Immature rat brain slices exposed to oxygen–glucose deprivation as an in vitro model of neonatal hypoxic–ischemic encephalopathy

Fernández-López, David; Martı́nez-Orgado, José; Casanova, Ignacio; Bonet, Bartolomé; Leza, Juan C.; Lorenzo, Pedro; Moro, Marı́a A.; Lizasoaín, Ignacio

doi:10.1016/j.jneumeth.2005.01.005

Cited by 32 publications

(26 citation statements)

References 33 publications

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“…Neither TNFa nor IL-6 was increased after pure OGD, which differs from previous studies, showing a prominent release of these cytokines after ischemia in vivo (Saito et al, 1996) and in hippocampal slices in vitro (Fernandez-Lopez et al, 2005). In contrast, the levels of MCP-1 increased after OGD in LPS preexposed slices, suggesting a role for this chemokine in postischemic processes.…”

Section: Lipopolysaccharide-induced Cytokine Release and Activation Ocontrasting

confidence: 99%

Tumor Necrosis Factor Receptor-1 is Essential for LPS-Induced Sensitization and Tolerance to Oxygen—Glucose Deprivation in Murine Neonatal Organotypic Hippocampal Slices

Markus

Cronberg

Cilio

et al. 2008

J Cereb Blood Flow Metab

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Inflammation and ischemia have a synergistic damaging effect in the immature brain. The role of tumor necrosis factor (TNF) receptors 1 and 2 in lipopolysaccharide (LPS)-induced sensitization and tolerance to oxygen-glucose deprivation (OGD) was evaluated in neonatal murine hippocampal organotypic slices. Hippocampal slices from balb/c, C57BL/6 TNFR1 À/À , TNFR2 À/À , and wild-type (WT) mice obtained at P6 were grown in vitro for 9 days. Preexposure to LPS immediately before OGD increased propidium iodide-determined cell death in regions CA1, CA3, and dentate gyrus from 4 up to 48 h after OGD (P < 0.001). Extending the time interval between LPS exposure and OGD to 72 h resulted in tolerance, that is reduced neuronal cell death after OGD (P < 0.05). Slices from TNFR1 À/À mice showed neither LPS-induced sensitization nor LPS-induced tolerance to OGD, whereas both effects were present in slices from TNFR2 À/À and WT mice. Cytokine secretion (TNFa and interleukin-6) during LPS exposure was decreased in TNFR1 À/À slices and increased in TNFR2 À/À as compared with WT slices. We conclude that LPS induces sensitization or tolerance to OGD depending on the time interval between exposure to LPS and OGD in murine hippocampal slice cultures. Both paradigms are dependent on signaling through TNFR1.

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Section: Lipopolysaccharide-induced Cytokine Release and Activation Ocontrasting

confidence: 99%

Tumor Necrosis Factor Receptor-1 is Essential for LPS-Induced Sensitization and Tolerance to Oxygen—Glucose Deprivation in Murine Neonatal Organotypic Hippocampal Slices

Markus

Cronberg

Cilio

et al. 2008

J Cereb Blood Flow Metab

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“…The experimental protocol has been described elsewhere (14). Briefly, newborn Wistar rats (7-10 d old) were killed by decapitation (according to the Committee of Animal Care at the Universidad Complutense of Madrid), the forebrain was removed and coronally cut (1 mm anterior and 3 mm posterior to the bregma), and the central portion was sliced (0.5 mm slice thickness) using a Vibroslice (WPI, Stevenage, UK) into cold (12-14°C) modified Krebs-Henseleit solution (preincubation solution) containing (mM): NaCl (120), KCl (2), CaCl2 (0.5), Abbreviations: iNOS, inducible nitric oxide synthase; LDH, lactate dehydrogenase; NHIE, neonatal hypoxic-ischemic encephalopathy; OGD, oxygen-glucose deprivation; WIN, R(ϩ)-WIN-55212-2…”

Section: Methodsmentioning

confidence: 99%

“…As a marker of necrotic tissue damage, LDH released from damaged cells in the slices was determined in the incubation solution, as previously described (14). LDH activity was measured spectrophotometrically at 340 nm by following the oxidation of NADH (decrease in absorbance) in the presence of pyruvate using a Thermomax microplate reader (Molecular Devices, Sunnyvale, CA).…”

Section: Methodsmentioning

confidence: 99%

“…Samples of the incubation solution were collected at t0. Analysis of glutamate in each sample was performed by HPLC with fluorimetric detection (Perkin Elmer Binary LC Pump 250 and Fluorescence Detector LC 240) following precolumn derivatization with the o-phtalaldialdehyde procedure, as previously described (14). Glutamate derivatives were separated isocratically on a reverse phase column (4.6 ϫ 150 mm, 5-mm particle diameter, Nucleosil 100-C18) using a mobile phase consisting of sodium acetate buffer (0.05 M, pH 6.5), 20% methanol and 2% tetrahydrofuran.…”

Section: Methodsmentioning

confidence: 99%

“…This involved an in vitro model using the exposure of newborn rat brain slices to OGD. This reproduces most the mechanisms of hypoxic-ischemic damage in the newborn brain, as glutamatergic excitotoxicity, cytokine release, or NO toxicity, with characteristics different from those of adult rat brain slice OGD; it is therefore considered an in vitro model of NHIE (14).…”

