow's milk, along with wheat gluten, is commonly suspected. However, the contributory role of cow's milk in these disorders has not been elucidated due to insufficient pathophysiological evidence. In the present study, we therefore investigated neuroinflammatory changes that are associated with behavioral abnormality using a non-anaphylactic mouse model of cow's milk allergy (CMA). Male and female C57BL/6J mice were subjected to a 5-week oral sensitization procedure without or with a major milk allergen, beta-lactoglobulin (BLG). All mice were then later challenged with BLG, and their anxiety- and depression-associated behaviors were subsequently assessed during the 6th and 7th weeks. We found that BLG-sensitized male mice exhibited significantly increased anxiety- and depression-like behavior, although they did not display anaphylactic reactions when challenged with BLG. Female behavior was not noticeably affected by BLG sensitization. Upon examination of the small intestines, reduced immunoreactivity to occludin was detected in the ileal mucosa of BLG-sensitized mice although the transcriptional expression of this tight-junction protein was not significantly altered when measured by quantitative RT-PCR. On the other hand, the expression of tumor necrosis factor alpha (TNFα) in the ileal mucosa was significantly elevated in BLG-sensitized mice, suggesting the sensitization had resulted in intestinal inflammation. Inflammatory responses were also detected in the brain of BLG-sensitized mice, determined by the hypertrophic morphology of GFAP-immunoreactive astrocytes. These reactive astrocytes were particularly evident near the blood vessels in the midbrain region, resembling the perivascular barrier previously reported by others in experimental autoimmune encephalitis (EAE) mouse models. Interestingly, increased levels of COX-2 and TNFα were also found in this region. Taken together, our results demonstrated that BLG sensitization elicits inflammatory responses in the intestine and brain without overt anaphylactic signs of milk allergy, signifying food allergy as a potential pathogenic factor of neuropsychiatric disorders.

Catfish (Arius bilineatus, Val.) skin protein preparation ameliorates neurobehavioral and axonal and neuronal histopathological alterations in a sciatic nerve crush injury rat model: a novel approach

Preliminary investigations showed that soluble protein fraction‐Fraction C (SPF‐FC) from catfish skin preparations (CSP) from the Arabian Gulf catfish (Arius bilineatus, Val.) exhibit potent anti‐inflammatory and healing properties as shown in our previous clinical trials for the healing of non‐healing diabetic foot ulcers, chronic back pain, and some other neurological disorders. The present study was designed to investigate the neuroprotective and neuroregenerative properties of SPF‐FC derived from CSP on the spinal neurons, regeneration of sciatic nerve fibers, and recovery of sensory‐motor functions in the sciatic nerve crush injury model. Male Wistar rats (weight 250–300 g) were randomly assigned to four groups: (I) SHAM, (II) CRUSH, (III) CRUSH+SPF treated intraperitoneally (IP) and (IV) CRUSH+SPF treated subcutaneously (SC) groups. Rats in CRUSH groups underwent surgery, where the sciatic nerve was crushed, followed by treatment with saline (II) or with 3–12 mg/Kg SPF IP (III) or SC (IV) daily for 14 days. All animals were tested for the neurobehavioral parameters throughout the 6 weeks of the study. Sciatic nerve and spinal cord tissues were processed for light and electron histological examinations, stereological analysis, immunohistochemical and biochemical examinations at Week 4 and Week 6 post‐injury. Administration of SPF IP or SC significantly (p&lt;0.05–0.0001) enhanced the sensory and motor performance on different neurobehavioral functional tests of the nerve‐injured rats. The stereological evaluation revealed significant (p&lt;0.000) histomorphological evidence of neuroregeneration in the sciatic nerve of the CRUSH+ SPF groups compared to controls at week 4 and 6. SPF‐FC treatment significantly prevented the decrease in the number of the NeuN‐immunoreactive spinal neurons ipsilateral to the crush injury, decreased the GFAP immunoreactive astrocytes and increased the GAP‐43 immunoreactivity. The data analysis supports our hypothesis that SPF‐FC immunoreactivity treatment lessens neurobehavioral deficits, enhances axonal regeneration and ameliorate histomorphological nerve alterations following nerve injury. Further, SPF‐FC protects spinal neurons and enhances subcellular recovery by decreasing the astrocytic activity and GAP‐43, thus improves nerve regeneration and functional recovery.Support or Funding InformationThis work was supported by Kuwait University Research Administration (KU Grants No. SL03/14 and SL04/09 and RCF/OMICSRU project SRUL02/13) and Kuwait Foundation for the Advancement of Sciences (Grant No. KFAS 2013‐1207‐01A‐D).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Kinetic deterioration of short memory in rat with acute hepatic encephalopathy: Involvement of astroglial and neuronal dysfunctions

