Under global cerebral ischemia, the effect of different brain temperature on cerebral ischemic injury was studied. Male Sprague-Dawley rats were divided into normothermic (37-38°C) ischemia, mild hypothermic (31-32°C) ischemia, hyperthermic (41-42°C) ischemia and sham-operated groups. Global cerebral ischemia was established using the Pulsinelli four-vessel occlusion model and brain temperature was maintained at defined level for 60 min after 20-min ischemia. The expression of c-fos protein and the levels of malondialdehyde (MDA) and lactate in brain regions were detected by immunochemistry and spectrophotometrical methods, respectively. C-fos positive neurons were found in the hippocampus and cerebral cortex after cerebral ischemia reperfusion. Mild hypothermia increased the expression of c-fos protein in both areas, whereas hyperthermia decreased the expression of c-fos protein in the hippocampus at 24 h reperfusion, and the cerebral cortex at 48 h reperfusion when compared to normothermic conditions. In normothermic, mild hypothermic and hyperthermic ischemia groups, the levels of MDA and lactate in brain tissue were increased at 24, 48 and 72 h reperfusion following 20-min ischemia as compared with the sham-operated group (P<0.01). The levels of MDA and lactate in mild hypothermic group were significantly lower than those in normothermic group (P<0.01). It is suggested that brain temperature influences the translation of the immunoreactive protein product of c-fos after global cerebral ischemia, and MDA and lactate are also affected by hypothermia and hyperthermia.
Animals ; Body Temperature ; Brain ; blood supply ; metabolism ; physiopathology ; Brain Ischemia ; metabolism ; physiopathology ; Cerebral Cortex ; blood supply ; metabolism ; physiopathology ; Hippocampus ; blood supply ; metabolism ; physiopathology ; Immunochemistry ; Lactic Acid ; metabolism ; Male ; Malondialdehyde ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Proto-Oncogene Proteins c-fos ; metabolism ; Rats ; Rats, Sprague-Dawley ; Reperfusion Injury ; metabolism ; physiopathology ; Spectrophotometry ; Temperature ; Time Factors ; Tumor Suppressor Protein p53 ; metabolism

OBJECTIVETo better assess the role of p38 MAPK, this project was designed to investigate whether intraventricular injection of antisense oligodeoxynucleotide (As-ODN) directed against the p38 MAPK of pyramidal neurons in hippocampus could affect the brain ischemic tolerance induced by limb ischemic preconditioning (LIP).

METHODSThe rat 4-vessel occlusion global cerebral ischemic model was used. Forty-eight male Wistar rats with permanently occlusion of the bilateral vertebral arteries were divided into 8 groups (n=6): sham, LIP, brain ischemic insult, LIP + brain ischemic insult, distilled water + LIP + brain ischemic insult, p38 MAPK As-ODN and p38 MAPK As-ODN + LIP + brain ischemic insult (two doses of 5 nmol/5 microl and 10 nmol/5 microl were used) groups. Thionin staining was used for observing histological changes of the hippocampus.

RESULTSNo significant delayed neuronal death (DND) was detected in the CA1 hippocampus of the rats that underwent sham and LIP operation. Brain ischemic insult for 8 min induced obvious DND as represented with the increase in histological grade (HG) and decrease in neuronal density (ND) significantly compared with sham and LIP groups. LIP protected the CA1 hippocampal pyramidal neurons against DND induced by global brain ischemic insult, suggesting the occurrence of brain ischemic tolerance. However, pretreatment with p38 MAPK As-ODN effectively blocked the ischemic tolerance induced by LIP in a dose dependent manner.

CONCLUSIONIt could be concluded that p38 MAPK plays an important role in the brain ischemic tolerance induced by LIP.

Animals ; Brain Ischemia ; physiopathology ; Cell Death ; Extremities ; blood supply ; Hippocampus ; pathology ; Ischemic Preconditioning ; methods ; Male ; Oligodeoxyribonucleotides, Antisense ; pharmacology ; Rats ; Rats, Wistar ; Reperfusion Injury ; prevention & control ; p38 Mitogen-Activated Protein Kinases ; metabolism ; physiology

OBJECTIVETo compare the different gene expression profiles among Qingkailing components of BA (baicalin), JA (jasminoidin), CA (cholic acid) and CM (concha margaritiferausta) in regulating hippocampus ischemia related genes of mice.

METHODThe hippocampus ischemia-reperfusion model mice were randomly divided into groups of BA, JA, CA, CM and M (model group), 15 mice for each group, and decapitated after 24 hours. Coronal brain slices were stained with TTC (2, 3, 5-Triphenylte trazolium Chloride) and the percentages of infarct volume were calculated. Meanwhile, total RNA were extracted from the hippocampus. We selected 374 genes which related to cerebral ischemia to find the different gene expression profiles among the Qingkailing components. Then T-tests was used to select different genes between BA and CM, JA and CM, CA and CM by Arraytrack software (P < 0. 05, Fold change > 1.5), and the pharmacodynamic characteristics were explored according to GO functional classification.

