| Objective: To observe the protective effect of local mild hypothermia against complete cerebral ischemia/ reperfusion (I/R) in rats and to investigate the protective effect of acivition of the mitochondria ATP-sensitive potassium channels of the underlying mechanism.Methods: Experiment protocol: The experiments were performed using 32 male Sprague-Dawley rats (weight 200～250g).The animals were housed in individual cages and allowed free access to water. Anesthesia was induced by an intraperitoneal injection of chloral hydrate (0.35 g/kg body weight), and the animals were premedicated with 0.1 mg of subcutaneously administered atropine. The SD rats were randomly divided into 4 groups (n=8 each) : (S group) sham operation :The vertebrobasilar artery and bilateral common carotid arteries will not be occlused, the temperature of the rats would not be mild hypotherature;( I/R group ) Ischemia/Reper- fusion group: The male SD rats were subjected to three-vein occlusion : the vertebrobasilar artery and bilateral common carotid arteries would be occlused for 10 min, the rats would not be local mild hypotherature;(H group) mild hypothermia+ I/R group : The vertebrobasilar artery and bilateral common carotid arteries would be occlused for 10 min, the rats would be locall mild hypotherature(32～35℃) by injecting the 10℃of sodium chloride, then the local mild hypotherature would be continue 3 hours ; ( 5-HD group ) 5-HD + mild hypothermia + I/R group: The vertebrobasilar artery and bilateral carotid arteries common will be occlused for 10 min, the rats will be locall mild hypotherature(32～35℃) by injecting the 10℃of sodium chloride, the 5-HD group that was used at the dose of 10 mg/kg was given by an intraperitoneal injection, before 30 min of cerebral ischemia. Rats of other groups were injected equal volume sodium chloride. Then the locall mild hypotherature will be continue 3 hours. The temperature was regulated by ice cubes and daylight lamp, which was modulated to an appropriate distance to the rats.To detect the change of blood gas at three time points, the blood sample was taken from femoral artery .The first point was taken at 5 min before the brain ischemia,the second was the time of 10 min after the brain ischemia, the third was the time of 10 min after the reperfusion .To detect the change of the open fild test(OPT) and the inclined plane test ( IPT) at 12 hours after reperfusion.The Rat NSE ELISA was used to detect the blood of rats. The blood samples were taken from rats at 12 hours after reperfusion. Then Animals were executed and the cortex of frontal lobe of the right brain were removed for microscopic examation of the change of pathology, and the other brain tissue was removed for ultrastrucrure examination and assaied of the content of maleic dialdehyde (MDA) with the method of Thibarbturic Acid, and assaied the activity of superoxide dismutase (SOD) with xanthine oxidase in brain tissue. The change of the left brain water content (BMC) was observed through wet dry weighing method.Result: The change of blood gas is normal at the three time points.The neuroethology of H group was better than I/R group and 5-HD group .Compared with I/R group,in S group and H group, the open fild test(OPT) was more (P<0.05) and the inclined plane test ( IPT) was less (P<0.05). Compared with H group,the I/R group and 5-HD group were less (P<0.05).The brain water content (BWC)of ischemia/reperfusion group and the level of NSE expression in the rat blood serum were higher than the sham group (P<0.05).In the mild hypothermia group,the BWC and NSE were lower than I/R group and 5-HD group (P<0.05). The activity of SOD of I/R group was lower than S group (P<0.05),In the hypothermia group ,the activity of SOD was higher than I/R group and 5-HD group(P<0.05).the concertration MDA of the cerebral tissue in I/R group was higher than S group and lower than H group (both P<0.05).The result of microscopic examation of the change of pathology:Sham group,The morphosis of nerve cell was common on the whole,and the endochy leme was plentiful andhad been dyed round or elipse.the clrromato spherite was clear;I/R group,The most of nerve cell were shrinkage and explarged the scope of injury.the tigroid body was broken down or lost;H group,the part of nerve cell were shrinke or lost .the caryoplasm and the chromatospherite were not clear;5-HD group, the part of nerve cell were shrinkage and explarged the scope of injury.The tigroid body was broken down or lost.Compared with I/R group and 5-HD group , H group had light injury.The cell nucleus and envelope were slight oedema. The number of cell nerve was not diminished.The result of transmission electron micotelecope: Ultrastructure change in various groups. Sham group , The morphosis of nerve cell was common on the whole. A little of critae and membrane of mitochondrium were fused or lost,and the rough endoplasmic reticulam was light degranulation.the Golgi's complex was common,and there were some secondary lysosome and free ribosome;I/R group, the nerve cytoplasm was height oedema and the number of cell organ was significantly decreased. The most or part of critae and membrane of mitochondrium were fused or lost,and the rough endoplasmic reticulam was heavy degranulation.the Golgi's complex was not common,and there were some free ribosome;H group , The morphosis of nerve cell was common on the whole. A part of critae and membrane of mitochondrium were fused or lost,and the rough endoplasmic reticulam was light degranulation.the Golgi's complex was common,and there were some secondary lysosome and free ribosome;5-HD group, the nerve cytoplasm was height oedema and the number of cell organ was significantly decreased. The most or part of critae and membrane of mitochondrium were fused or lost,and the rough endoplasmic reticulam was heavy degranulation.the Golgi's complex was not common,and there were some free ribosome and the secondary lysosome was decreased. Compare with I/R group and 5-HD group ,there was light injury in H group ,but compare with sham group, H group had a heavy injury,there was inditical injury between I/R group and 5-HD group.Conclusion: Local mild hypothermia has protective effect on the brain against I/R injury. Activation of mito-KATP channel is involved in the mechanism.
|
PDF Full Text Request