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Transient focal cerebral ischemia induces long-term cognitive function deficit in an experimental ischemic stroke model.

ABSTRACT: Vascular dementia ranks as the second leading cause of dementia in the United States. However, its underlying pathophysiological mechanism is not fully understood and no effective treatment is available. The purpose of the current study was to evaluate long-term cognitive deficits induced by transient middle cerebral artery occlusion (tMCAO) in rats and to investigate the underlying mechanism. Sprague-Dawley rats were subjected to tMCAO or sham surgery. Behavior tests for locomotor activity and cognitive function were conducted at 7 or 30days after stroke. Hippocampal long term potentiation (LTP) and involvement of GABAergic neurotransmission were evaluated at 30days after sham surgery or stroke. Immunohistochemistry and Western blot analyses were conducted to determine the effect of tMCAO on cell signaling in the hippocampus. Transient MCAO induced a progressive deficiency in spatial performance. At 30days after stroke, no neuron loss or synaptic marker change in the hippocampus were observed. LTP in both hippocampi was reduced at 30days after stroke. This LTP impairment was prevented by blocking GABAA receptors. In addition, ERK activity was significantly reduced in both hippocampi. In summary, we identified a progressive decline in spatial learning and memory after ischemic stroke that correlates with suppression of hippocampal LTP, elevation of GABAergic neurotransmission, and inhibition of ERK activation. Our results indicate that the attenuation of GABAergic activity or enhancement of ERK/MAPK activation in the hippocampus might be potential therapeutic approaches to prevent or attenuate cognitive impairment after ischemic stroke.

SUBMITTER: Li W

PROVIDER: S-EPMC3790570 | biostudies-literature | 2013 Nov

REPOSITORIES: biostudies-literature

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Transient focal cerebral ischemia induces long-term cognitive function deficit in an experimental ischemic stroke model.

Li Wenjun W Huang Renqi R Shetty Ritu A RA Thangthaeng Nopporn N Liu Ran R Chen Zhenglan Z Sumien Nathalie N Rutledge Margaret M Dillon Glenn H GH Yuan Fang F Forster Michael J MJ Simpkins James W JW Yang Shao-Hua SH

Neurobiology of disease 20130709

Vascular dementia ranks as the second leading cause of dementia in the United States. However, its underlying pathophysiological mechanism is not fully understood and no effective treatment is available. The purpose of the current study was to evaluate long-term cognitive deficits induced by transient middle cerebral artery occlusion (tMCAO) in rats and to investigate the underlying mechanism. Sprague-Dawley rats were subjected to tMCAO or sham surgery. Behavior tests for locomotor activity and ...[more]

PMID: 23845275

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Project description:Estrogen reduces brain injury after experimental cerebral ischemia in part through a genomic mechanism of action. Using DNA microarrays, we analyzed the genomic response of the brain to estradiol, and we identified a transcript, cocaine- and amphetamine-regulated transcript (CART), that is highly induced in the cerebral cortex by estradiol under ischemic conditions. Using in vitro and in vivo models of neural injury, we confirmed and characterized CART mRNA and protein up-regulation by estradiol in surviving neurons, and we demonstrated that i.v. administration of a rat CART peptide is protective against ischemic brain injury in vivo. We further demonstrated binding of cAMP response element (CRE)-binding protein to a CART promoter CRE site in ischemic brain and rapid activation by CART of ERK in primary cultured cortical neurons. The findings suggest that CART is an important player in estrogen-mediated neuroprotection and a potential therapeutic agent for stroke and other neurodegenerative diseases. Experiment Overall Design: affy platform; replicate samples; two treated samples, two placebo. Experiment Overall Design: MIAME compliance: All samples were run in commercial arrays from Affymetrix, using Rat Genome U34A GeneChip arrays as described in the Affymetrix web site. The sample information is described below. Likewise, the Affymetrix protocol is detailed and referenced above. These descriptions include all information currently considered under the MIAME supportive guidelines, with which the JHMI Microarray Core Facility abides in all its procedures. Experiment Overall Design: Methodology. Expression profiling using DNA microarrays (One round amplification): Experiment Overall Design: Experimental Design and Sample Information. Transient focal cerebral ischemia was induced in ovariectomized female rats with and without replacement with 17beta-estradiol by intraluminal middle cerebral artery occlusion (MCAO), as described in the manuscript. At 6 and 24 hours after 2-hour MCAO, total RNA was prepared from the ischemic and contralateral cerebral cortex and striatum between coronal levels 2 and 3 mm relative to Bregma, which represent the core of MCA territory. RNA samples from 3 animals were combined into one pooled sample and hybridized to Affymetrix RG-U34A GeneChip, as described below. Each microarray experiment was performed in duplicate using two independent pooled samples per group (6 animals per group at each time point, 2 pooled samples for each brain region per time point). Experiment Overall Design: Microarray Hybridization Protocol. Total RNA was isolated using the RNeasy Mini Kit (QIAGEN). RNA from control and experimental tissue are processed using the RNA amplification protocol described by Affymetrix (Affymetrix Expression Manual). Briefly, 5 micrograms of total RNA were used to synthesize first strand cDNA using oligonucleotide probes with 24 oligo-dT plus T7 promoter as primer (Proligo LLC, Boulder, Colorado) and the SuperScript Choice System (Invitrogen, Carlsbad, California). Following the double stranded cDNA synthesis, the product was purified by phenol-chloroform extraction, and biotinilated anti-sense cRNA was generated through in vitro transcription using the BioArray RNA High Yield Transcript Labeling kit (ENZO Life Sciences Inc., Farmingdale, New York). 15 ug of the biotinilated cRNA were fragmented at 94 C for 35 min (100mM Tris-acetate, pH 8.2, 500mM KOAc, 150mM MgOAC), and 10ug of total fragmented cRNA were hybridized to the Affymetrix U34A GeneChip arrays for 16hr at 45 C with constant rotation (60 rpm). Affymetrix Fluidics Station 450 was then used to wash and stain the Chips, removing the non-hybridized target and incubating with a streptavidin-phycoerythrin conjugate to stain the biotinilated cRNA. The staining was then amplified using goat IgG as blocking reagent and biotinilated anti-streptavidin antibody (goat), followed by a second staining step with a streptavidin-phycoerythrin conjugate. Experiment Overall Design: Quality Control. To ascertain the quality control of the total RNA from the samples, we used the Agilent Bioanalyzer, Lab on a Chip technology, and confirmed the rRNA ratios and clean run patterns of the samples. Likewise, this technology is used to confirm the quality of the RNA in the form of cRNA and fragmented cRNA. To assess the QC of the hybridization, GeneChip image, and comparison between chips, we confirmed the following parameters: scaling factor values within comparable range; low background values (50-95); high percentage of present calls (between 30 and 55%); consistent 3â/5â ratios of GAPDH as representation of housekeeping genes, and presence or absence of Bio B and C as internal spike controls.

Dataset Information

Transient focal cerebral ischemia induces long-term cognitive function deficit in an experimental ischemic stroke model.

Publications

Transient focal cerebral ischemia induces long-term cognitive function deficit in an experimental ischemic stroke model.

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