左归降糖解郁方调控AMPK/MFF/DRP1信号改善糖尿病并发抑郁症大鼠海马线粒体动力学

Effect of Zuogui Jiangtang Jieyu decoction on hippocampal mitochondrial dynamics in diabetic rats with depression based on AMPK/MFF/DRP1 Signaling

关键词:  左归降糖解郁方  糖尿病并发抑郁症  线粒体  腺苷酸活化蛋白激酶

Key words:  Zuogui Jiangtang Jieyu decoction  Diabetes complicated with depression  Mitochondria  AMPK

摘要:

目的:探讨左归降糖解郁方改善糖尿病并发抑郁症大鼠海马神经元线粒体动力学的作用机制。方法:对雄性SD大鼠进行高脂饲养4周后尾静脉注射链脲佐菌素,根据空腹血糖值筛选糖尿病大鼠,随后对其进行28天慢性温和不可预知应激,建立糖尿病并发抑郁症模型并随机分为模型组、AMPK抑制剂组、左归降糖解郁方高、低剂量组。另设等数量健康SD大鼠为正常组。药物干预4周后,血糖试纸检测空腹血糖,强迫游泳实验评价动物的抑郁样情绪,Morris水迷宫评价动物的学习记忆能力,尼氏染色和电镜检测海马神经元及胞内线粒体形态,比色法检测能量代谢产物ATP含量和线粒体呼吸链复合物Ⅰ-Ⅴ含量,免疫荧光法检测神经元中线粒体动力学相关蛋白MFN1、MFN2、DRP1表达,Western blot法检测AMPK、MFN2、MFFser173、DRP1蛋白表达。结果:与正常组比较,模型组大鼠表现出明显的高血糖和抑郁样行为,大鼠海马线粒体呼吸链复合物Ⅰ-Ⅴ含量及能量产物ATP均减少,神经元中MFN1、MFN2表达减少,AMPK、MFFser173、DRP1表达增加。与模型组比较,AMPK抑制剂可显著降低AMPK、MFFser173、DRP1表达,改善线粒体氧化磷酸化功能,增加ATP含量,保护线粒体形态,但对MFN1、MFN2无显著影响；左归降糖解郁方可显著降低糖尿病并发抑郁症大鼠外周血糖,改善动物的抑郁样行为和认知障碍,改善海马线粒体形态,增加神经元线粒体呼吸链复合物Ⅰ-Ⅴ和ATP含量,增加线粒体MFN1、MFN2表达,减少AMPK、MFFser173、DRP1的表达。结论:左归降糖解郁方可通过调节AMPK/MFF/DRP1信号,抑制神经元线粒体分裂,改善其动力学,增加神经元氧化磷酸化水平和能量代谢。

Abstract:

Objective To explore the mechanism of Zuogui Jiangtang Jieyu decoction on the mitochondrial dynamics of hippocampal neurons in diabetic rats with depression. Methods The male SD rats were fed with high-fat diet fed for 4 weeks, and then injected with streptozotocin through tail vein. According to the level of fasting blood glucose, diabetic rats were definited. Then they were subjected to 28 days of chronic mild unpredictable stress to establish the model of diabetes complicated with depression. The model rats were randomly divided into model group, AMPK inhibitor group, Zuogui Jiangtang Jieyu decoction high and low dose groups. The equal number of healthy SD rats were set as the normal group. After 4 weeks of intervention, fasting blood glucose levels were measured by blood glucose test strips. The behaviors of rats were evaluated by forced swimming test and Morris water maze. Hippocampal neurons and intracellular mitochondrial morphology were detected by Nissl staining and electron microscopy. The level of ATP and mitochondrial respiratory chain complex I-V were detected by colorimetric methods. Mitochondrial dynamics related proteins MFN1, MFN2 and DRP1 were detected by immunofluorescence. The expression of AMPK, MFN2, MFFser173, and DRP1 were detected by Western blot. Result Compared with the normal group, the model group rats showed significant hyperglycemic and depressive behavior. The content of mitochondrial respiratory chain complexes I-V and energy product ATP in the hippocampus of rats were significantly decreased, and the expression of MFN1 and MFN2 were decreased. The expression of AMPK, MFFser173, and DRP1 were significantly increased. Compared with the model group, AMPK inhibitors significantly reduced the expression of AMPK, MFFser173, and DRP1. The mitochondrial oxidative phosphorylation function were improved, and the content of ATP was increased. But there was no significant effect on MFN1 and MFN2. Zuogui Jiangtang Jieyu decoction significantly reduced the peripheral blood glucose of diabetic rats with depression. The depression like behavior and cognitive function were improved. The morphology of hippocampus mitochondria were improved. The contents of neuronal mitochondrial respiratory chain complexes I-V and ATP were increased. The expression of MFN1 and MFN2 were increased and the expression of AMPK, MFFser173, DRP1 were reduced. Conclusion Zuogui Jiangtang Jieyu decoction can inhibit neuronal mitochondrial division by regulating AMPK/MFF/DRP1 signaling, improve mitochondrial dynamics and increase the oxidative phosphorylation levels.