电针“风池”“四神聪”介导NMDAR/CREB/BDNF信号通路改善血管性痴呆大鼠的学习记忆障碍  

Exploration of electroacupuncture at"Fengchi"(GB 20)and"Sishencong"(EX-HN 1)for attenuating learning and memory impairment in vascular dementia rats based on NMDAR/CREB/BDNF signaling pathway

作　　者：宋元毓 陈英华[2] 孙玮 李长青 李俊峰 王浩宇 秦瑞琦 苏晓庆 吴彤 赵红旭 韩玉生[1] SONG Yuanyu;CHEN Yinghua;SUN Wei;LI Changqing;LI Junfeng;WANG Haoyu;QIN Ruiqi;SU Xiaoqing;WU Tong;ZHAO Hongxu;HAN Yusheng(First Clinical Medicine School,Heilongjiang University of CM,Harbin 150040,China;Fifth Department of Acupuncture and Moxibustion,First Affiliated Hospital of Heilongjiang University of CM,Harbin 150040,China)

机构地区：[1]黑龙江中医药大学第一临床医学院,哈尔滨150040 [2]黑龙江中医药大学附属第一医院针灸五科,哈尔滨150040

出　　处：《中国针灸》2024年第12期1409-1417,共9页Chinese Acupuncture & Moxibustion

基　　金：国家自然科学基金资助项目:82074530;黑龙江省自然科学基金项目:ZD2021H007;黑龙江省中医药科研项目:ZHY2023-189、ZHY2023-209、ZHY2024-090;黑龙江中医药大学研究生创新科研项目:2024yjscx001。

摘　　要：目的:观察电针“风池”“四神聪”对血管性痴呆(VD)大鼠海马N-甲基-D-天冬氨酸受体(NMDAR)/环磷腺苷效应元件结合蛋白(CREB)/脑源性神经营养因子(BDNF)信号通路及海马区钙超载所致兴奋性毒性的影响,探究电针“风池”“四神聪”改善大鼠学习记忆障碍的作用机制。方法:选用SPF级雄性SD大鼠32只,随机分为正常组(6只)、假手术组(6只)和手术组(20只)。手术组采用改良的Pulsinelli四血管阻断(4-VO)法制备VD模型,将造模成功的12只大鼠随机分为模型组和电针组,每组6只。电针组于双侧“风池”及左右“四神聪”进行电针干预,予连续波,频率2 Hz,电流1 mA,以大鼠头部微颤为宜。电针干预每日1次,每次30 min,持续21d。Morris水迷宫实验检测大鼠学习记忆能力;HE染色观察大鼠海马CA1区神经元形态;比色法检测大鼠血清、海马组织谷氨酸(GLU)含量及海马组织Ca^(2+)-三磷酸腺苷(ATP)酶活力;免疫组化法检测大鼠海马CA1区NMDAR、钙调蛋白依赖性蛋白激酶Ⅱ(Ca MKⅡ)、磷酸化钙调蛋白依赖性蛋白激酶Ⅱ(p-Ca MKⅡ)、磷酸化环磷腺苷效应元件结合蛋白(p-CREB)、CREB、BDNF蛋白表达;Western blot法检测大鼠海马组织NMDAR、CREB、p-CREB、BDNF蛋白表达。结果:与假手术组比较,模型组大鼠逃避潜伏期延长、穿越平台次数减少(P<0.01);海马神经元结构不同程度受损,海马CA1区组织结构疏松,排列结构散乱,网格状结构明显,神经元形态不规则,核仁固缩甚至溶解,胶质细胞增多,毛细血管扩张,散见炎性细胞浸润;血清及海马组织GLU含量、海马组织NMDAR蛋白表达升高(P<0.01),海马组织Ca^(2+)-ATP酶活力及Ca MKⅡ、p-Ca MKⅡ、CREB、p-CREB、BDNF蛋白表达水平降低(P<0.01,P<0.05),p-CaMKⅡ/CaMKⅡ、p-CREB/CREB值降低(P<0.05)。与模型组比较,电针组大鼠逃避潜伏期缩短、穿越平台次数增加(P<0.01);大鼠海马CA1区组织排列散乱程度稍有好转,神经�Objective To explore the mechanism of electroacupuncture(EA)at"Fengchi"(GB 20)and"Sishencong"(EX-HN 1)on learning and memory impairment in vascular dementia(VD)rats by observing the influences on the N-methyl-D-aspartate receptor(NMDAR)/cyclic adenosine monophosphate response element-binding protein(CREB)/brain-derived neurotrophic factor(BDNF)signaling pathway and the excitotoxicity induced by hippocampal calcium overload.Methods Thirty-two male SD rats of SPF grade were selected and randomized into a normal group(6 rats),a sham-operation group(6 rats)and an operation group(20 rats).VD model was established with the modified Pulsinelli's four-vessel occlusion(4-VO)method.Twelve rats after successfully modeled were assigned randomly into a model group and an EA group,6 rats in each one.In the EA group,EA was delivered at bilateral"Fengchi"(GB 20)and"Sishencong"(EX-HN 1),with the continuous wave,the frequency of 2 Hz and the electric current of 1 mA.Stimulation intensity was adjusted depending on the slightly trembling of rat head.EA was given once daily,30 min each time;and EA intervention was delivered for 21 days continuously.Using Morris water maze test,the learning and memory function was assessed.The neuronal morphology in the hippocampal CA1 was observed with HE staining;the level of glutamate(GLU)in serum and hippocampal tissue,as well as the activity of calcium pump(Ca^(2+)-ATP)in the hippocampus were detected using colorimetric method.The protein expression of NMDAR,calmodulin-dependent protein kinaseⅡ(Ca MKⅡ),phosphorylated calmodulin-dependent protein kinaseⅡ(p-Ca MKⅡ),phosphorylated cyclic phosphoradenosine effector element binding proteins(p-CREB),CREB,and BDNF in the hippocampal CA1 was detected using immunohistochemistry.The protein expression of NMDAR,CREB,p-CREB and BDNF in the hippocampal tissue was detected using Western blot method.Results Compared to the sham-operation group,in the model group,the escape latency was prolonged and the platform crossing times of rats were reduced(P<0.01

关 键 词：血管性痴呆 电针 N-甲基-D-天冬氨酸受体(NMDAR)/环磷腺苷效应元件结合蛋白(CREB)/脑源性神经营养因子(BDNF)信号通路 谷氨酸 学习记忆障碍 

分 类 号：R245[医药卫生—针灸推拿学]