Glial Fibrillary Acidic Protein Intensity Research Articles

Adolescent alcohol binge drinking and withdrawal: behavioural, brain GFAP-positive astrocytes and acute methamphetamine effects in adult female rats

RationaleAlcopop beverages are generally the first alcoholic beverage that young females drink which contain high levels of sugar and alcohol. The over-consumption of these drinks may encourage alcohol co-administration with methamphetamine (METH) impacting on drinking behaviour and glial function.AimsThe aims of this study were to evaluate the effect of adolescent binge alcohol exposure on consumption level, anxiety-like behaviour, cross-sensitization with METH and on astrocyte expression in reward related brain regions.MethodsAdolescent female Sprague-Dawley rats had daily 1-hour oral alcohol consumption of alcopop (ALCP; with sucrose) or ethanol-only (ETOH; without sucrose), transitioned from 5 to 15% (v/v) ethanol content for 34 days. Water and sucrose groups act as controls. During alcohol withdrawal, rats were tested for anxiety on the elevated plus maze (EPM) and locomotor activity following saline or METH (1 mg/kg i.p) treatment. Brains were then collected to assess astrocyte immunofluorescence for glial fibrillary acidic protein (GFAP) in reward-related brain regions.ResultsRats pretreated with 5% ALCP consumed significantly more volume and ethanol intake when compared to 5% EtOH rats. Both ALCP and EtOH groups had a higher preference ratio for 5% than 15% alcohol solutions and ALCP rats had greater ethanol intake at 15% than EtOH rats. Alcohol withdrawal showed no significant differences between groups on anxiety, METH cross-sensitization effects or GFAP intensity in the regions studied.ConclusionsOverall, the addition of sucrose to alcoholic solutions encouraged female rats to consume larger volumes and greater ethanol intake compared to ethanol-only solutions, yet did not have long lasting effects on behaviour and astrocytes.

Sera from people with HIV and depression induce commensurate metabolic alterations in astrocytes: toward precision diagnoses and therapies.

People with HIV (PWH) have high rates of depression and neurocognitive impairment (NCI) despite viral suppression on antiretroviral therapy (ART). Mounting evidence suggests that immunometabolic disruptions may contribute to these conditions in some PWH. We hypothesized that metabolic dysfunction in astrocytes is associated with depressive symptoms and cognitive function in PWH. Human astrocytes were exposed to sera from PWH (n=40) with varying degrees of depressive symptomatology and cognitive function. MitoTrackerTM Deep Red FM (MT) was used to visualize mitochondrial activity and glial fibrillary acidic protein (GFAP) as an indicator of astrocyte reactivity using the high-throughput fluorescent microscopy and image analyses platform, CellInsight CX5 (CX5). The Seahorse platform was used to assess glycolytic and mitochondrial metabolism. More severe depression, as indexed by higher Beck's Depression Inventory (BDI-II) scores, was associated with lower MT signal measures. Better cognitive function, as assessed by neuropsychiatric testing t-scores, was associated with increased MT signal measures. GFAP intensity negatively correlated with several cognitive t-scores. Age positively correlated with (higher) MT signal measures and GFAP intensity. Worse depressive symptoms (higher BDI-II scores) were associated with decreased oxygen consumption rate and spare respiratory capacity, concomitant with increased extracellular acidification rate in astrocytes. These findings show that factors in the sera of PWH alter mitochondrial activity in cultured human astrocytes, suggesting that mechanisms that alter mitochondrial and astrocyte homeostasis can be detected peripherally. Thus, in vitro cultures may provide a model to identify neuropathogenic mechanisms of depression or neurocognitive impairment in PWH and test personalized therapeutics for neurologic and psychiatric disorders.

Differences in Anatomical Outcomes Between Early Chronic and Far Chronic Time-Points After Transplantation of Spinal Cord Neural Progenitor Cells in Mice.

