海马BDNF-p11信号通路在氯胺酮抗抑郁中的作用

发布时间：2018-08-07 06:55

【摘要】：目的：N-甲基-D-天冬氨酸(N-methyl-D-aspartate, NMDA)受体阻滞剂氯胺酮可以产生快速、有效、较持久的抗抑郁作用。不同分子机制、神经环路、信号通路和相关脑区在氯胺酮抗抑郁作用中具有一定的作用,但其具体作用机制尚不清楚。新近研究表明,小分子蛋白p11能够调节许多离子通道和五羟色胺(5-hydroxytryptamine,5-HT)受体在细胞膜上的表达,且促进突触再生,在抑郁症的神经病理学机制中具有重要作用。在抑郁症患者外周血液和抑郁模型动物相关脑区中,p11mRNA和蛋白表达均减少,而且p11的表达水平可作为判断抑郁症患者预后的指标。离体和在体实验均证实脑源性神经营养因子(Brain-derived neurotrophic factor, BDNF)能够诱导并促进海马p11表达,产生抗抑郁作用；而BDNF在氯胺酮抗抑郁作用中亦具有关键作用。因此,我们推测海马BDNF-p11信号通路参与氯胺酮的抗抑郁作用。本研究拟观察海马BDNF-p11信号通路在氯胺酮抗抑郁中的变化及其作用,并探讨相关机制。方法：采用慢性不可预知温和应激(Chronic unpredicted mild stress, CUMS)建立大鼠慢性抑郁模型。腹腔注射氯胺酮10 mg/kg或氯胺酮10 mg/kg联合ANA-12(特异性酪氨酸激酶B (Tyrosine kinases, TrkB)受体阻滞剂)0.5 mg/kg后0.5 h和72 h,行旷场实验(Open field test, OFT)、强迫游泳实验(Forced swimming test,FST)和蔗糖偏好实验(Sucrose preference test, SPT)以检测大鼠抑郁样行为。构建并在离体大鼠海马神经元培养中筛选p11慢病毒载体,将高效能p11慢病毒颗粒在体注射至大鼠海马中以沉默海马p11的表达,10天后行OFT.FST及SPT。行为学检测结束后,取大鼠海马组织,采用Western blotting检测大鼠海马中BDNF和p11蛋白表达。结果：与对照组相比,CUMS大鼠在FST中不动时间增加,在SPT中糖水偏好百分比降低,表现出抑郁样行为。注射氯胺酮后0.5 h和72 h,CUMS大鼠在FST中不动时间减少,在SPT中糖水偏好百分比增加,且与对照组差异无统计学意义,说明氯胺酮产生了快速和持续抗抑郁作用。联合注射氯胺酮和ANA-12后0.5 h和72 h,与单独注射氯胺酮相比,CUMS大鼠在FST中不动时间增加,在SPT中蔗糖偏好百分比减少；与注射生理盐水相比,CUMS大鼠在FST及SPT中差异无统计学意义,说明ANA-12可阻断氯胺酮的抗抑郁作用。各种处理措施对大鼠在OFT中运动总距离的影响差异均无统计学意义(P0.05)。与对照组相比,CUMS大鼠海马中BDNF和p11表达显著减少(P0.05)。氯胺酮注射后0.5 h和72 h,CUMS大鼠海马中BDNF显著增加,且与对照组差异无统计学意义。p11仅在氯胺酮注射后72 h,而不是0.5 h,显著增加(P0.05)。联合注射氯胺酮和ANA-12后72 h,与单独注射氯胺酮72 h相比,CUMS大鼠海马中p11表达显著减少(P0.05)；与注射生理盐水相比,CUMS大鼠海马P11表达差异无统计学意义(P0.05)。与对照组相比,在体海马注射慢病毒空载体对大鼠在FST中不动时间和SPT中蔗糖偏好百分比影响差异无统计学意义(P0.05)。与在体海马注射慢病毒空载体相比,在体海马注射p11慢病毒后13天,p11表达显著减少,大鼠表现出不动时间增加和蔗糖偏好百分比降低等抑郁样行为。与注射生理盐水相比,注射氯胺酮对在体海马注射p11慢病毒大鼠在FST中不动时间和SPT中蔗糖偏好百分比影响差异无统计学意义(P0.05),说明将大鼠海马p11沉默后,氯胺酮抗抑郁作用消失。结论：在CUMS大鼠抑郁模型中,海马BDNF-p11信号通路在氯胺酮抗抑郁作用的维持中起关键作用。
[Abstract]:Objective: N- methyl -D- aspartic acid (N-methyl-D-aspartate, NMDA) receptor blocker, ketamine, can produce rapid, effective and lasting antidepressant effects. Different molecular mechanisms, neural circuits, signal pathways and related brain regions have a certain role in the antidepressant effect of ketamine, but the specific mechanism of its action is not yet clear. It shows that small molecular protein P11 can regulate the expression of many ion channels and five serotonin (5-hydroxytryptamine, 5-HT) receptors on the cell membrane and promote synapse regeneration, which plays an important role in the neuropathological mechanism of depression. In the peripheral blood and depression model animal related brain regions of the depressive patients, the expression of p11mRNA and protein are all Decrease, and the expression level of P11 can be used as an indicator to judge the prognosis of patients with depression. In vitro and in vivo, Brain-derived neurotrophic factor (BDNF) can induce and promote the expression of P11 in hippocampus and produce antidepressant effect. BDNF also plays a key role in the antidepressant effect of ketamine. Therefore, we speculate that the hippocampal BDNF-p11 signaling pathway participates in the antidepressant effect of ketamine. This study intends to observe the changes and effects of the hippocampal BDNF-p11 signaling pathway in ketamine antidepressants and to explore the mechanisms. Methods: chronic unpredictable mild stress (Chronic unpredicted mild stress, CUMS) is used to establish chronic depression models in rats. Type. Intraperitoneal injection of ketamine 10 mg/kg or ketamine 10 