人参-知母-赤芍提取物对血管性痴呆大鼠海马神经元的保护作用及机制

目的:观察人参-知母-赤芍提取物对血管性痴呆大鼠海马N-甲基-D-天冬氨酸受体1(NMDAR1)的影响,探讨其保护海马神经元的作用机制。方法:60只SPF级雄性SD大鼠,随机分为正常组,假手术组,模型组,人参-知母-赤芍提取物组(0. 20 g·kg-1)和美金刚组(2. 1 mg·kg-1),每组12只。采用双侧颈总动脉反复夹闭合并腹腔注射硝普钠的方法建立血管性痴呆大鼠模型,造模后,正常组,假手术组,模型组给予同等体积生理盐水,每日1次,连续14 d。采用Morris水迷宫评价各组大鼠学习记忆能力;苏木素-伊红(HE)染色观察海马CA1区病理改变;蛋白免疫印迹法(Western blot)检测海马神经元胞膜NMDAR1的蛋白表达水平;免疫组化法(IHC)检测海马NMDAR1的表达情况;实时荧光定量PCR(Real-time PCR)检测海马组织中NMDAR1 mRNA的表达水平。结果:与正常组和假手术组比较,模型组大鼠逃避潜伏期显著延长(P 0. 01),在平台所在象限停留时间及穿越平台的次数显著减少(P 0. 01),海马CA1区神经细胞层次减少,排列紊乱、出现核固缩、神经元丢失,NMDAR1蛋白及mRNA表达显著升高(P 0. 01);与模型组比较,人参-知母-赤芍提取物组,美金刚组逃避潜伏期明显缩短(P 0. 05,P 0. 01),在平台所在象限停留时间及穿越平台的次数明显增加(P 0. 05,P 0. 01),海马CA1区神经元数目与形态有明显改善,海马神经元NMDAR1蛋白及NMDAR1 mRNA表达明显降低(P 0. 05)。结论:人参-知母-赤芍提取物能够改善血管性痴呆大鼠的学习记忆能力,减轻海马CA1区神经元的损伤,其机制可能与下调海马神经元NMDAR1的表达有关。

Protective Effect and Mechanism of Extracts from Ginseng Radix et Rhizoma,Anemarrhenae Rhizoma and Paeoniae Radix Rubra on Hippocampal Neurons in Rats with Vascular Dementia

Objective: To observe the effect of extracts from Ginseng Radix et Rhizoma,Anemarrhenae Rhizoma and Paeoniae Radix Rubra on N-methyl-D-aspartate receptors(NMDAR1) in hippocampal neurons in rats with vascular dementia and investigate its possible mechanism. Method: The 60 SPF male rats were randomly divided into normal group, sham-operated group,model group, traditional Chinese medicine group(0.20 g·kg^-1)and memantine group(2.1 mg·kg^-1),with 12 rats in each group. The model was established by repeated ischemia-reperfusion combined with intraperitoneal injection of sodium nitroprusside. After modelling, normal group, sham-operated group and model group were dosed the similar volume of normal saline once a day for 14 days. The learning and memory capacity was assessed by Morris water maze; pathologic change in the CA1 district of hippocampus was assessed by hematoxylin-eosin (HE) staining, and the expression level of NMDAR1 in hippocampal neuron membrane protein was detected by Western blot and immunohistochemistry(IHC),the NMDAR1 mRNA in hippocampal tissue was detected by Real-time PCR. Result:Compared with normal and sham-operated group, the latency period was prolonged in model group(P<0.01), the time in the platform quadrant and the frequency of crossing the platform were lessened significantly(P<0.01), the disorder of the neurons, the decrease of the neuronal number, and the neuronal necrosis and apoptosis were obversed in the CA1 district, expression of membrane NMDAR1 of the hippocampal neuron and NMDAR1 mRNA were increased significantly(P<0.01).Compared with model group, the latency period of extracts from Ginseng Radix et Rhizoma,Anemarrhenae Rhizoma and Paeoniae Radix Rubra group and memantine group were shortened significantly(P<0.05,P<0.01), the time and frequency were increased(P<0.05,P<0.01), the pathologic change was improved markedly, the protein expression of membrane NMDAR1 in hippocampal neuron and NMDAR1 mRNA were decreased significantly(P<0.01). Conclusion:The extracts from Ginseng Radix et Rhizoma,Anemarrhenae Rhizoma and Paeoniae Radix Rubra can improve the learning and memory capacity of rats with vascular dementia, and alleviate the injury in CA1 district of hippocampus. The mechanism may be related to the down-regulation of NMDAR1 expression in hippocampal neurons.