RESULTCompared with the model group BA, JA and CA could effectively reduce infarct size of hippocampus ischemic (P < 0.05). the numbers of significantly differentially expressed genes were 41 (24 up, 17 down) between BA and CM, 22 (13 up, 9 down) between JA and CM, and 11(8 up, 3 down) between CA and CM. All of BA, JA and CA could inhibit the expression of Myb gene.

CONCLUSIONWhen exerting its pharmacological effects, BA, JA, CA not only have common gene targets but also have diversity in pharmacological character.

Animals ; Brain Ischemia ; drug therapy ; genetics ; metabolism ; Disease Models, Animal ; Drugs, Chinese Herbal ; pharmacology ; Gene Expression Regulation ; drug effects ; Hippocampus ; blood supply ; drug effects ; metabolism ; Humans ; Male ; Mice ; Random Allocation

OBJECTIVETo investigate the expression of vascular endothelial growth factor (VEGF) and microvessel density (MVD) in hippocampus of rats with aging.

METHODSParaffin sections of brain tissue of rats at the age of 3, 18, 24, 30 months were stained by immunohistochemistry, the expression of VEGF and MVD was quantitatively analyzed.

RESULTSInnunohistochemical staining showed that the VEGF-positive cells were mainly pyramidal neuron in hippocampus; the intensity of VEGF-positivity in neuron cells was decreased with the aging (P<0.05). The MVD in hippocampus was also decreased with the aging of rats (P<0.05).

CONCLUSIONIncreasing VEGF contents and improving blood circulation in brain tissue may prevent or treat vascular dementia and cerebrovascular diseases.

Aging ; Animals ; Capillaries ; pathology ; Hippocampus ; blood supply ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley ; Vascular Endothelial Growth Factor A ; genetics ; metabolism

OBJECTIVETo investigate the effect of acupuncture serum on Ca2+ content in the cultured nervous cells of hippocampus after ischemia-reperfusion, so as to probe into humoral factors in acupuncture treatment.

METHODSThe neurons of the hippocampus from the new born rats were cultured for 9-11 days. Fluorescein-molecular probe Fluo-3 AM was used for staining of intracellular Ca2+. Fluorescent levels in the nervous cells cultured with the serum of the normal rats or the rats given electroacupuncture at "Baihui" (GV 20), "Zusanli" (ST 36), "Quchi" (LI 11) and "Sanyinjiao" (SP 6) for 2 weeks were determined by using a laser confocal microscope.

RESULTSAfter the normal serum was added, the intracellular Ca2+ fluorescent levels increased to 697 +/- 113 from 461 +/- 96, while after acupuncture serum was added, the Ca2+ fluorescent levels decreased to 584 +/- 103 from 673 +/- 108, indicating that after addition of acupuncture serum, the increased intracellular Ca2+ content could be decreased.

CONCLUSIONThere are some active substances in acupuncture serum which can obviously decrease intracellular Ca2+ content after ischemia-reperfusion, so as to provide a direct evidence for role of humoral factor in acupuncture treatment.

Acupuncture Therapy ; Animals ; Blood Proteins ; pharmacology ; Brain Ischemia ; metabolism ; therapy ; Calcium ; metabolism ; Cells, Cultured ; Hippocampus ; blood supply ; drug effects ; metabolism ; Humans ; Neurons ; drug effects ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Reperfusion Injury ; metabolism ; prevention & control ; therapy ; Serum ; chemistry

AIMTo study the protective effects and the mechanisms of sevoflurane on ischemic cerebral neurons.

METHODSWith electrophysiological microelectrode recoding technique, the OPS of hippocampal slices deprived with oxygen and glucose (OGD) and injured from toxicity of glutamate (Glu) in the control group, 2% sevoflurane group and 4% sevoflurane group were observed. The changes of ultrastructure in the three groups were also observed respectively.

RESULTSIn the control group and 2% sevoflurane group it didn't show the improvement of recovery in OPS of hippocampal slices injured from OGD and Glu. In 4% sevoflurane group the recovery degree and the recovery rate of OPS were obversely. With electricmicroscope, it was founded that in the control group and 2% sevoflurane group, the pyramidal neurons in CA1 regions deprived with glucose and oxygen and exposured by Glu were damaged. Intercellular edema were severe, the nucleus membranes were not complete, the chromatin formed mass, the endoplasmic reticulum in the cytoplasm were degenerate, mitochondrion swelled. In 4% sevoflurane group, the pyramidal neurons in CA1 regions did not swell obviously, the nucleus was clear, the nucleus membranes were complete and the mitochondria swelled lightly.

