Focal Cerebral Ischemia-reperfusion In Rats Research Articles

SIRT1 / NF-κB pathway on neuronal apoptosis in rats with ischemic stroke.

Recent studies have shown that neuronal apoptosis is one of the important mechanisms leading to cerebral ischemic injury. SIRT1/NF-κB is a carrier of protein units in the body, participates in the synthesis of DNA, RNA and important neurotransmitters in the brain, and is an important nutrient necessary to maintain the normal growth and development of the nervous system and the normal function of cells. This article aims to study the effect of the SIRT1/NF-κB pathway on neuronal apoptosis in rats with ischemic stroke. Rats were divided into sham operation group, model group, SIRT1/NF-κB group, each group was given intragastric administration at different doses 3 days before the operation and after the operation, and brain tissue was taken at 24, 48, 72, 96 h after operation HE staining, TTC staining and dynamic changes of SIRT1/NF-κB expression were measured. The results of the experiment showed that the result of Nao Xintong was low, and the SIRT1/NF-κB group could reduce the volume of cerebral infarction in rats (P <0.05), and reduce the expression of SIRT1/NF-κB positive cells. The experimental results show that Naoxintong SIRT1/NF-κB has a protective effect on neuronal apoptosis after focal cerebral ischemia-reperfusion in rats.

Neuroprotective effects of Senkyunolide I against glutamate-induced cells death by attenuating JNK/caspase-3 activation and apoptosis

Glutamate-induced neurotoxicity is one of the most important pathogenic mechanisms in neurological diseases and is widely used as an in vitro model for ischemic stroke. Senkyunolide I (SEI), an active constituent derived from traditional Chinese medicine Ligusticum chuanxiong Hort. and Angelica sinensis (Oliv.) Diels, has been shown to have beneficial effects against focal cerebral ischemia-reperfusion in rats. However, the mechanisms underlying SEI-mediated neuroprotection remain not well understood. Thus, we explored the influence of SEI in glutamate-mediated injury to mouse neuroblastoma (Neuro2a) cells and determined the mechanisms involved. Neuro2a cells were treated with SEI under exposure to glutamate for 24 h. Cell viability was assessed by using WST-1 reagents, and apoptosis was evaluated using Annexin V-FITC and a PI double staining kit. The protein expression levels of p-AKT, AKT, p-GSK3β, GSK3β, p-p38, p38, p-ERK, ERK, p-JNK, JNK, Bcl-2, Bax, Bcl-xl, p-Bad, Bad, p53, and cleaved caspase-3 were determined by Western blot analysis. Glutamate significantly decreased cell viability and elevated the level of apoptosis. Treatment with SEI reversed those effects. Furthermore, the expression of p-JNK/JNK and cleaved caspase-3 were also reduced after treatment with SEI. Our findings demonstrate that SEI protected Neuro2a cells against glutamate toxicity by regulating JNK/caspase-3 pathway and apoptosis. Thus, SEI maybe a promising candidate for neuroprotection.

Effect of sevoflurane preconditioning on expression of metabotropic glutamate receptor type II during focal cerebral ischemia-reperfusion in rats

Objective To evaluate the effect of sevoflurane preconditioning on the expression of metabotropic glutamate receptor type Ⅱ (mGluRⅡ) during focal cerebral ischemia-reperfusion (I/R) in rats. Methods Forty-eight clean-grade healthy male Sprague-Dawley rats were divided into 3 groups (n=16 each) using a random number table method: sham operation group (group S), cerebral I/R group (group I/R) and sevoflurane preconditioning group (group Sev). Rats were anesthetized with 10% chloral hydrate 3 ml/kg.Focal cerebral I/R was produced by occlusion of the right middle cerebral artery for 2 h followed by 24 h reperfusion.In group Sev, 2.7% sevoflurane was inhaled for 1 h and 24 h later focal cerebral I/R was produced.At 24 h after reperfusion, neurological deficit was scored, the cerebral infarct size was determined by TTC staining, the cell apoptosis in ischemic penumbra was observed by TUNEL, IκB-α expression was detected by Western blot, and mGluRⅡexpression was determined by immunofluorescent staining.The apoptosis rate was calculated. Results Compared with group S, the neurological deficit score, cerebral infarct size and apoptosis rate were significantly increased, the expression of mGluRⅡwas up-regulated, and the expression of IκB-α was down-regulated in I/R and Sev groups (P<0.05). Compared with group I/R, the neurological deficit score, cerebral infarct size and apoptosis rate were significantly decreased, the expression of mGluRⅡwas down-regulated, and the expression of IκB-α was up-regulated in group Sev (P<0.05). Conclusion Sevoflurane preconditioning reduces focal cerebral I/R injury through inhibiting the expression of mGluRⅡ in rats. Key words: Anesthetics, inhalation; Ischemic preconditioning; Brain; Reperfusion injury; Receptors, glutamate

