(D-Ser2) oxyntomodulin recovers hippocampal synaptic structure and theta rhythm in Alzheimer’s disease transgenic mice

In our previous studies,we have shown that(D-Ser2)oxyntomodulin(Oxm),a glucagon-like peptide 1(GLP-1)receptor(GLP1R)/glucagon receptor(GCGR)dual agonist peptide,protects hippocampal neurons against Aβ1-42-induced cytotoxicity,and stabilizes the calcium homeostasis and mitochondrial membrane potential of hippocampal neurons.Additionally,we have demonstrated that(D-Ser2)Oxm improves cognitive decline and reduces the deposition of amyloid-beta in Alzheimer’s disease model mice.However,the protective mechanism remains unclear.In this study,we showed that 2 weeks of intraperitoneal administration of(D-Ser2)Oxm ameliorated the working memory and fear memory impairments of 9-month-old 3×Tg Alzheimer’s disease model mice.In addition,electrophysiological data recorded by a wireless multichannel neural recording system implanted in the hippocampal CA1 region showed that(D-Ser2)Oxm increased the power of the theta rhythm.In addition,(D-Ser2)Oxm treatment greatly increased the expression level of synaptic-associated proteins SYP and PSD-95 and increased the number of dendritic spines in 3×Tg Alzheimer’s disease model mice.These findings suggest that(D-Ser2)Oxm improves the cognitive function of Alzheimer’s disease transgenic mice by recovering hippocampal synaptic function and theta rhythm.

电针心包经穴对MCAO大鼠急性期D-ser及NMDAR的影响

目的:观察电针心包经穴对大脑中动脉闭塞(MCAO)大鼠神经功能缺损、缺血脑组织D-丝氨酸(D-Ser)含量、NMDAR表达的影响。方法:将36只SD大鼠随机分为正常组、假手术组、模型组、电针心包经组、模型+D-Ser组、电针心包经+D-Ser组。采用Zea Longa法制备MCAO模型,模型+D-Ser组、电针心包经+D-Ser组在插入线栓成功后30 min予外源性D-Ser试剂(160 mg/kg i.p.);电针心包经组、电针心包经+D-Ser组于次日行电针治疗30 min;其余组仅捆绑固定30 min。在大鼠造模成功后6 h和第2天干预后分别进行2次行为学评定,随后处死取材。应用液相色谱串联质谱法测缺血脑组织D-Ser含量、Western Blot测NR1、NR2A、NR2B的蛋白表达。结果:处理前与正常组、假手术组比,造模后各组的行为学评分均升高(P<0.05);与模型组相比,模型+D-Ser组和电针心包经+D-Ser组的行为学评分均升高(P<0.05)。处理后与模型组相比,电针心包经组差异无统计学意义(P>0.05),模型+D-Ser组和电针心包经+D-Ser组的行为学评分仍升高(P<0.05);与模型+D-Ser组相比,电针心包经+D-Ser组行为学评分无统计学意义(P>0.05)。组内处理前后比较,除正常组和假手术组外,各组行为学评分均略升高,差异无统计学意义(P>0.05)。与模型组相比,模型+D-Ser组的脑组织D-Ser含量及NR1、NR2A、NR2B的蛋白表达显著增加(P<0.05);电针心包经组的NR1、NR2A、NR2B的蛋白表达下降(P<0.05);与模型+D-Ser组相比,电针心包经+D-Ser组的D-Ser含量及NR1、NR2A、NR2B的蛋白表达均下降(P<0.05)。结论:电针心包经穴可降低急性期MCAO大鼠脑组织中D-Ser含量,调控NR1、NR2A、NR2B蛋白表达下降,抑制NMDA受体活性,发挥神经保护作用。

Effect of electroacupuncture at pericardium meridian on D‑ser and NMDAR in acute phase of MCAO rats