Spinal cord injury (SCI) affects millions of people worldwide. Neural progenitor cell (NPC) transplantation is a promising treatment for regenerating lost spinal cord tissue and restoring neurological function after SCI. We conducted a literature search and found that less than a quarter of experimental rodent cell and tissue transplantation studies have investigated anatomical outcomes at longer than 4 months post-transplantation. This is a critical topic to investigate, given that stem and progenitor cell therapies would need to remain in place throughout the lifetime of an individual. We sought to determine how commonly assessed anatomical outcomes evolve between early and far chronic time-points post-NPC transplantation. At either 8 weeks or 26 weeks following transplantation of NPCs into sites of cervical SCI, we evaluated graft neuronal density, astroglial cell density, graft axon outgrowth, and regeneration of host axon populations into grafts in male and female mice. We found that graft neuronal density does not change over time, but the numbers of graft-associated astrocytes and glial fibrillary acidic protein intensity is significantly increased in the far chronic phase compared with the early chronic time-point. In addition, graft axon outgrowth was significantly decreased at 26 weeks post-transplantation compared with 8 weeks post-transplantation. In contrast, corticospinal axon regeneration into grafts was not diminished over time, but rather increased significantly from early to far chronic periods. Interestingly, we found that graft neuronal density is significantly influenced by sex of the host animal, suggesting that sex-dependent processes may shape graft composition over time. Collectively, these results demonstrate that NPC transplants are dynamic and that commonly assessed outcome measures associated with graft efficacy evolve over the weeks to months post-transplantation into the spinal cord.

Optic Nerve Head Myelin-Related Protein, GFAP, and Iba1 Alterations in Non-Human Primates With Early to Moderate Experimental Glaucoma.

PurposeThe purpose of this study was to test if optic nerve head (ONH) myelin basic protein (MBP), 2′,3′-cyclic nucleotide 3′-phosphodiesterase (CNPase), glial fibrillary acidic protein (GFAP), and ionized calcium binding adaptor molecule 1 (Iba1) proteins are altered in non-human primate (NHP) early/moderate experimental glaucoma (EG).MethodsFollowing paraformaldehyde perfusion, control and EG eye ONH tissues from four NHPs were paraffin embedded and serially (5 µm) vertically sectioned. Anti-MBP, CNPase, GFAP, Iba1, and nuclear dye-stained sections were imaged using sub-saturating light intensities. Whole-section images were segmented creating anatomically consistent laminar (L) and retrolaminar (RL) regions/sub-regions. EG versus control eye intensity/pixel-cluster density data within L and two RL regions (RL1 [1-250 µm]/RL2 [251-500 µm] from L) were compared using random effects models within the statistical program “R.”ResultsEG eye retinal nerve fiber loss ranged from 0% to 20%. EG eyes’ MBP and CNPase intensity were decreased within the RL1 (MBP = 31.4%, P < 0.001; CNPase =62.3%, P < 0.001) and RL2 (MBP = 19.6%, P < 0.001; CNPase = 56.1%, P = 0.0004) regions. EG eye GFAP intensity was decreased in the L (41.6%, P < 0.001) and RL regions (26.7% for RL1, and 28.4% for RL2, both P < 0.001). Iba1+ and NucBlue pixel-cluster density were increased in the laminar (28.2%, P = 0.03 and 16.6%, P = 0.008) and both RL regions (RL1 = 37.3%, P = 0.01 and 23.7%, P = 0.0002; RL2 = 53.7%, P = 0.002 and 33.2%, P < 0.001).ConclusionsRetrolaminar myelin disruption occurs early in NHP EG and may be accompanied by laminar and retrolaminar decreases in astrocyte process labeling and increases in microglial/ macrophage density. The mechanistic and therapeutic implications of these findings warrant further study.

Time Course of Astrocyte Activation in Mouse Nucleus Tractus Solitarius During Exposure to Hypoxia

Chronic sustained hypoxia (CH) occurs in populations living at high altitude and in patients with chronic pulmonary disease. Exposure to CH produces ventilatory acclimatization to hypoxia (VAH) and increases the hypoxic ventilatory response (HVR) by mechanisms that involve areas of the brainstem that control breathing. The nucleus tractus solitarius (NTS) is a sensory integrative center in the medulla receiving carotid body afferents and known to be crucial for VAH. Our previous results demonstrated that glia cells in the rat NTS contribute to VAH but these mechanisms have not been studied in mice exposed to CH beyond 24 hours. We hypothesized that CH produces an early transient activation of astrocytes and microglia in the mouse brainstem as observed in rats. To study the activation of glial cells with CH, we exposed mice to normobaric hypoxia (10% FiO2) for 0.5, 1 and 4 hours and 1 and 7 days. Mice were perfused with 4% paraformaldehyde and brainstem sections were obtained. We quantified astrocyte activation by measuring glial fibrillary acidic protein (GFAP) intensity with immunofluorescence, and microglial activation by measuring branch morphology using Iba‐1 marker. Exposure to CH significantly increased GFAP intensity after 30 minutes and 1day while microglia morphology did not change significantly. The results differ from those in rats (J. Neurophysiol.117: 1625–1635, 2017) and suggest that time‐dependent astrocyte activation in the NTS may contribute to VAH.Support or Funding InformationSupported by NIH RO1 HL‐081823

Diabetic ketoacidosis in juvenile rats is associated with reactive gliosis and activation of microglia in the hippocampus.