Effect of dexmedetomidine pretreatment on contents of glutamic acid and γ-aminobutyric acid dur-ing focal cerebral ischemia-reperfusion in rats

Objective To evaluate the effect of dexmedetomidine pretreatment on contents of glutamic acid and γ-aminobutyric acid during focal cerebral ischemia-reperfusion (I/R) in rats. Methods Thirty-six clean-grade healthy male Wistar rats, aged 10 months, weighing 250-300 g, were divided into 3 groups using a random number table method: sham operation group (group S), I/R group, and dexmedetomidine group (group D). Focal cerebral I/R was induced by occlusion of the left middle cerebral artery for 30 min followed by reperfusion in anesthetized rats.The left middle cerebral artery was only isolated but not occluded in group S. Sterile normal saline 1 ml was intraperitoneally injected, and 30 min later the model of focal cerebral I/R was established in group I/R.Rats were sacrificed at the end of reperfusion, and brains were removed for determination of contents of glutamic acid and γ-aminobutyric acid in brain tissues and for examination of ultrastructure (with an electron microscope). Results Compared with group S, the content of glutamic acid was significantly increased, and the content of γ-aminobutyric acid was decreased in I/R and D groups (P<0.05). Compared with group I/R, the content of glutamic acid was significantly decreased, and the content of γ-aminobutyric acid was increased in group D (P<0.05). The damage to the ultrastructure of brain tissues was significantly attenuated in group D when compared with group I/R. Conclusion Dexmedetomidine pretreatment can reduce focal cerebral I/R injury through decreasing the content of glutamic acid and increasing the content of γ-aminobutyric acid in rats. Key words: Dexmedetomidine; Brain; Reperfusion injury; Glutamic acid; gamma-Aminobutyric acid

Effects of chrysin and early exercise on NF-κB signaling pathway during lung injury induced by focal cerebral ischemia-reperfusion in rats

Inflammation and oxidative stress play an integral role in the pathogenesis of cerebral ischemia. Chrysin exhibits anti-inflammatory and neuroprotective effects. Early exercise is an effective strategy for stroke treatment. However, the signaling pathways that link these events are not fully understood. We investigated the effects signaling pathways of chrysin treatment with early exercise in a rat model of lung injury induced by transient focal cerebral ischemia. Forty female healthy SD rats were divided into four s randomly ( n = 10): sham operation (A), lung injury induced by ischemia-reperfusion (B), chrysin treatment (C), and chrysin + early exercise (D). In C, rats were treated intragastrically with chrysin every day at a dose of 20 mg/kg while rats were treated intragastrically with the same volume of NS in A and B. In D, rats received forced treadmill training and were treated intragastrically with chrysin (20 mg/kg). Fourteen days later, all rats were sacrificed. Brain and lung tissues were obtained for pathological analysis, lung water content and lung function were assessed. Meanwhile, qRT–PCR and western blotting were employed to detect the expression of NF-κB and TNF–α in lung. Neurobehavioral deficits and lung injury could be seen in B. Compared with B, chrysin (C and D) successfully decreased the infarct size, relieved pulmonary edema and improves lung function, while downregulated expression of NF-κB and TNF–α. Notably, neurobehavioral deficits and lung injury were significantly improved in D, as well as the decreased expression of NF-κB and TNF–α. Chrysin treatment with early exercise improved functional outcomes and abrogated the lung injury induced by focal cerebral ischemia-reperfusion on rats, which might be associated with inhibition of NF-κB-mediated neuroinflammation. Chrysin combined with early exercise may be used as a potential treatment in patients at high risk of lung injury induced by ischemic stroke.