CONCLUSION4% sevoflurane could protect hippocampal neurons deprived with glucose and oxygen from the damage. The probable mechanism is 4% sevoflurane reduced the excitatory of Glu.

Anesthetics, Inhalation ; pharmacology ; Animals ; Brain Ischemia ; physiopathology ; Excitatory Amino Acid Antagonists ; pharmacology ; Hippocampus ; blood supply ; In Vitro Techniques ; Male ; Methyl Ethers ; pharmacology ; Neuroprotective Agents ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Reperfusion Injury ; prevention & control

OBJECTIVETo investigate the effect of ischemic preconditioning on chaperone hsp70 expression and protein aggregation in the CA1 neurons of rats, and to further explore its potential neuroprotective mechanism.

METHODSTwo-vesseloccluded transient global ischemia rat model was used. The rats were divided into sublethal 3-min ischemia group, lethal 10-min ischemia group and ischemic preconditioning group. Neuronal death in the CA1 region was observed by hematoxylineosin staining, and number of live neurons was assessed by cell counting under a light microscope. Immunochemistry and laser scanning confocal microscopy were used to observe the distribution of chaperone hsp70 in the CA1 neurons. Differential centrifuge was used to isolate cytosol, nucleus and protein aggregates fractions. Western blot was used to analyze the quantitative alterations of protein aggregates and inducible chaperone hsp70 in cellular fractions and in protein aggregates under different ischemic conditions.

RESULTSHistological examination showed that ischemic preconditioning significantly reduced delayed neuronal death in the hippocampus CA1 region (P < 0.01 vs 10-min ischemia group). Sublethal ischemic preconditioning induced chaperone hsp70 expression in the CA1 neurons after 24 h reperfusion following 10-min ischemia. Induced-hsp70 combined with the abnormal proteins produced during the secondary lethal 10-min ischemia and inhibited the formation of cytotoxic protein aggregates (P < 0.01 vs 10-min ischemia group).

CONCLUSIONIschemic preconditioning induced chaperone hsp70 expression and inhibited protein aggregates formation in the CA1 neurons when suffered secondary lethal ischemia, which may protect neurons from death.

Animals ; Brain Ischemia ; pathology ; Cell Count ; methods ; Cell Death ; Disease Models, Animal ; Gene Expression Regulation ; physiology ; HSP70 Heat-Shock Proteins ; metabolism ; Hippocampus ; blood supply ; metabolism ; pathology ; Ischemic Preconditioning ; Male ; Neurons ; metabolism ; Proteins ; metabolism ; Rats ; Rats, Wistar ; Time Factors

AIMTo investigate the role of extracellular-signal regulated kinase (ERK) cascade on cerebral ischemia and ischemic preconditioning in hippocampal neuron.

METHODSMale gerbils were randomly divided into sham group (SH), ischemia/reperfusion group (I/ R), ischemia preconditioning group (IP), specific antagonist of ERK-PD98059 (PD), solvent control groups (VE group), PD98059 combined with IP group (PIP). Forebrain ischemia was induced by occlusion of bilateral common carotid arteries and confirmed by isoelectricity of EEG. Observations were carried out in each group 15 min, 2 h, 4 h, 6 h, 1 d, 3 d, 5 d and 7 d after ischemia. Open field test was used to examine the spontaneous motor activity, the survival and apoptotic neurons, Fos and NF-kappaB masculine neurons in hippocampal CA1 region were counted, the expression of HSP70 in hippocampal CA1 region and p-ERK in hippocampal CA3/DG regions were detected by SABC immunocytochemical technique.

RESULTSThe spontaneous motor activity, the number of apoptotic neurons and NF-kappaB masculine neurons at 1 d, 3 d, 5 d, 7 d in CA1 region were much less in IP group than in I/R group (P < 0.01). The number of Fos masculine neurons at 15 min, 2 h, 4 h, 6 h, 1 d in CA1 region were significant more in IP group than in I/R group (P < 0.01). The expressions of p-ERK and HSP70 were significantly higher in IP group than in I/R group. The number of Fos masculine neurons at each point were more and apoptotic neurons at 1 d, 3 d were less in PD group than in I/R group. Results of observation in PIP group were within IP group and I/R group.

CONCLUSIONActivation of ERK in CA3/DG regions were related to ischemic tolerance. Induction of the expression of Fos and HSP70, decreasing of the product of NF-kB which might be one of the molecule mechanisms playing an important role in neural protection of ischemic preconditioning.