Objective :To observe the effect of electroacupuncture at pericardium meridian on neurological deficit, D-serine ( D-Ser ) content and NMDAR expression in ischemic brain tissue of rats with middle cerebral artery occlusion ( MCAO ). Methods:A total of 36 SD rats were randomly divided into normal group, sham operation group, model group, EA pericardium meridian group, model+D-Ser group,and EA pericardium meridian+D-Ser group. The MCAO model was prepared by Zea Longa method. The model+D-Ser group and the EA pericardium meridian+D-Ser group were given exogenous D-Ser reagent(160 mg/kg i.p.) 30 minutes after the success of the insertion of the suture. EA pericardium meridian group and EA pericardium meridian+D-Ser group were treated with electroacupuncture for 30 min on the next day. The remaining groups were only bound for 30 min.Behavioral evaluation was performed at 6 h after successful modeling and 2 d after intervention, and then the rats were sacrificed. The content of D-Ser in ischemic brain tissue was measured by liquid chromatography tandem mass spectrometry, and the protein expression of NR1, NR2A and NR2B was detected by Western Blot. Results : Before treatment, compared with the normal group and the sham operation group, the behavioral scores of each group increased after modeling (P<0.05 );Compared with the model group, the behavioral scores of the model+D-Ser group and the EA pericardium meridian+D-Ser group were increased (P<0.05).After treatment, compared with the model group, there was no significant difference in the EA pericardium meridian group (P>0.05),The behavioral scores of the model+D-Ser group and the EA pericardium meridian+D-Ser group were still increased (P<0.05);Compared with the model+D-Ser group, the behavioral score of the EA pericardium meridian+D-Ser group was not statistically significant (P>0.05).Comparison before and after treatment in the group,except for the normal group and the sham operation group, the behavioral scores of each group were slightly increased, and the difference was not statistically significant (P>0.05).Compared with the model group, the content of D-Ser and the protein expression of NR1, NR2A and NR2B in the brain tissue of the model+D-Ser group were significantly increased (P<0.05);The protein expression of NR1, NR2A and NR2B decreased in EA pericardium meridian group (P<0.05);Compared with the model+ D-Ser group, the D-Ser content and the protein expression of NR1, NR2A and NR2B in the EA pericardium meridian+D-Ser group were decreased (P<0.05). Conclusion : EA at pericardium meridian points can reduce the D-Ser content in brain tissue of rats with acute MCAO, regulate the decrease of NR1, NR2A and NR2B protein expression, inhibit the NMDA receptor activity, and play a neuroprotective role.

抗糖尿病新药(D-Ser2)Oxm拮抗淀粉样β蛋白细胞毒性的神经保护效应观察

淀粉样β蛋白(amyloidβ-protein,Aβ)在脑内的沉积及其神经毒性是阿尔茨海默病(Alzheimer’s disease,AD)的主要原因之一,目前仍缺乏拮抗Aβ的有效药物。最新报道表明,一种新的抗糖尿病药物(D-Ser2)Oxm不仅可以改善2型糖尿病(T2DM)大鼠的血糖和胰岛素水平,也具有促进皮层和海马神经元及其突触发生的效应。然而,(D-Ser2)Oxm是否能拮抗AD时Aβ所致的细胞损伤仍缺乏实验依据。本研究在培养原代大鼠海马神经细胞基础上,通过细胞活性和早期凋亡测定、细胞内钙成像以及线粒体膜电位检测,研究了(D-Ser2)Oxm对Aβ1-42所致细胞毒性的拮抗效应。结果显示,与单独给予Aβ1-42处理的细胞相比,(D-Ser2)Oxm+Aβ1-42处理组的细胞活力明显提高,而加入GLP-1受体抑制剂exendin(9-39)后,细胞活力则显著下降;(D-Ser2)Oxm可有效拮抗Aβ1-42导致的细胞凋亡,并使凋亡相关蛋白caspase3含量显著降低;(D-Ser2)Oxm处理还有效阻止了Aβ1-42引起的海马细胞内钙水平升高、线粒体膜电位去极化以及糖原合成酶激酶-3β(glycogen synthase kinase-3β,GSK-3β)(Y216)的活化。以上结果表明,(D-Ser2)Oxm可能是通过激动GLP-1受体对抗Aβ1-42的神经毒性,并且这种保护效应可能与细胞内钙稳态调节和线粒体膜电位稳定有关。