Effect of hydrogen on mitochondrial membrane potential during focal cerebral ischemia-reperfusion in rats

Objective To investigate the effect of hydrogen on mitochondrial membrane potential during focal cerebral ischemia-reperfusion (I/R) in rats. Methods Eighty-four healthy adult male SPF Sprague-Dawley rats, weighing 220-240 g, were divided into 3 groups (n=28 each) using a random number table: sham operation group (Sham group), I/R group and hydrogen group (H2 group). Focal cerebral I/R was produced by mid-cerebral artery occlusion in I/R and H2 groups.Hydrogen 10 ml/kg was intraperitoneally injected immediately after onset of reperfusion and at 12 h of reperfusion in group H2.Neurological deficit was scored at 24 h of reperfusion.The rats were then sacrificed and brain tissues in cortex were removed for microscopic examination of the pathological changes and for determination of cerebral infarct size (by TTC staining), nerve cell apoptosis (by TUNEL), mitochondrial membrane potential(by JC-1 staining) and expression of Bcl-2 and Bax (by Western blot). Apoptotic index (AI) was calculated. Results Compared with Sham group, the neurological deficit score, percentage of cerebral infarct size and AI were significantly increased, the mitochondrial membrane potential was decreased, Bax expression in brain tissues was up-regulated, and Bcl-2 expression in brain tissues was down-regulated in I/R and H2 groups (P<0.05). Compared with I/R group, the neurological deficit score, percentage of cerebral infarct size and AI were significantly decreased, the mitochondrial membrane potential was increased, Bax expression in brain tissues was down-regulated, and Bcl-2 expression in brain tissues was up-regulated (P<0.05), and the pathological changes of brain tissues were significantly attenuated in H2 group. Conclusion The mechanism by which hydrogen reduces focal cerebral I/R injury is related to decreasing dissipation of mitochondrial membrane potential and inhibiting nerve cell apoptosis in rats. Key words: Hydrogen; Reperfusion injury; Brain; Membrane potential, mitochondrial

Neurochemical changes in unilateral cerebral hemisphere during the subacute stage of focal cerebral ischemia-reperfusion in rats: An ex vivo 1H magnetic resonance spectroscopy study

Understanding the subacute may shed light on the mechanism of cerebral ischemia. The present study aimed to explore metabolic features underlying subacute stage of ischemia-reperfusion injury and developing effective treatments. Rats were divided into three groups: the permanent middle cerebral artery occlusion (pMCAO), transient cerebral focal ischemia (tMCAO) and sham group. Evaluation of animal models was performed by the neurological deficit, MR images and pathological morphological abnormality. To elucidate metabolic changes, we conducted a comparative analysis of metabolic composition of unilateral brain tissue using 1H nuclear magnetic resonance spectroscopy. The successful model was observed low signal on T1WI and high signal on T2WI lesions in the left cerebral. Histopathological results confirmed the formation of apparent lesions in the left striatum, hippocampus CA1 and cortex tissues of subacute cerebral ischemia rats and showed that rats with focal cerebral ischemia-reperfusion could alleviate the extent of pathological damage degree. In pMCAO rats 7 days after surgery, decreased levels of N-acetyl aspartate (NAA), γ-aminobutyric acid (GABA), glutamate (Glu) and succinate (Suc) concomitantly with increased levels of glutamine (Gln), myo-inositol (m-Ins) and lactate (Lac) were observed compared to the control. Whereas, increased level of Lac with decreased levels of NAA, GABA, Glu, Suc, creatine (Cre) were observed in the tMCAO rats. This demonstrated that experimental subacute ischemic stroke in rats caused extensive perturbation in energy metabolism, the tricarboxylic acid cycle and GABA shunt, which provided essential information for understanding the pathogenesis of subacute cerebral ischemia-reperfusion and provided guidance in choosing the suitable therapeutic schedule.

Thrombopoietin could protect cerebral tissue against ischemia-reperfusion injury by suppressing NF-κB and MMP-9 expression in rats

Objective: To determine the neuroprotective effects and underpinning mechanisms of thrombopoietin (TPO), Matrix Metalloproteinase-9(MMP-9) and Nuclear Factor-κB (NF-κB) after focal cerebral ischemia-reperfusion in rats.Methods: Male rats underwent 2 hours of right middle cerebral artery occlusion (MCAO) followed by 22 hours of reperfusion. PBS or TPO (0.1ug/kg) was administered from caudal vein before reperfusion. Neurologic deficits, brain edema, Evans blue (EB) extravasation, NF-κB and MMP-9 expression were subsequently examined.Results: Ischemia-reperfusion injury produced a large area of edema. TPO significantly reduced edema and alleviated neurologic deficits after ischemia-reperfusion. Ischemia-induced increases of NF-κB, MMP-9 and Evans blue extravasation were reduced by TPO intervention.Conclusion: TPO improved neurological function and ameliorated brain edema after stroke, partly by reducing the ischemia-induced increase of NF-κB and MMP-9.