Animals ; Brain Ischemia ; metabolism ; prevention & control ; Extracellular Signal-Regulated MAP Kinases ; metabolism ; Gerbillinae ; Hippocampus ; blood supply ; cytology ; Ischemic Preconditioning ; Male ; NF-kappa B ; metabolism ; Neurons ; metabolism ; Signal Transduction ; physiology

AIMTo study the effect of different intervals between occlusions of vertebral arteries and bilateral common carotid arteries on the Pulsinelli 4-vessel occlusion global cerebral ischemic model, and the features of ischemia of the brainstem and hippocampus induced by occulusion of bilateral common carotid arteries under the condition of occlusion of unilateral vertebral artery.

METHODSEighty four adult male Wistar rats were divided into 4 groups randomly: control group, bilateral vertebral artery occluding group, global brain ischemic insult group, and unilateral vertebral artery occluding plus bilateral common carotid arteries occluding group. In the global brain ischemic insult group, rats were further divided into 24 h, 48 h, and 72 h interval subgroups according to the interval between the occlusion of the vertebral arteries and bilateral common carotid arteries. The responses including enlarging of pupils and the light reflex during the brain ischemia were observed. The duration of right reflex disappearing, the general state, and the delayed neuronal death (DND) of pyramidal neurons in the CA1 hippocampus of the rats after the brain ischemia were also observed.

RESULTSAmong the global brain ischemic insult group, both the responses and DND were more severe in 72 h interval subgroup than those in 24 h and 48 h interval subgroups. There was no significant difference between 24 h and 48 h interval subgroups. When the bilateral common carotid arteries were occluded under the condition of occlusion of unilateral vertebral artery, severe DND was observed in the CA1 hippocampus ipsilateral to the occluding vertebral artery, but no significant DND was observed in the contralateral CA1 hippocampus.

CONCLUSIONThe results suggested that the prior occlusion of the bilateral vertebral arteries during producing Pulsinelli 4-vessel occlusion global cerebral ischemic model might be a cerebral ischemic preconditioning that could protect to some extent pyramidal neurons of the hippocampus against severe ischemic insult induced by occlusion of bilateral common carotid arteries within 48 h. Moreover, There is ipsilateral predominance of blood perfusion from one side of vertebral artery to the brainstem and hippocampus, although there was Willis artery circle in rats.

Animals ; Brain Ischemia ; prevention & control ; Hippocampus ; blood supply ; Ischemic Preconditioning ; methods ; Male ; Rats ; Rats, Wistar ; Vertebral Artery ; pathology

Several studies have demonstrated that ischemic preconditioning increases superoxide dismutase activity, but it is unclear how ischemic preconditioning affects events downstream of hydrogen peroxide production during subsequent severe ischemia and reperfusion in the hippocampus. To answer this question, we investigated whether ischemic preconditioning in the hippocampal CA1 region increases the activities of antioxidant enzymes glutathione peroxidase and catalase, resulting in a decrease in the level of hydroxyl radicals during subsequent severe ischemia-reperfusion. Transient forebrain ischemia was induced by four-vessel occlusion in rats. Ischemic preconditioning for 3 min or a sham operation was performed and a 15-min severe ischemia was induced three days later. Ischemic preconditioning preserved the CA1 hippocampal neurons following severe ischemia. The concentration of 2,3-dihydroxybenzoic acid, an indicator of hydroxyl radical, was measured using in vivo microdialysis technique combined with HPLC. The ischemia-induced increase in the ratio of 2,3-dihydroxybenzoic acid concentration relative to baseline did not differ significantly between preconditioned and control groups. On the other hand, activities of the antioxidant enzymes glutathione peroxidase-1 and catalase were significantly increased at 3 days after ischemic preconditioning in the hippocampus. Our results suggest that, in preconditioned rats, while hydrogen peroxide is generated from severe ischemia, the activity of catalase and glutathione peroxidase-1 is correspondingly increased to eliminate the excessive hydrogen peroxide. However, our results show that the enhanced activity of these antioxidant enzymes in preconditioned rats is not sufficient to decrease hydroxyl radical levels during subsequent severe ischemia-reperfusion.
Animals ; Antioxidants/*metabolism ; Catalase/metabolism ; Enzyme Activation ; Glutathione Peroxidase/metabolism ; Hippocampus/*blood supply ; Hydrogen Peroxide/metabolism ; Hydroxybenzoic Acids/metabolism ; Hydroxyl Radical/*metabolism ; Ischemic Attack, Transient/*metabolism/physiopathology/prevention & control ; *Ischemic Preconditioning ; Male ; *Prosencephalon ; Rats ; Rats, Sprague-Dawley ; Reperfusion Injury/metabolism/prevention & control