shRNA敲降CaMKⅡβ对抑郁症小鼠行为及神经递质水平的影响

Effects of shRNA-mediated knockdown of CaMKⅡβ on behavior and neurotransmitter levels in depressed mice

目的:探讨shRNA敲降CaMKⅡβ对抑郁症小鼠行为及神经递质水平的影响。方法:45只小鼠随机分为对照组、模型组和实验组。实验组小鼠经皮层注射CaMKⅡβshRNA腺相关病毒1×10^9CFU/只,对照组和模型组经皮层注射对照shRNA腺相关病毒1×10^9CFU/只。感染4周后模型组和实验组采用慢性不可预见性应激制备抑郁症模型,然后进行悬尾、强迫游泳、糖水消耗和蔽箱实验测试;采用酶联免疫反应(enzyme-linked immunosorbent assay,ELISA)测定3组小鼠前额叶皮层多巴胺(dopamine,DA)、5-羟色胺(5-hydroxytryptamine,5-HT)和去甲肾上腺素(norepinephrine,NE)水平。结果:与对照组和模型组比较,实验组小鼠采用Ca MKⅡβshRNA明显降低了CaMKⅡβ蛋白的表达水平,差异有统计学意义(P<0.05)。3组小鼠水平运动和垂直运动次数比较,差异无统计学意义(P>0.05)。与对照组比较,模型组和实验组小鼠悬尾实验和强迫游泳实验持续不动时间明显增加,糖水消耗量显著减少,差异有统计学意义(P<0.05)。与模型组比较,实验组小鼠悬尾实验和强迫游泳实验持续不动时间明显增加,糖水消耗量明显减少,差异有统计学意义(P<0.05)。与对照组比较,模型组和实验组小鼠DA、5-HT和NE水平明显下调,差异有统计学意义(P<0.05)。与模型组比较,实验组小鼠皮层DA、5-HT和NE的水平明显升高,差异有统计学意义(P<0.05)。结论:腺相关病毒介导CaMKⅡβshRNA可明显下调脑组织CaMKⅡβ蛋白水平,并上调DA、5-HT和NE水平,进而达到治疗抑郁症的效果。

Objective:To investigate the effects of short hairpin RNA(shRNA)-mediated knockdown of calcium/calmodulin-dependent protein kinase Ⅱ beta(Ca MKⅡβ)on behavior and neurotransmitter levels in depressed mice. Methods:Forty-five mice were randomly divided into control group,model group,and experimental group. The experimental group received an intracortical injection of CaMKⅡβ shRNA adeno-associated virus at 1×10^9 CFU/mouse,and both the control group and model group received an intracortical injection of control shRNA adeno-associated virus at 1×10^9 CFU/mouse. After four weeks of infection,chronic unpredictable stress was applied in both the model group and experimental group to develop an animal model of depression,followed by tail suspension,forced swimming,sugar water consumption,and covered box tests. The levels of dopamine(DA),5-hydroxytryptamine(5-HT),and norepinephrine(NE) in the prefrontal cortex of three groups of mice were determined by enzyme-linked immunosorbent assay.Results:Compared with the control group and model group,the experimental group had significantly lower CaMKⅡβ expression with CaMKⅡβ shRNA adeno-associated virus(P<0.05). There was no significant difference in the number of horizontal and vertical movements between the three groups(P>0.05). Compared with the control group,both the model group and experimental group had significantly longer immobility time in tail suspension and forced swimming tests and less sugar water consumption(P<0.05). Compared with the model group,the experimental group had significantly longer immobility time in tail suspension and forced swimming tests and less sugar water consumption(P<0.05). Compared with the control group,both the model group and experimental group had significantly lower levels of DA,5-HT,and NE(P<0.05). Compared with the model group,the experimental group had significantly higher levels of DA,5-HT,and NE(P<0.05). Conclusion:Adeno-associated virus-mediated CaMKⅡβ shRNA can significantly down-regulate the level of CaMKⅡβ and up-regulate the levels of DA,5-HT,and NE in brain tissue,thus achieving the effect of treating depression.