Type 1 diabetes may be associated with structural and functional alterations in the brain. The role of diabetic ketoacidosis (DKA) in causing these alterations has not been well explored. We used immunohistochemical staining to investigate cellular alterations in brain specimens from juvenile rats with DKA before, during, and after treatment with insulin and saline, and compared these to samples from diabetic rats and normal controls. Glial fibrillary acidic protein (GFAP) staining intensity was increased in the hippocampus during DKA and increased further during insulin/saline treatment. Twenty-four and 72 h after treatment, hippocampal GFAP intensity declined but remained above control levels. There were no significant changes in GFAP intensity in the cortex or striatum. OX42 staining intensity was increased during untreated DKA and increased further during insulin/saline treatment in the hippocampus and cortex. NeuN staining intensity was decreased after DKA treatment in the striatum but not in other regions. DKA causes inflammatory changes in the brain including reactive gliosis and activation of microglia. These findings are present during untreated DKA, but intensify during insulin/saline treatment. The hippocampus was disproportionately affected, consistent with previous studies showing deficits in hippocampal functions in rats after DKA recovery and decreased memory capacity in children with a history of DKA.

Gestational treatment of folic acid attenuates blood–brain barrier leakage in pregnant- and prepubertal rats after pentylenetetrazole-induced seizure

Objectives: Folic acid (FA) is physiologically important in mammals and is a common vitamin supplement used during pregnancy and lactation. Numerous studies have reported that FA significantly improves endothelial function. The blood–brain barrier (BBB) plays an important role in maintaining the microenvironment required for neuronal function, but its unique structure is damaged by epileptic seizures. The aim of this study was to evaluate the potential protective role of FA on BBB leakage, as well as on the reactive astrogliosis in pregnant rats and their prepubertal offspring during pentylenetetrazole (PTZ)-induced epileptic seizure.Methods: Pregnant rats were treated with FA (5 mg/kg) and PTZ on gestational days 0–19 and 19, respectively. The pups were treated with PTZ at puberty. Evans blue was used to evaluate BBB integrity. Reactive astrogliosis was defined using immunohistochemical analysis for glial fibrillary acidic protein (GFAP). Mean arterial blood pressure (MABP) was measured at the femoral artery.Results: A moderate decrease in BBB leakage was observed in FA-treated pregnant and prepubertal animals (P < 0.05). MABP was decreased significantly in pregnant rats (P < 0.05). The epilepsy-induced increase in MABP was less prominent in pregnant animals (P < 0.05). GFAP intensity decreased in PTZ-treated pregnant animals (P < 0.01) and FA-treated prepubertal rats.Discussion: Our findings suggest that FA, which is used as a maternal vitamin to promote normal fetus development, may be beneficial against seizure-induced neuronal damage by decreasing BBB leakage and reactive astrogliosis in pregnant and prepubertal rats.

Amyloid beta(1-40)-induced astrogliosis and the effect of genistein treatment in rat: a three-dimensional confocal morphometric and proteomic study.

Astrocytes are highly involved in regulation and homeostasis of the extracellular environment in the healthy brain. In pathological conditions, these cells play a major role in the inflammatory response seen in CNS tissues, which is called reactive astrogliosis and includes hypertrophy and proliferation of astrocytes. Here, we performed 3D confocal microscopy to evaluate the morphological response of reactive astrocytes positive for glial fibrillary acidic protein (GFAP) in rats, to the presence of Aβ1–40 in the rat brain before and after treatment with genistein. In 50 astrocytes per animal, we measured the volume and surface area for the nucleus, cell body, the entire cell, the tissue covered by single astrocytes and quantified the number and length of branches, the density of the astrocytes and the intensity of GFAP immunoreactivity. Injecting Aβ1–40 into the brain of rats caused astrogliosis indicated by increased values for all measured parameters. Mass spectrometric analysis of hippocampal tissue in Aβ1–40-injected brain showed decreased amounts of tubulins, enolases and myelin basic protein, and increased amounts of dihydropyrimidinase-related protein 2. In Aβ1–40-injected rats pretreated with genistein, GFAP intensity was decreased to the sham-operated group level, and Aβ1–40-induced astrogliosis was significantly ameliorated.

