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OBJECTIVE@#To improve the classical 4-vessel occlusion (4VO) model established by Pulsinelli and Brierley.@*METHODS@#Thirty-two male SD rats were randomized into sham operation group, I4VO-Con10 group, I4VO-Int10 group and I4VO-Int15 group. The sham surgery group underwent exposure of the bilateral vertebral arteries and carotid arteries without occlusion to block blood flow. The I4VO-Con10 group experienced continuous ischemia by occluding the bilateral vertebral arteries and carotid arteries for 10 minutes followed by reperfusion for 24 hours. The I4VO-Int10 and I4VO-Int15 groups were subjected to intermittent ischemia. The I4VO- Int10 group underwent 5 minutes of ischemia, followed by 5 minutes of reperfusion and another 5 minutes of ischemia, and then reperfusion for 24 hours. The I4VO-Int15 group experienced 5 minutes of ischemia followed by two cycles of 5 minutes of reperfusion and 5 minutes of ischemia, and then reperfusion for 24 hours. The regional cerebral blood flow (rCBF) was monitored with laser Doppler scanning, and survival of the rats was observed. HE staining was used to observe hippocampal pathologies to determine the optimal method for modeling. Another 48 rats were randomized into 6 groups, including a sham operation group and 5 model groups established using the optimal method. The 5 I4VO model groups were further divided based on the reperfusion time points (1, 3, 7, 14, and 28 days) into I4VO-D1, I4VO-D3, I4VO-D7, I4VO- D14, and I4VO- D28 groups. Body weight changes and survival of the rats were recorded. HE staining was used to observe morphological changes in the hippocampal, retinal and optic tract tissues. The Y-maze test and light/dark box test were used to evaluate cognitive and visual functions of the rats in I4VO-D28 group.@*RESULTS@#Occlusion for 5 min for 3 times at the interval of 5 min was the optimal method for 4VO modeling. In the latter 48 rats, the body weight was significantly lower than that of the sham-operated rats at 1, 3, 7, 14 and 28 days after modeling without significant difference in survival rate among the groups. The rats with intermittent vessel occlusion exhibited progressive deterioration of hippocampal neuronal injury and neuronal loss. Cognitive impairment was observed in the rats in I4VO-D28 group, but no obvious ischemic injury of the retina or the optic tract was detected.@*CONCLUSION@#The improved 4VO model can successfully mimic the main pathological processes of global cerebral ischemia-reperfusion injury without causing visual impairment in rats.
Subject(s)
Rats , Male , Animals , Rats, Sprague-Dawley , Brain Ischemia , Cerebral Infarction , Reperfusion Injury , Body WeightABSTRACT
ObjectiveTo investigate the effects of dopamine receptor agonist pramipexole and levodopa on emotion and cognition, and mitochondrial membrane potential of rats after global cerebral ischemia-reperfusion injury. MethodsA total of 80 male Sprague-Dawley rats were divided into sham group (n = 20), model group (n = 20), pramipexole group (n = 20) and combined group (n = 20). The latter three groups were used to prepare the model of global cerebral ischemia-reperfusion injury with Pulsinelli's four-vessel occlusion. The pramipexole group was intraperitoneally injected pramipexole 0.5 mg/kg once a day, while the combined group was injected levodopa 50 mg/kg and pramipexole 0.5 mg/kg, for 14 days. Five rats in each group were tested with open field test three, seven and 14 days after modeling; five were tested with Y-maze test seven and 14 days after modeling; five were detected mitochondrial membrane potential three, seven and 14 days after modeling; and five were observed under Nissl's staining14 days after modeling. ResultsCompared with the model group, the number of entries into the central zone (P < 0.05), total distance travelled (P < 0.05) and average velocity (P < 0.05) in the open field test increased in the pramipexole and combined groups seven and 14 days after modeling, duration spent in the central zone increased in the pramipexole and combined groups seven days after modeling (P < 0.05); the rate of spontaneous alternation of Y-maze test increased in the pramipexole and combined groups 14 days after modeling (P < 0.05); mitochondrial membrane potential in hippocampus increased in the pramipexole and combined groups seven and 14 days after modeling (P < 0.05), and it was less in the pramipexole group than in the combined group 14 days after modeling (P < 0.05); and the number of surviving neurons in the hippocampal CA1 increased in the pramipexole and combined groups 14 days after modeling (P < 0.05). ConclusionPramipexole may improve emotion and cognition of rats after global cerebral ischemia-reperfusion injury, and it may be helpful for restoring mitochondrial membrane potential as combining with levodopa.
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Objective To investigate the effect of high concentration hydrogen gas on neurons in the rat hippocampus CA1 region during global cerebral ischemic/reperfusion injury (GCIR) Methods Four-vessel occlusion was used to establish rat model with GCIR injury. One hundred and five healthy male Sprague-Dawley rats were randomly divided into sham operation group(SH group, n = 15), model group(4-VO group, n = 45) and treatment group(4-VO+H2 group,n = 45). After 72 h and 9 d reperfusion, hippocampal CA1 region pyramidal neurons in every group were detected with Nissle staining , immunohistochemical neuron-specific nuclear protein (NeuN), specific protein antibody microglial cells (Iba1) staining and the relationship of position between neurons and microglia was observed through fluorescence double staining. We used Morris water maze to test the space orientation ability and the learning and memory ability in rats after 9 d reperfusion. Results Compared with those of 4-VO group,the neurons of hippocampus CA1 region were closer to normal in 72 h and 9 d in 4-VO+H2 group and neuron form and the number of neuron survival were increased significantly (P < 0.05);immunohistochemical staining showed that the number of neuron survival in 4-VO+H 2 group was obviously higher than that in 4-VO group (P < 0.05) and the number of microglia in 4-VO group was obviously higher than that in 4-VO+H2 group (P < 0.05). Water maze experiment showed that the swimming time in quadrant Ⅳ in 4-VO+H2 group was longer than that in 4-VO group (P < 0.05). Conclusion Inhalation of high concentration hydrogen gas has prominent protective effect on neurons of rat hippocampal CA1 region during reperfusion. The mechanism may be related with inhibiting the microglia excitation and activation during GCIR.