Hepatic encephalopathy (HE) is a complex neuropsychiatric syndrome resulting from acute or chronic hepatic impairments. The clinical features of HE include attention as well as a mild cognitive deficits associated with impaired attentional and executive networks in patients as well as in animal models of HE. The underlining pathomechanism of memory impairment in HE patients is still not fully understood; however, it may involve a possible gliopathy as well as neuropathy. The aim of the present investigation is to assess progression of short working memory deterioration in acute HE and to delineate the glial and the neuronal alteration which may underlie such cognitive impairment. The study was carried out in male Sprague-Dawley rats with acute liver failure induced by thioacetamide (TAA). The study was performed on different stages of acute HE; 12 h, 24 h and 36 h following administration of TAA. The liver functions were assessed via different biochemical markers (ALT, AST, bilirubin, urea and creatinine) and an histopathological examination of the liver tissue. While for the behavioral study, we used T-Maze test to assess short working memory using the percentage of alternation behavior, together with an immunohistochemical analysis of the Glial Fibrillary Acidic Protein (GFAP) as the key marker of astrocytes in the hippocampus, as well as serotonin (5-HT) for 5-HTergic neurons within the dorsal Raphe nucleus (DRN). Our data revealed a progressive loss of liver tissue integrity with inflammation and hepatocytes degeneration which was associated to obvious loss of the liver function. In parallel, we observed a gradual alteration of the alternation behavior, as a sign of altered short working memory in the acute HE rats. At the central level, the immunohistochemical study showed a time dependent region-specific changes of GFAP-immunoreactive astrocytes within the hippocampus. While within the DRN, serotonin levels declined progressively in a time-dependant manner. Our data revealed for the first time, a gradual loss of short memory function in acute HE, resulting from liver dysfunction. Such cognitive deterioration may involve a possible gliopathy as well as a 5-HTergic dysfunction which could be considered as a new key element for understanding the basis of memory and attention loss in HE patients.

Fate of Astrocytes in The Gerbil Hippocampus After Transient Global Cerebral Ischemia

Neuronal death and reactive gliosis are major features of brain tissue damage following transient global cerebral ischemia (tgCI). This study investigated long-term changes in neuronal death and astrogliosis in the gerbil hippocampus for 180 days after 5 min of tgCI. A massive loss of pyramidal neurons was found in the hippocampal CA1 area (CA1) area between 5 and 30 days after tgCI by Fluoro-Jade B (FJB, a marker for neuronal degeneration) histofluorescence staining, but pyramidal neurons in the CA2/3 area did not die. The reaction of astrocytes (astrogliosis) was examined by glial fibrillary acidic protein (GFAP) immunohistochemistry. Morphological change or degeneration (death) of the astrocytes was found in the CA1 area after tgCI, but, in the CA2/3 area, astrogliosis was hardly shown. GFAP immunoreactive astrocytes in the CA1 area was significantly increased in number with time and peaked at 30 days after tgCI, and they began to be degenerated or dead from 40 days after tgCI. The effect was examined by double immunofluorescence staining for FJB and GFAP. The number of FJB/GFAP+ cells (degenerating astrocytes) was gradually increased with time after tgCI. At 180 days after tgCI, FJB/GFAP+ cells were significantly decreased, but FJB+ cells (dead astrocytes) were significantly increased. In brief, 5 min of tgCI induced a progressive degeneration of CA1 pyramidal neurons from 5 until 30 days with an increase of reactive astrocytes, and, thereafter, astrocytes were degenerated with time and dead at later times. This phenomenon might be shown due to the death of neurons.