Reaction of Müller cells in an experimental rat model of increased intraocular pressure following timolol, latanoprost and brimonidine

The aim of this study was to evaluate the reaction of Müller cells in an experimental rat model of intraocular pressure (IOP) and their response to treatment with ocular hypotensive drugs. Episcleral vein cauterization in unilateral eyes of Wistar rats was performed to produce elevated IOP. The animals were divided into five groups: control, experimental, and experimental treated with timolol, latanoprost or brimonidine. Histological sections of retina were studied by immunochemistry with antibodies to glial fibrillary acidic protein (GFAP), and the percentage of labeled area was measured to evaluate the degree of reactive gliosis. In the experimental group, the Müller cells showed hypertrophy and a significant increase in GFAP (4.39 ± 0.32%) in relation to retinas of the control group (2.05 ± 0.14%). Gliosis was detected in all three treated groups, with a varying increase in GFAP intensity. The timolol-treated group showed the most intense and persistent glial reactivity after 3 months of treatment (13.89 ± 0.63%). Treatment with brimonidine, however, resulted in a decrease in the level of GFAP immunoreactivity (8.37 ± 0.4%). The group treated with latanoprost showed the lowest glial reactivity (4.8 ± 0.36%). Given that all three drugs are effective hypotensive agents, their neuroprotective effect could be related with other factors, such as gliosis, which, over long periods may have noxious effects on the neurons. Thus, hypotensives like brimonidine, and specially latanoprost, may afford greater neuroprotection to the ganglion cells by attenuating the retinal glial reaction.

Brain-Specific Deletion of Extracellular Signal-Regulated Kinase 2 Mitogen-Activated Protein Kinase Leads to Aberrant Cortical Collagen Deposition

The mitogen-activated protein kinases extracellular signal-regulated kinase (ERK)1 and 2 are essential intracellular mediators of numerous transmembrane signals. To investigate neural-specific functions of ERK2 in the brain, we used a Cre/lox strategy using Nestin:Cre to drive recombination in neural precursor cells. Nestin:Cre;ERK2(fl/fl) conditional knockout (cKO) mice have architecturally normal brains and no gross behavioral deficits. However, all cKO mice developed early-onset (postnatal day 35 to 40) frontal cortical astrogliosis, without evidence of neuronal degeneration. Frontoparietal cortical gray matter, but not underlying white matter, was found to contain abundant pericapillary and parenchymal reticulin fibrils, which were shown by immunohistochemistry to contain fibrillar collagens, including type I collagen. ERK1 general KO mice showed neither fibrils nor astrogliosis, indicating a specific role for ERK2 in the regulation of brain collagen. Collagen fibrils were also observed to a lesser extent in GFAP:Cre;ERK2(fl/fl) mice but not in CamKII-Cre;ERK2(fl/fl) mice (pyramidal neuron specific), consistent with a possible astroglial origin. Primary astroglial cultures from cKO mice expressed elevated fibrillar collagen levels, providing further evidence that the phenotype may be cell autonomous for astroglia. Unlike most other tissues, brain and spinal cord parenchyma do not normally contain fibrillar collagens, except in disease states. Determining mechanisms of ERK2-mediated collagen regulation may enable targeted suppression of glial scar formation in diverse neurological disorders.

Elevated pressure induced astrocyte damage in the optic nerve

Astrocytes maintain an intimate relationship with central nervous system (CNS) neurons and play a crucial role in regulating their biochemical environment. A rise in neural tissue pressure in the CNS is known to lead to axonal degeneration however the response of astrocytes during the early stages of neural injury has not been studied in great detail. The optic nerve is a readily accessible model in which to study CNS axonal injury. Previous work from our laboratory has shown that an acute increase in intraocular pressure (IOP) results in axonal cytoskeleton changes and axonal transport retardation within the optic nerve head. Axonal changes occurred in a time-dependent manner with the magnitude of change being proportional to the duration of the IOP rise. Using glial fibrillary acidic protein (GFAP) as a marker of astrocytes we have now studied pressure induced changes in astrocyte structure in the optic nerve head. Using confocal microscopy we found that an increase in IOP resulted in morphological changes in the astrocytes that were consistent with previous reports of swelling. In addition there was also a decrease in GFAP intensity within these astrocytes. These changes occurred in a time-dependent manner with the chronology of change coinciding with that of axonal change. There was no evidence of apoptosis in regions where astrocyte changes were found. The present results provide evidence that in the early stages of neural tissue pressure rise there are both astrocyte and axonal injury.