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Aim To investigate the effects of baicalin on cognitive function in global cerebral ischemia reper-fusion rats, and the probable mechanism involved. Methods Sixty male Sprague-Dawley(SD) rats were randomly divided into Sham operation group ( S group) , global cerebral ischemia reperfusion group ( I/R group) , global cerebral ischemia reperfusion + ba-icalin treatment group ( I/RB group) , twenty in each. Model was induced via the bilateral occlusion of the common carotid arteries plus hemorrhagic hypotension. 12 h after reperfusion, rats in I/RB group were given baicalin (100 mg·kg-1 ) saline solution by intragas-tric administration twice per day for 7 days. Rats in S group and I/R group were given the corresponding dose of saline infusion at the same time. Morris water maze test was employed to detect spatial learning and memo-ry. BrdU immunohistochemistry was used to detect the proliferation of neural precursor cells ( NPCs ) in the brain. Expression of COX-2 in the brain tissue was measured by Western blot. Results Compared to I/R group, baicalin improved spatial learning and memory damage ( P nitive function in the rats with global cerebral ischemia reperfusion, which might be associated with its inhibi-tory effects on the expression of COX-2 , thereby in-creasing the proliferation of NPCs in the brain.
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Objective To investigate the effects of the FKBP51 · PHLPP · AKT signal module on the phosphorylation of Akt and hippocampal neuronal injury after the cerebral ischemia / reperfusion induced neuronal death in rat hippocampus.Methods Transient(15 min)brain ischemia was induced by the four-vessel occlusion in Sprague-Dawley rats.6 rats were used in each group.The antisense oligodeoxynucletides(AS ODN)of PHLPP2 (PH domain and leucine rich repeat protein phosphatases) was used to suppress the assembly of FKBP51 · PHLPP · Akt signal module by intracerebroventricular infusion once per day for 3 days before ischemia.After 6 hours reperfusion,interactions of PHLPP2 and FKBP51 (FK506 binding protein 5) with Akt were detected by immunoprecipitation (IP) and the phosphorylation of Akt was detected by western blot (IB).After 5 days reperfusion,rats were perfusion-fixed with paraformaldehyde and Hematoxylin-Eosin staining was used to examine the survival number of CA1 pyramidal cells of hippocampus.Results Compared to PHLPP2 MS ODN group(1.24±0.24,1.68±0.11,0.58±0.01),PHLPP2 AS ODN suppressed the assembly of the FKBP51 · PHLPP · Akt signaling module(1.06±0.01,1.04±0.13),and increased the phosphorylation of Akt(0.76±0.02) (P＜0.05).Furthermore,compared to PHLPP2 MS ODN group (20.1±2.5),the number of surviving neurons significantly increased in PHLPP2 AS ODN group(88.3±2.7)(P＜0.05).Conclusion The increasing assembly of FKBP51 · PHLPP · Akt signal module can damage CA1 pyramidal cells of hippocampus by inhibiting the phosphorylation level of Akt.
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Objective To explore the role of parecoxib in protection of learning and memory ablility in rats with global cerebral ischemia/reperfusion injury. Methods Male Wistar rats were randomly divided into a sham group (S group), a ischemia/reperfusion group (I/R group), and a ischemia/reperfusion with parecoxib group (I/R+PA group). Global cerebral ischemia was induced by Pulsinelli 4-vessel occlusion. 72 h after reperfusion, Morris water maze was used to assess spatial learning and memory ability for consecutive five days. The histological changes in the hippocampus were detected by Nissl staining. Results As compared with the I/R group, rats in S guoup and I/R+PA group had a significantly shortened escape latency (P<0.05), and had significantly longer dwelling time in the former platform quadrant and more frequent cross-platform movement in the Morris water maze test. Conclusions Parecoxib plays a role in protection of learning and memory ability in rats with global cerebral ischemia/reperfusion injury.
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Aim To investigate the effects and mecha-nism of nuclear factor-κ B inhibitor, PDTC, on global cerebral ischemia reperfusion ( GCIR ) rat hippocam-pus. Methods Forty-eight adult male Sprague-Daw-ley rats were randomly divided into one control group receiving sham operation and three experimental groups all receiving global cerebral ischemia for 20 min. In PDTC 100 mg·kg-1 group ( P100 ) and PDTC 200 mg ·kg-1 group ( P200 ) , PDTC 100 mg · kg-1 or PDTC 200 mg·kg-1 was injected ip one hour before ischemi-a respectively. Spatial learning and memory function of rats were tested using Morris water maze. HE staining was employed to observe pathological changes of hipp-ocampal neurons. Expression of COX2 was measured by Western blot, and the content of PGI2 and TXA2 in rat hippocampus was detected by enzyme-linked immu-nosorbent assay. Results A significant increase of es-cape latency was observed in GCIR group compared to the sham operation group(P<0.05). PDTC 100 mg· kg-1 and PDTC 200 mg · kg-1 significantly reduced escape latency ( P <0.05 ) and histopathological injury in CA1 region of hippocampus. PDTC 100 mg · kg-1 and PDTC 200 mg · kg-1 also reduced COX2 expres-sion, PGI2 content, TXA2 content and PGI2/TXA2 . Conclusion Pretreatment with PDTC can protect hip-pocampus from GCIR injury through inhibition of COX2 expression and PGI2/TXA2 .