Role of PI3K/Akt/GSK-3β signaling pathway in hydrogen-induced inhibition of neuronal apoptosis induced by focal cerebral ischemia-reperfusion in rats

Objective To investigate the role of phosphatidylinositol 3-kinase/protein kinase B/glycogen synthase kinase-3β (PI3K/Akt/GSK-3β) signaling pathway in hydrogen-induced inhibition of neuronal apoptosis induced by focal cerebral ischemia-reperfusion (I/R)in rats. Methods One hundred and twenty healthy adult male Sprague-Dawley rats, weighing 200-220 g, were divided into 5 groups (n=24 each) using a random number table: sham operation group (group S); cerebral I/R group (group I/R); hydrogen group (group H2); LY294002 (specific PI3K inhibitor) group (group LY); LY294002+ hydrogen group (group LY+ H2). Focal cerebral I/R was induced by occlusion of the middle cerebral artery for 1 h followed by 24 h reperfusion.In H2 and H2+ LY groups, the animals inhaled 67% H2+ 33% O2 for 2 h starting from onset of reperfusion, and then inhaled H2 for 2 h every 6 h. In LY and LY+ H2 groups, LY294002 (10 mmol/L) 10 μl was injected into the lateral cerebral ventricle at 10 min before reperfusion.Neurologic deficit was evaluated and scored (NDS) at 24 h of reperfusion.The rats were then sacrificed, and the brains were removed for measurement of the cerebral infarct size (by TTC staining) and apoptosis in cortical neurons (by TUNEL) and for determination of the expression of Akt in the ischemic cerebral cortex, phosphorylated Akt (p-Akt), GSK-3β and phosphorylated GSK-3β (p-GSK-3β) and Bcl-2 and Bax positive cell count in the ischemic cerebral cortex (by immuno-histochemistry). The apoptosis index (AI), p-Akt/Akt ratio and p-GSK-3β/GSK3-β ratio were calculated. Results Compared with group S, the NDS, cerebral infarct size, AI, p-Akt/Akt ratio, p-GSK-3β/GSK-3β ratio and Bax positive cell count were significantly increased, and the Bcl-2 positive cell count was significantly decreased in group I/R (P 0.05). Compared with group H2, the NDS, cerebral infarct size, AI and Bax positive cell count were significantly increased, and the p-Akt/Akt ratio, p-GSK-3β/GSK-3β ratio and Bcl-2 positive cell count were significantly decreased in group LY+ H2(P<0.05). Conclusion The mechanism by which hydrogen inhibits focal cerebral I/R-induced neuronal apoptosis is associated with the activation of PI3K/Akt/GSK-3β signaling pathway in rats. Key words: Hydrogen; 1-Phosphatidylinositol 3-kinase; Protein-serine-threonine kinases; Glycogen synthase kinase 3; Apoptosis; Reperfusion injury; Brain

Diosgenin attenuates the brain injury induced by transient focal cerebral ischemia-reperfusion in rats

The aim of the present study is to explore the potential cerebroprotection of diosgenin against the transient focal cerebral ischemia-reperfusion (I/R) injury and its possible underlying mechanisms. The diosgenin at two dose levels, namely 100 and 200mgkg−1, was intragastrically administrated once daily for 7-day period prior to the surgery. Then, the rats were subjected to middle cerebral artery occlusion (MCAO) using the intraluminal thread for 90min. After 24h reperfusion, several diagnostic indicators were evaluated and all animals were sacrificed to harvest their brains and blood for subsequent biochemical analyses. The results indicated that diosgenin treatment significantly inhibited the death rate and improved the impaired neurological functions along with neurological deficit scores and cerebral infarct size as compared with the rats exposed to I/R insult without agents administration. The increase in the number of apoptotic cells determined by TUNEL in the hippocampus CA1 and cortex was also apparently attenuated in the diosgenin treatment group, which was closely correlated with suppression of Caspase-3 activity and Bax/Bcl-2 ratio. In addition the elevated concentrations of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6) in blood serum of the I/R treated rats were reduced almost to their normal level. Further results obtained from the Western blotting analysis revealed that the protein expression of IκBα in the injured brain was up-regulated, while the p65 subunit of NF-κB was down-regulated in nucleus after the treatment. Collectively, this neuroprotection of diosgenin against I/R injury may be attained through its anti-apoptosis, anti-inflammation and intervening the NF-κB signal pathway properties. Due to the satisfactory findings, diosgenin might be a powerful therapeutic agent to combat the similar disease in future clinic.

