FKBP12.6对小鼠学习记忆的影响

发布时间：2018-09-13 07:02

【摘要】：目的:FK506结合蛋白12.6(FK506 binding protein 12.6,FKBP12.6),属于FKBP蛋白家族成员之一,是免疫抑制剂FK506的胞内作用靶点。FKBP12.6由108个氨基酸组成,分子量为12.6KD,生理状态下能够与内质网和肌浆网上的RyR2结合,稳定钙离子通道,进而减少内质网和肌浆网中钙离子外泄;同时FKBP12.6与FK506结合,则可抑制钙调磷酸酶(CaN)活性,从而抑制免疫反应。FKBP12.6主要分布于心肌、脑组织和平滑肌等。FKBP12.6在中枢神经系统有着广泛的表达,包括一些与学习记忆相关的脑区,如海马、杏仁核;然而,其在学习记忆方面的功能还不清楚。本文旨在研究FKBP12.6敲除对新物体识别记忆、物体位置识别记忆及条件恐惧记忆的影响,并从电生理和蛋白表达方面探讨其可能的作用机制。方法:1.对小鼠尾部DNA进行基因鉴定,判断其是否成功敲除FKBP12.6基因。2.依据小鼠对新事物的探究特性建立实验模型,研究FKBP12.6敲除小鼠和WT(野生型)小鼠在新物体识别记忆和物体位置识别记忆上的差异。3.通过依据巴甫洛夫条件反射原理建立的恐惧记忆测试系统,研究FKBP12.6敲除小鼠和WT小鼠在条件恐惧记忆上的差异。4.通过膜片钳技术,研究FKBP12.6敲除小鼠和WT小鼠杏仁核神经元在电生理性质上的差异。5.通过免疫组化方法,研究FKBP12.6敲除小鼠和WT小鼠记忆相关脑区c-fos蛋白表达水平的差异。结果:1.实验中所用的小鼠确定已成功敲除FKBP12.6基因。2.与WT小鼠相比,FKBP12.6敲除小鼠的新物体识别记忆、物体位置识别记忆和条件恐惧记忆均下降。3.与WT小鼠相比,FKBP12.6敲除小鼠脑部杏仁核区域微小抑制性突触后电位(miniature inhibitory excitatory post synaptic potential,mIPSP)的频率没有显著变化,微小兴奋性突触后电位(miniature excitatory post synaptic potential,mEPSP)的频率显著下降。4.与WT小鼠相比,FKBP12.6敲除小鼠脑部杏仁核区域c-fos蛋白表达水平明显下降。结论:FKBP12.6基因敲除会损伤小鼠的新物体识别记忆、物体位置识别记忆和条件恐惧记忆;初步结果表明杏仁核mEPSP和c-fos蛋白表达水平的变化可能与条件恐惧记忆的损伤有关。
[Abstract]:Objective: FK506 binding protein 12.6 (FK506 binding protein 12.6FKBP12.6), a member of FKBP protein family, is the intracellular target of immunosuppressant FK506. FKBP12.6 is composed of 108 amino acids and has a molecular weight of 12.6KD. in physiological state, it can bind to endoplasmic reticulum (ER) and sarcoplasmic reticulum (RyR2). Calcium channel can be stabilized to reduce the leakage of calcium ions in endoplasmic reticulum and sarcoplasmic reticulum, meanwhile, FKBP12.6 combined with FK506 can inhibit the activity of calmodulin (CaN), thus inhibit the immune response. FKBP12.6 mainly distributes in myocardium. FKBP12.6 is widely expressed in the central nervous system, including some brain regions related to learning and memory, such as hippocampus and amygdala, but its function in learning and memory is not clear. The purpose of this paper is to study the effects of FKBP12.6 knockout on new object recognition memory, object position recognition memory and conditional fear memory, and to explore the possible mechanism of FKBP12.6 knockout in terms of electrophysiology and protein expression. Method 1: 1. The mouse tail DNA gene was identified to determine whether it successfully knocked out the FKBP12.6 gene. 2. An experimental model was established to study the difference between FKBP12.6 knockout mice and WT mice in new object recognition memory and object position recognition memory. The fear memory test system based on Pavlov's conditioned reflex principle was established to study the difference of conditional fear memory between FKBP12.6 knockout mice and WT mice. The electrophysiological properties of amygdala neurons in FKBP12.6 knockout mice and WT mice were studied by patch clamp technique. The difference of c-fos protein expression in memory related brain region between FKBP12.6 knockout mice and WT mice was studied by immunohistochemical method. The result is 1: 1. The mice used in the experiment confirmed that they had successfully knocked out the FKBP12.6 gene. 2. 2. Compared with WT mice, the new object recognition memory, object position recognition memory and conditional fear memory of FKBP12.6 knockout mice decreased by .3. Compared with WT mice, the frequency of (miniature inhibitory excitatory post synaptic potential,mIPSP) in the amygdala region of FKBP12.6 knockout mice was not significantly changed, but the frequency of (miniature excitatory post synaptic potential,mEPSP) was significantly decreased by .4.The frequency of (miniature excitatory post synaptic potential,mEPSP in the brain of FKBP12.6 knockout mice was significantly lower than that of FKBP12.6 knockout mice. Compared with WT mice, the expression of c-fos protein in the amygdala of FKBP12.6 knockout mice decreased significantly. Conclusion the new object recognition memory, the object position recognition memory and the conditioned fear memory can be damaged by the knockout of the 1: FKBP12.6 gene, and the preliminary results indicate that the changes of mEPSP and c-fos protein expression levels in the amygdala may be related to the damage of conditioned fear memory.
【学位授予单位】：南昌大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R338.64

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