[Effects of Yizhi Tiaoshen acupuncture on learning and memory function and the expression of phosphorylated tau protein in the hippocampus of Alzheimer's disease model rats].

To observe the effects of Yizhi Tiaoshen (benefiting mental health and regulating the spirit) acupuncture on learning and memory function, and the expression of phosphorylated tubulin-associated unit (tau) protein in the hippocampus of Alzheimer's disease (AD) model rats, and explore the effect mechanism of this therapy on AD.A blank group and a sham-operation group were randomly selected from 60 male SD rats, 10 rats in each one. AD models were established in the rest 40 rats by the intraperitoneal injection of D-galactose and okadaic acid in the CA1 region of the bilateral hippocampus. Thirty successfully-replicated model rats were randomly divided into a model group, a western medication group and an acupuncture group, 10 rats in each one. In the acupuncture group, acupuncture was applied to "Baihui" (GV 20), "Sishencong" (EX-HN 1), "Neiguan" (PC 6), "Shenmen" (HT 7), "Xuanzhong" (GB 39) and "Sanyinjiao" (SP 6); and the needles were retained for 10 min. Acupuncture was given once daily. One course of treatment was composed of 6 days, with the interval of 1 day; the completion of treatment included 4 courses. In the western medication group, donepezil hydrochloride solution (0.45 mg/kg) was administrated intragastrically, once daily; it took 7 days to accomplish one course of treatment and a completion of intervention was composed of 4 courses. Morris water maze (MWM) and novel object recognition test (NORT) were used to assess the learning and memory function of the rats. Using HE staining and Nissl staining, the morphological structure of the hippocampus was observed. With Western blot adopted, the protein expression of the tau, phosphorylated tau protein at Ser198 (p-tau Ser198), protein phosphatase 2A (PP2A) and glycogen synthase kinase-3β (GSK-3β) in the hippocampus was detected.There were no statistical differences in all of the indexes between the sham-operation group and the blank group. Compared with the sham-operation group, in the model group, the MWM escape latency was prolonged (P<0.05), the crossing frequency and the quadrant stay time in original platform were shortened (P<0.05), and the NORT discrimination index (DI) was reduced (P<0.05); the hippocampal cell numbers were declined and the cells arranged irregularly, the hippocampal neuronal structure was abnormal and the numbers of Nissl bodies decreased; the protein expression of p-tau Ser198 and GSK-3βwas increased (P<0.05) and that of PP2A decreased (P<0.05). When compared with the model group, in the western medication group and the acupuncture group, the MWM escape latency was shortened (P<0.05), the crossing frequency and the quadrant stay time in original platform were increased (P<0.05), and DI got higher (P<0.05); the hippocampal cell numbers were elevated and the cells arranged regularly, the damage of hippocampal neuronal structure was attenuated and the numbers of Nissl bodies were increased; the protein expression of p-tau Ser198 and GSK-3β was reduced (P<0.05) and that of PP2A was increased (P<0.05). There were no statistically significant differences in the above indexes between the acupuncture group and the western medication group (P>0.05).Acupuncture therapy of "benefiting mental health and regulating the spirit" could improve the learning and memory function and alleviate neuronal injure of AD model rats. The effect mechanism of this therapy may be related to the down-regulation of GSK-3β and the up-regulation of PP2A in the hippocampus, and then to inducing the inhibition of tau protein phosphorylation.目的：观察“益智调神”法针刺对阿尔茨海默病（AD）模型大鼠学习记忆功能及海马微管相关蛋白（tau）蛋白磷酸化表达的影响，探讨“益智调神”法针刺治疗AD的作用机制。方法：从60只雄性SD大鼠中随机选出空白组和假手术组，每组10只。剩余40只大鼠采用腹腔注射D-半乳糖与双侧海马CA1区注射冈田酸建立AD模型。将模型复制成功的30只大鼠随机分为模型组、西药组和针刺组，每组10只。针刺组针刺“百会”“四神聪”“内关”“神门”“悬钟”“三阴交”，留针10 min，每日1次，每周6次，休息1 d，7 d为一疗程，共治疗4个疗程。西药组给予盐酸多奈哌齐溶液（0.45 mg/kg）灌胃治疗，每日1次，7 d为一疗程，共治疗4个疗程。干预结束后，采用Morris水迷宫实验、新物体识别实验评估大鼠学习记忆功能，利用HE染色法、尼氏染色法观察大鼠海马组织形态，采用Western blot法检测海马tau、磷酸化tau蛋白丝氨酸198位点（p-tau Ser198）、蛋白磷酸酶2A（PP2A）、糖原合成酶激酶-3β（GSK-3β）蛋白表达。结果：假手术组与空白组各项指标比较差异均无统计学意义（P>0.05）。与假手术组比较，模型组逃避潜伏期延长（P<0.05），穿越原平台次数、原平台所在象限停留时间减少（P<0.05），物体辨别指数（DI）值降低（P<0.05）；海马细胞数量减少、排列不整齐，海马神经元结构异常，尼氏体数量减少；p-tau Ser198、GSK-3β蛋白表达增多（P<0.05），PP2A蛋白表达减少（P<0.05）。与模型组比较，西药组和针刺组逃避潜伏期缩短（P<0.05），穿越原平台次数、原平台所在象限停留时间增加（P<0.05），DI值升高（P<0.05）；海马细胞数量增加、排列尚整齐，海马神经元结构损害减轻，尼氏体数量增加；p-tau Ser198、GSK-3β蛋白表达减少（P<0.05），PP2A蛋白表达增多（P<0.05）。针刺组与西药组以上指标比较差异无统计学意义（P>0.05）。结论：“益智调神”法针刺可改善AD模型大鼠学习记忆功能，减轻神经元损伤，其机制可能与下调海马GSK-3β表达，上调PP2A表达，进而抑制tau蛋白磷酸化有关。.