Effect of dexmedetomidine on expression of iNOS and COX-2 during focal cerebral ischemia-reperfusion in rats

Objective To evaluate the effect of dexmedetomidine on the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) during focal cerebral ischemia-reperfusion (I/R) in rats. Methods Thirty pathogen-free male Sprague-Dawley rats, aged 12-16 months, weighing 300-360 g, were randomly divided into 3 groups (n =10 each) using a random number table: sham operation group (group Sham), focal cerebral I/R group (group I/R), and dexmedetomidine group (group D). Focal cerebral I/R was induced by occlusion of the right middle cerebral artery.In group D, dexmedetomidine was given as a loading dose of 1 μg/kg (over 10 min) starting from 1 h of ischemia, followed by an infusion of 0.05 μg · kg-1 · h-1 until 2 h of reperfusion.Neurological deficit was assessed and scored at 24 h of reperfusion, and then the rats were sacrificed.Brains were removed for determination of cerebral infarct size and expression of iNOS and COX-2 in the hippocampus (by Western blot). The percentage of cerebral infarct size was calculated. Results Compared with group Sham, the neurological deficit score, percentage of head swing to the left, percentage of cerebral infarct size, and expression of iNOS and COX-2 in the hippocampus were significantly increased in I/R and D groups (P<0.05). Compared with group I/R, the neurological deficit score, percentage of head swing to the left, percentage of cerebral infarct size, and expression of iNOS and COX-2 in the hippocampus were significantly decreased in group D (P<0.05). Conclusion The mechanism by which dexmedetomidine mitigates focal cerebral I/R injury may be related to inhibition of iNOS and COX-2 expression in rats. Key words: Dexmedetomidine; Reperfusion injury; Brain; Nitric oxide synthase; Cyclooxygenase 2

Effect of exogenous hydrogen sulfide on neuronal apoptosis during focal cerebral ischemia-reperfusion in rats

Objective To investigate the effect of exogenous hydrogen sulfide (H2S) on neuronal apoptosis during focal cerebral ischemia-reperfusion (I/R) in rats. Methods One hundred eight male Sprague-Dawley rats, aged 6-8 weeks, weighing 250-270 g, were randomly divided into 3 groups (n = 36 each) using a random number table: control group (group C), I/R group and H2S group.Focal cerebral ischemia was induced by middle cerebral artery occlusion in anesthetized rats.A nylon thread with a rounded tip was inserted into the left internal carotid artery and advanced intracranially to block blood flow into the middle cerebral artery.Middle cerebral artery occlusion was maintained for 90 min followed by reperfusion.In group H2S, 0.25% NaSH (a donor of exogenous H2S) 10 mg/kg was injected intraperitoneally at the onset of reperfusion.The equal volume of normal saline was given in C and I/R groups.At 1, 3 and 7 days of reperfusion, neurological deficit was scored, and corner test was performed.Brains were removed for determination of myocardial infarct size, Bax-, Bcl-2- and caspase-3-positive cells, and cell apoptosis.The percentage of myocardial infarct size, rate of Bax-, Bcl-2-and caspase-3-positive cells and apoptosis rate were calculated. Results Compared with group C, the neurological deficit score was significantly decreased, and the corner score, percentage of myocardial infarct size, rate of Bax-, Bcl-2- and caspase-3-positive cells and apoptosis rate were increased at each time point of reperfusion in I/R and H2S groups (P<0.05). Compared with group I/R, the neurological deficit score and Bcl-2-positive cells were significantly increased, and the corner score, percentage of myocardial infarct size, rate of Bax- and caspase-3-positive cells and apoptosis rate were decreased at each time point of reperfusion in group H2S (P<0.05). Conclusion The mechanism by which exogenous H2S attenuates focal cerebral I/R is related to inhibition of neuronal apoptosis in rats. Key words: Hydrogen sulfide; Reperfusion injury; Brain; Apoptosis

Influence of edaravone on growth arrest and DNA damage-inducible protein 34 expression following focal cerebral ischemia-reperfusion in rats

ABSTRACT Objective To investigate the influence of edaravone on the expression of growth arrest and DNA damage-inducible protein 34 (GADD34). Methods A total of 108 healthy male Sprague-Dawley rats were randomly divided into sham operation group, model group and edaravone group (36 cases for each group). Transient focal cerebral ischemia was induced by middle cerebral artery occlusion for 2 h followed by reperfusion in Sprague-Dawley rats. Then, GADD34 expression was measured with immunohistochemistry at different time-points after reperfusion in the peri-infarct regions of all rats. Results The GADD34 expression was detected in the peri-infarct regions of rats 1 h after reperfusion, which reached its peak 24 h after reperfusion. And edaravone could significantly down-regulate the GADD34 expression. Conclusions Edaravon could down-regulate GADD34 expression, which suggests that edaravone may exert an important function in inhibiting endoplasmic reticulum stress reaction by scavenging free radicals in the upper stream.