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confidence: 99%

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Characterization of the Neuroprotective Effect of the Cannabinoid Agonist WIN-55212 in an In Vitro Model of Hypoxic-Ischemic Brain Damage in Newborn Rats

et al. 2006

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Brain slices from 7-d-old Wistar rats were exposed to oxygen-glucose deprivation (OGD) for 30 min. OGD slices were incubated with vehicle or with the CB1/CB2 cannabinoid agonist WIN55212 (50 M), the CB1 agonist arachidonyl-2-chloroethylamide (ACEA) (50 M), or the CB2 agonist JW133 (50 M), alone or combined with the CB1 and CB2 receptor antagonist SR 141716 (50 M) or SR 144528 (50 M), respectively. Neuronal damage was assessed by histologic analysis and spectrophotometric determination of lactate dehydrogenase (LDH) efflux into the incubation medium. Additionally, medium glutamate levels were determined by highperformance liquid chromatography (HPLC) and those of tumor necrosis factor ␣ (TNF-␣) by enzyme-linked immunosorbent assay. Finally, inducible nitric oxide synthase (iNOS) and CB1/CB2 receptor expression were determined in slices homogenate by Western blot. Both CB1 and CB2 receptors were expressed in slices. OGD increased CB1 expression, cellular damage, LDH efflux, glutamate and TNF-␣ release, and inducible nitric oxide synthase (iNOS) expression; WIN55212 inhibited all these actions. SR141716 and SR144528 inhibited the effect of R(ϩ)-WIN-55212-2 (WIN), as well as the reduction of LDH efflux by ACEA and JW133, respectively. In conclusion, WIN55212 afforded robust neuroprotection in the forebrain slices exposed to OGD, by acting on glutamatergic excitotoxicity, TNF-␣ release, and iNOS expression; this neuroprotective effect seemed to be mediated by CB1 and CB2 receptors. P erinatal hypoxia-ischemia remains the single most important cause of brain injury in the newborn, leading to death or lifelong sequelae (1,2). The complexities of neonatal hypoxic-ischemic encephalopathy (NHIE) pathophysiology suggest that successful neuroprotection could be achieved only with a multitherapeutic approach (2). In recent years, interest in the neuroprotective possibilities of cannabinoids has grown (3-5). Endocannabinoids emerge as natural brain protective substances in different damaging situations (3,4); in newborn rats, enhanced levels of cannabinoids have been observed in the brain after acute excitotoxic insult (6). Exogenous cannabinoid agonists are neuroprotective in different paradigms of brain injury (7,8); they inhibit intracellular calcium influx, reduce glutamate and TNF-␣ release, decrease stimulated iNOS expression, induce hypothermia, and exert immunomodulatory and antioxidant actions (3-5,9,10). Some of these effects are dependent on the activation of the principal brain cannabinoid receptors, the CB1 receptors, and others are dependent on the molecular properties of the cannabinoid or the activation of non-CB1 receptors (3). There are few studies regarding the possible neuroprotective effect of cannabinoids in newborns. Early studies describe that the administration of anandamide (11) or the cannabinoid agonist ⌬ (9)-tetrahydrocannabinol (12) affords neuroprotection in a newborn rat model with excitotoxic neuronal injury. It has also been demonstrated that the cannabinoid agonist WIN, prevents ea...

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Nanoparticle‐microglial interaction in the ischemic brain is modulated by injury duration and treatment

Joseph

Liao

Zhang

et al. 2020

Bioengineering & Transla Med

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Cerebral ischemia is a major cause of death in both neonates and adults, and currently has no cure. Nanotechnology represents one promising area of therapeutic development for cerebral ischemia due to the ability of nanoparticles to overcome biological barriers in the brain. ex vivo injury models have emerged as a highthroughput alternative that can recapitulate disease processes and enable nanoscale probing of the brain microenvironment. In this study, we used oxygen-glucose deprivation (OGD) to model ischemic injury and studied nanoparticle interaction with microglia, resident immune cells in the brain that are of increasing interest for therapeutic delivery. By measuring cell death and glutathione production, we evaluated the effect of OGD exposure time and treatment with azithromycin (AZ) on slice health. We found a robust injury response with 0.5 hr of OGD exposure and effective treatment after immediate application of AZ. We observed an OGD-induced shift in microglial morphology toward increased heterogeneity and circularity, and a decrease in microglial number, which was reversed after treatment. OGD enhanced diffusion of polystyrene-poly(ethylene glycol) (PS-PEG) nanoparticles, improving transport and ability to reach target cells. While microglial uptake of dendrimers or quantum dots (QDs) was not enhanced after injury, internalization of PS-PEG was significantly increased. For PS-PEG, AZ treatment restored microglial uptake to normal control levels. Our results suggest that different nanoparticle platforms should be carefully screened before application and upon doing so; disease-mediated changes in the brain microenvironment can be leveraged by nanoscale drug delivery devices for enhanced cell interaction.

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Immature rat brain slices exposed to oxygen–glucose deprivation as an in vitro model of neonatal hypoxic–ischemic encephalopathy

Cited by 32 publications

References 33 publications

Tumor Necrosis Factor Receptor-1 is Essential for LPS-Induced Sensitization and Tolerance to Oxygen—Glucose Deprivation in Murine Neonatal Organotypic Hippocampal Slices

Tumor Necrosis Factor Receptor-1 is Essential for LPS-Induced Sensitization and Tolerance to Oxygen—Glucose Deprivation in Murine Neonatal Organotypic Hippocampal Slices

Characterization of the Neuroprotective Effect of the Cannabinoid Agonist WIN-55212 in an In Vitro Model of Hypoxic-Ischemic Brain Damage in Newborn Rats

Nanoparticle‐microglial interaction in the ischemic brain is modulated by injury duration and treatment

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