电针刺激百会及大椎穴对新生鼠缺氧缺血性脑损伤的保护

背景:中国传统医学针灸治疗缺血性脑损伤可提高脑的抗损伤能力并加快损伤修复.目的:观察百会、大椎两穴同时给予电针刺激后大鼠脑源性神经生长因子及胆碱乙酰转移酶的表达水平,探讨电针对缺氧缺血性脑损伤的保护作用.设计:随机对照实验. 单位:郑州师范高等专科学校生命科学系.材料:实验于2000-06/2001-09在郑州大学基础医学院人体解剖教研室完成.实验选取出生后7 d的清洁级Wistar大鼠50只,随机分为假手术组10只、模型对照组20只、电针治疗组20只.自制常压缺氧舱,40 cm×50 cm×60 cm,有两个2 em×2 cm的小孔与外界相通,钠石灰吸收舱内水分和CO2.方法:①模型建立及电针刺激:假手术组不造模,模型对照组及电针治疗组建立缺氧缺血模型.造模后两组大鼠在室温中恢复1～4 h后,进行低氧处理,即置于恒温37℃的常压缺氧舱内,以1.5 L/min的速度通人8 mL/L O2,920 mL/L混合气体,2 h后将动物取出,返回母鼠身边继续哺乳喂养.电针治疗组从造模后第2天开始,用一寸毫针,沿皮斜刺百会穴(顶骨正中)及大椎穴(第7颈椎与第1胸椎间,背部正中),两穴同时给予电针刺激,施以连续波,频率16Hz,强度10V,留针10 min,1次/d,10 d为1个疗程,共两个疗程,两个疗程间隔2 d.模型对照组动物造模后未进行任何治疗.②细胞记数:模型对照组及电针治疗组大鼠于缺氧缺血后22 d给予麻醉处死,取左侧脑组织制作石蜡切片,光镜下对各组切片的免疫阳性细胞进行记数.评估脑神经细胞功能状态,选择海马区进行胆碱乙酰转移酶阳性细胞数计数,每张脑片随机选取5个视野,求出阳性细胞的算术平均值.评估神经组织损伤修复作用,选择皮质和海马区脑源性神经生长因子阳性细胞数计数,方法同上.主要观察指标:新生大鼠电刺激后脑组织胆碱乙酰转移酶和脑源性神经生长因子免疫阳性细胞的表达.结果:①脑海马区胆碱乙酰转移酶免疫阳性细胞的表达:与假手术组比较,模型对照组明显降低,而电针治疗组则无明显变化(24.46±8.24),(13.96±7.62),(25.54±5.05)个/视野,P＜0.05,P＞0.05];电针治疗组高于模型对照组(P＜0.05).②脑皮质和海马区脑源性神经生长因子免疫阳性细胞的表达:与假手术组比较,模型对照组和电针治疗组均明显升高(14.14±6.11),(24.49±8.31),(31.35±9.92)个/视野,P均＜0.05;(13.42±5.56),(21.93±5.12),(27.63±7.15)个/视野,P均＜0.05];电针治疗组高于模型对照组(P＜0.05).结论:电针刺激使缺氧缺血动物中枢胆碱能神经系统处于积极活动状态,使脑源性神经营养因子数量上升增进缺氧缺血动物的神经修复功能.

Protection of electroacupuncture at Baihui and Dazhui on neonatal rats with hypoxia-ischemic brain injury

BACKGROUND: Acupuncture in Chinese traditional medicine improves capacity of brain on resisting injury and accelerates injury repair in treatment of ischemic brain injury.OBJECTIVE: To observe the expressions of cerebral nerve growth factor (NGF) and choline acetyltransferase after simultaneous stimulation with electroacupuncture on Baihui (GV 20) and Dazhui (GV 14) so as to probe into the protection of electroacupuncture on hypoxia-ischemia brain injury.DESIGN: Randomized controlled experiment.SETTING: Department of Life Science in Zhengzhou Normal High Training School.MATERIALS: The experiment was performed in Human Anatomy Department of Basic Medical College of Zhengzhou University, in which, 50 cleangrade neonatal Wistar rats of 7 days old were employed and randomized into sham-operation group (10 rats), model control (20 rats) and electroacupuncture group (20 rats). Hypoxia cabin was self-made with constant pressure, 40 cm ×50 cm×60 cm in size, with two small holes of 2 cm ×2 cm for each to connect with the external. Soda lime was used to absorb moisture and CO2 in the cabin.The model was not prepared in sham-operation group. In model control and electroacupuncture group, hypoxia-ischemia model was set up. After modeling, the rats in two groups were recovered for 1 to 4 hours at room temperature; afterwards, hypoxia management was performed. The rats were placed in hypoxia cabin with constant pressure at constant temperature of 37 ℃, inputting O2 8 mL/L and mixed gas 920 mL/L, 1.5 L/minute; 2hours later, the rats were returned back to female rats for lactation continuously. In electroacupuncture group, on the 2nd day after modeling, a filiform needle of one cun was used to insert Baihui (GV 20) (midpoint of parietal bone) subcutaneously and Dazhui (GV 14) (between C7 and T1, on the midline of back). Electric stimulation was done simultaneously on two points with continuous wave, 16 Hz in frequency, 10V in intensity, retaining for 10 minutes, once per day, 10 days made 1 course, totally two courses at interval of 2 days. In model control, no any treatment was given group, 22 days after hypoxia and ischemia, the rats were anesthetized and sacrificed and brain tissue on the left side was collected to prepare paraffin slices. Immune positive cell was counted on slices of each group under optic microscope. For evaluation on the function of brain nerve cell, hippocampus was selected to count the positive cells of choline acetyltransferase. On each brain slice, 5 visual fields were randomized to calculate the average of positive cells. For evaluation on injury repair of nerve tissue, cortex and hippocampus were selected to count positive cells of cerebral NGF. The method was same as the above.MAIN OUTCOME MEASURES: Immune positive cell expressions of choline acetyltransferase and cerebral NGF in brain tissue after electric stimulation in neonatal rats.ferase in brain hippocampus: Compared with sham-operation group, that in model control was lower remarkably, but, there was no obvious change in electroacupuncture group (24.46±8.24), (13.96±7.62), (25.54±5.05) pcs/visual field, P ＜ 0.05, P ＞ 0.05]; that in electroacupuncture group was of cerebral NGF in cerebral cortex and hippocampus: Compared with sham-operation group, that in both model control and electroacupuncture was increased remarkably (14.14±6.11), (24.49±8.31), (31.35±9.92) pcs/visual field, P ＜ 0.05; (13.42±5.56), (21.93±5.12), (27.63±7.15) pcs/visual field, P ＜ 0.05], of which, that in electroacupuncture group was higher than model control (P ＜ 0.05).CONCLUSION: Electroacupuncture stimulates central cholinergic nerve system into positive active state in hypoxia-ischemia animals, increases cerebral nerve growth factor in quantity and enhances nerve repair of hypoxia-ischemia animals.