Effect of sevoflurane postconditioning on oxidative stress responses during focal cerebral ischemia-reperfusion in rats

Objective To investigate the effect of sevoflurane postconditioning on oxidative stress responses during focal cerebral ischemia-reperfusion (I/R) in rats.Methods Twenty-four male Wistar rats,weighing 240-280 g,were randomly assigned into 3 groups:sham operation group (group S),focal cerebral I/R group (group I/R) and sevoflurane postconditioning group (group SP).The animals were anesthetized with intraperitoneal 10％ chloral hydrate 350 mg/kg.Focal cerebral I/R was produced by middle cerebral artery occlusion.In group SP,3.9％ sevoflurane (1.5 MAC) was inhaled starting from 20 min before reperfusion until 10 min after reperfusion.While 100％ O2 and air were given instead of sevoflurane in groups I/R and S,respectively.Six rats chosen from each group at 24 h of reperfusion were sacrificed and brains were removed for determination of malondialdehyde (MDA),glutathione (GSH),superoxide dismutase (SOD),catalase (CAT),glutathione peroxidase (GSH-Px) and glutathione reductase (GR) levels and for microscopic examination.The cerebral infarct size was measured by TTC staining.Results Compared with group S,MDA level and cerebral infarct size were significantly increased in groups I/R and SP,and GSH,SOD,CAT,GSH-Px and GR levels were decreased in group I/R,and GSH-Px level was decreased in group SP (P ＜ 0.05).Compared with group I/R,cerebral infarct size and MDA level were decreased,and GSH,SOD,CAT,GSH-Px and GR levels were decreased in group SP (P ＜ 0.05).The pathological changes were significantly attenuated in group SP compared with group I/R.Conclusion The mechanism by which sevoflurane postconditioning mitigates focal cerebral I/R injury in rats is related to enhanced antioxidase activity and inhibition of oxidative stress responses. Key words: Anesthetics, inhalation ; Oxidative stress ; Reperfusion injury ; Brain

Effect of parecoxib pretreatment on expression of aquaporin-4 during focal cerebral ischemia-reperfusion in rats

Objective To investigate the effect of parecoxib pretreatment on the expression of aquaporin-4 (AQP4) during focal cerebral ischemia-reperfusion (I/R) in rats.Methods One hundred and twenty-eight adult male Sprague-Dawley rats,aged 6-8 weeks,weighing 230-280 g,were randomly divided into 4 groups (n =32 each):sham operation group (S group),I/R group,parecoxib 5 mg/kg group (group P5) and parecoxib 10 mg/kg group (group P10).The rats were anesthetized with 10％ chloral hydrate 3.5 ml/kg.Focal cerebral I/R was induced by 2 h middle cerebral artery occlusion followed by reperfusion.Parecoxib 5 and 10 mg/kg were injected via the right internal jugular vein at 30 min before middle cerebral artery occlusion in groups P5 and P10,respectively.The equal volume of normal saline was injected instead of parecoxib in groups S and I/R.Neurological deficit score (NDS) was assessed at 2 and 24 h of reperfusion.The rats were then sacrificed and brains were removed for determination of infract volume (by TTC staining).Brain water content was measured by wet/dry brain weight ratio.Their brains were cut into sections which were stained with haematoxylin and eosin and examined under microscope.The expression of AQP4 in brain tissues was measured.Results Compared with S group,NDS and brain water content were significandy increased,the infarct volume was enlarged,and the expression of AQP4 in brain tissues was up-regulated in I/R group (P ＜ 0.05 or 0.01).NDS and brain water content were significantly lower,the infarct volume was smaller,and the expression of AQP4 in brain tissues was lower in groups P5 and P10 than in I/R group (P ＜ 0.05 or 0.01).Microscopic examination showed that brain injury was significantly attenuated in groups P5 and P10 as compared with I/R group.Conclusion The mechanism by which parecoxib pretreatment alleviates the focal cerebral I/R injury in rats is related to down-regulation of the expression of AQP4. Key words: Cyclooxygenase inhibitors; Reperfusion injury; Brain; Aquaporin 4