mg/kg combined with ANA-12 (specific tyrosine kinase B (Tyrosine kinases, TrkB) receptor blocker) 0.5 h and 72 h after 0.5 mg/kg (Open field), forced swimming test and sucrose preference test The P11 lentivirus vector was screened in the cultured rat hippocampal neurons in vitro, and the high efficient P11 lentivirus particles were injected into the hippocampus of the rat to silence the expression of P11 in the hippocampus. After 10 days of OFT.FST and SPT. behavior detection, the hippocampus tissue was taken and B in the hippocampus of rats was detected by Western blotting. DNF and P11 protein expression. Results: compared with the control group, the duration of CUMS rats increased in FST, and the percentage of sugar water preference decreased in SPT, showing depressive behavior. After injection of ketamine, 0.5 h and 72 h, CUMS rats were less active in FST, and the ratio of sugar to water in SPT increased, and there was no significant difference from the control group. The combination of ketamine and ANA-12 0.5 h and 72 h after injection of ketamine and ANA-12, compared with the single injection of ketamine, increased the duration of CUMS rats in FST and decreased the percentage of sucrose preference in SPT; compared with the injection of saline, the difference in FST and SPT in CUMS rats was not statistically significant, indicating ANA-12 can be found. The antidepressant effect of ketamine was blocked. There was no significant difference in the effect of various treatment measures on the total distance of exercise in OFT (P0.05). Compared with the control group, the expression of BDNF and P11 in the hippocampus of CUMS rats decreased significantly (P0.05). The BDNF of 0.5 h and 72 h in the rats after injection of ketamine, and the BDNF of the hippocampus in the rats of CUMS significantly increased, and there was no difference between the control group and the control group. .p11 was only 72 h after ketamine injection, but not 0.5 h, significantly increased (P0.05). Compared with ketamine and ANA-12 72 h after injection of ketamine and ANA-12, the expression of P11 in hippocampus of CUMS rats decreased significantly (P0.05). Compared with the injection of saline, there was no significant difference in the expression of hippocampal P11 expression in CUMS rats. There was no significant difference in the effect of lentivirus no-load in the body hippocampus on the duration of FST and the percentage of sucrose preference in SPT (P0.05). Compared with the lentivirus empty vector injected in the hippocampus, the expression of P11 was significantly reduced at 13 days after the injection of P11 lentivirus in the hippocampus, and the rats showed increased time and sucrose preference. Compared with the injection of physiological saline, there was no significant difference in the effect of injection of ketamine on the duration of P11 lentivirus injection in FST rats in the hippocampus and the percentage of sucrose preference in SPT in the hippocampus of the rats (P0.05), indicating that the antidepressant effect of chloramine ketamine disappeared after the hippocampal P11 was silent in the rat. Conclusion: depression in CUMS rats In the model, the hippocampal BDNF-p11 signaling pathway plays a key role in the maintenance of ketamine antidepressant effect.
【学位授予单位】：南京大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R614

【共引文献】