Remote ischemic preconditioning improves spatial learning and memory ability after focal cerebral ischemia-reperfusion in rats

Using a rodent model of middle cerebral artery occlusion (MCAO), we investigated the effect of remote ischemic preconditioning (RIPC) on spatial learning and memory ability after focal cerebral ischemia-reperfusion (I/R). Thirty-six male Sprague-Dawley rats were randomly allocated into 3 groups (n=12, each): sham group, rats were subjected to sham operations without MCAO and RIPC; I/R group, rats were subjected to 1h of MCAO followed by 3 days of reperfusion; I/R+RIPC group, rats were subjected to RIPC and 1h MCAO followed by 3 days of reperfusion. The spatial learning and memory ability of the rats was measured by the Morris water maze task.The activity of cholineacetyltransferase (ChAT) in the hippocampus CA1 region was observed by an immunohistochemistry method. In the Morris water maze task, MCAO elicited a significant decrease of the ability of spatial learning and memory in contrast to the sham group. Meanwhile, RIPC induced a significantly shorter prolongation of the escape latency (p<0.05); greater number passing through the platform (p<0.05) and less time for exploring the target quadrant (p<0.05) as compared with values for the I/R group. Furthermore, the number of ChAT positive cells in the CA1 region in the I/R+RIPC group was strikingly more than those of the I/R group (p<0.05). RIPC could improve the ability of spatial learning and memory after focal cerebral ischemia-reperfusion probably due to its protection of the cholinergic neurons in the hippocampal CA1 region.

English

Caspase-3 is a key enzyme to execute apoptosis and may be cause internucleosomal DNA fragmentation in ischemic neurons. However, whether caspase-3 inhibitor can directly inhibit caspase-3&ndash;dependent deoxyribonuclease activity and prevent neuronal apoptosis following cerebral ischemic is unknown. In this study, we detected the caspase-3 and CAD protein, as well as the frequencies of neuronal apoptosis in both model control group (DMSO injection) and treatment group (Ac-DEVD-CHO injection) after focal cerebral ischemia-reperfusion in rats. Our data showed that caspase-3 and CAD protein were detectable, and apoptotic-like neuronal death occurred following cerebral ischemic in both groups. However, these results obtained were inhibited by Ac-DEVD-CHO in treatment group as compared with model control group. Taken together, these data further support that the pathway of caspase-3&ndash;dependent CAD activity and neuronal apoptosis is an important mechanism in ischemic neuronal injury, and Ac-DEVD-CHO has the neuroprotective effect to a certain degree. &nbsp; Key words:&nbsp;Caspase-3, caspase-activated deoxyribonuclease, Ac-DEVD-CHO, apoptosis, cerebral ischemia.

Effects of emulsified isoflurane preconditioning on expression of platelet-activating factor and its receptor in brain during focal cerebral ischemia-reperfusion in rats

Objective To investigate the effects of emulsified isoflurane preconditioning on the expression of platelet-activating factor (PAF) and PAF receptor during focal cerebral ischemia-reperfusion (I/R) in rats.Methods Thirty-two healthy adult male SD rats weighing 250-300 g were randomly divided into 4 groups ( n =8each):group sham operation (group S); group I/R; group emulsified isoflurane preconditioning( group EI) and group lipid emulsion (group LE).Focal cerebral I/R was induced by 2 h occlusion of middle cerebral artery followed by reperfusion in groups I/R,EI and LE.8％ emulsified isoflurane 10.5 ml/kg and 30％ lipid emulsion 10.5 ml/kg were injected intraperitoneally at 24 h before cerebral ischemia in groups EI and LE respectively.The neurologic deficit score (NDS) (0 =no deficit,4 =unable to control) was evaluated at 12 h of reperfusion.Venous blood samples were collected for measurement of plasma PAF concentration.The animals were then sacrificed and their brains removed for determination of infarct size (by TTC staining) and PAF receptor expression in hippocampus and cerebral cortex (by Western blot).Results Focal cerebral I/R significantly increased NDS,the infarct size,plasma PAF concentration and PAF receptor expression in cerebral cortex and hippocampus in group I/R as compared with group S.Emulsified isoflurane preconditioning significantly attenuated the focal cerebral I/R induced above changes in group EI as compared with group I/R,but there was no significant difference between groups I/R and LE.Conclusion Emulsified isoflurane preconditioning can attenuate focal cerebral I/R injury by inhibiting PAF and PAF receptor expression. Key words: Isoflurane; Fat emulsions, intravenous; Reperfusion injury; Platelet activating factor; Receptors, cell surface; Brajn

Effect of emulsified isoflurane preconditioning on adenosine A1 receptor expression in hippocampus during focal cerebral ischemia-reperfusion in rats

目的 探讨乳化异氟醚预处理对大鼠局灶性脑缺血再灌注时海马腺苷A1(AA1)受体表达的影响.方法 健康雄性SD大鼠88只,体重280～320 g,采用随机数字表法,将其随机分为7组:假手术组(S组,n=16)、缺血再灌注组(I/R组,n=16)、乳化异氟醚预处理组(EI组,n=16)、乳化异氟醚溶剂组(LE组,n=16)、乳化异氟醚+AA1受体拮抗剂8-环戊基-1,3-二丙基黄嘌呤(DPCPX)组(ED组,n=8)、DPCPX组(DP组,n=8)和DPCPX溶剂二甲基亚砜组(DM组,n=8).采用大脑中动脉阻塞法制备局灶性脑缺血再灌注损伤模型.于缺血前24h,I/R组、EI组和LE组分别腹腔注射生理盐水、8％乳化异氟醚和脂肪乳剂10.5 ml/kg,DP组和DM组分别腹腔注射DPCPX 1 mg/kg和二甲基亚砜2.5mg/kg,S组于相应时点腹腔注射生理盐水10.5 ml/kg; ED组于腹腔注射8％乳化异氟醚前30 min腹腔注射DPCPX 1 mg/kg.各组于再灌注24h时取8只大鼠,进行神经功能缺陷评分,然后处死,取脑组织,测定脑梗死体积；S组、I/R组、EI组和LE组于再灌注24 h时处死8只大鼠,取脑组织,采用Western blot法检测海马AA1受体的表达水平.结果 与S组比较,其余各组神经功能缺陷评分和脑梗死体积百分比升高,I/R组和LE组海马AA1受体表达下调,EI组海马AA1受体表达上调(P＜0.01)；与I/R组比较,EI组神经功能缺陷评分和脑梗死体积百分比降低,海马AA1受体表达上调(P＜0.05或0.01)；与EI组比较,LE组、ED组、DP组和DM组神经功能缺陷评分和脑梗死体积百分比升高(P＜0.05或0.01).结论 乳化异氟醚预处理减轻大鼠局灶性脑缺血再灌注损伤的机制可能与上调海马AA1受体表达有关。

Regulation of BK channel on intracellular Ca2 + level and neuronal cells apoptosis after focal cerebral ischemia-reperfusion in rats

Objective To investigate the regulation of BK channel on intracelullar free calcium concentration[Ca2 +]i and apoptosis in neuronal cells in the rat brain following focalcerebral ischemia injury. Methods 108 SD rats were randomly assigned to sham group (n =36) , ischemic reperfusion group ( n =36), IBTX group ( n =36), compared neurological function defect and infarct size of different groups at the same reperfusing time. Laser scanning confocal microscope imaging of the calcium fluorescent indicator dye Fluo-2/AM was used to determine the changes of[Ca2+]i in neurons, the expression of BK tunnel was investigated by immunohistochemistry and neuronal cells apoptosis were measured by TUNEL. Results At 24 h after ischemic reperfusion, the neurological deficit scores in IBTX group were (2. 17 ± 0. 44),which obviously higher than ischemic reperfusion group ( 1.83 ± 0. 42, P ＜ 0.05 ) ;The brain infarct volume in IBTX group was (27. 97 ± 5.84 ) ％, which larger than ischemic reperfusion group ( 22. 83 ± 4. 74 ) ％( P ＜ 0. 05 ). The[Ca2 +]i and the cell apoptosis rate of IBTX group had significant difference at 24 h after ischemic reperfusion compared with ischemic reperfusion group: (914. 50 ± 86. 57 ) nmol/L VS (732. 09 ±51.30) nmol/L (P＜0. 01); (15.20±6. 11)％ vs (10.49±1.91)％ (P＜0.05).The results of immunohistochemistry indicate that the expression of BK channel in ischemic reperfusion group decreased, but the defference of three groups was not different significantly (P ＞ 0. 05). Conclusion BK channels could play a role in protecting the neuronal cells in mice following cerebral ischemia-reperfusion by inhibited the ascension of[Ca2+]i and decrease neurons apotosis. Key words: BK channel ; Cerebral ischemia-reperfusion; Calcium overload; Neurons apoptosis