主 办:北 京 中 医 药 大 学
ISSN 1006-2157 CN 11-3574/R

北京中医药大学学报 ›› 2021, Vol. 44 ›› Issue (5): 444-453.doi: 10.3969/j.issn.1006-2157.2021.05.009

• 中药药理 • 上一篇    下一篇

复方柴金解郁方对抑郁症失眠大鼠海马、下丘脑CaMKⅡ和CREB表达影响及药效研究*

王叶情1, 王宇红1#, 黄会珍1, 邹蔓姝1, 刘灿1, 李春艳1, 赵洪庆1, 刘检2   

  1. 1 湖南中医药大学科技创新中心 湖南 410208;
    2 湖南中医药大学第一附属医院
  • 收稿日期:2020-09-05 出版日期:2021-05-30 发布日期:2021-06-01
  • 通讯作者: #王宇红,女,博士,研究员,博士生导师,主要研究方向:中药神经药理,E-mail:wyh107@126.com
  • 作者简介:王叶情,女,在读硕士生
  • 基金资助:
    *国家“重大新药创制”科技重大专项(No.2017ZX09309026),湖南省自然科学基金项目(No.2019JJ50428),湖南省教育厅一般项目(No.18C0371,No.18C0392),湖南省科技厅重点项目(No.2018DK2014),湖南省研究生创新项目(No.CX20200763),湖南中医药大学中医学国内一流学科建设项目(No.2018-2020)

Effects and efficacy of Compound Chaijin Jieyu Formula on the expression of CaMKⅡ and CREB in hypothalamus and hippocampus of rats with depression and insomnia*

Wang Yeqing1, Wang Yuhong1#, Huang Huizhen1, Zou Manshu1, Liu Can1, Li Chunyan1, Zhao Hongqing1, Liu Jian2   

  1. 1 Institute of Scientific Innovation and Applied Research,Hunan University of Chinese Medicine, Hunan 410208, China;
    2 The First Hospital of Hunan University of Chinese Medicine, Hunan 410007, China
  • Received:2020-09-05 Online:2021-05-30 Published:2021-06-01
  • Contact: Prof.Wang Yuhong, M.D., Doctoral Supervisor. Institute of Scientific Innovation and Applied Research, Hunan University of Chinese Medicine. No. 300 Xueshi Road, Hanpu Science and Education Park, Yuelu District, Changsha 410208. E-mail: wyh107@126.com
  • Supported by:
    National Science and Technology Major Project for Research and Development of Major New Drugs (No.2017ZX09309026), Natural Science Foundation of Hunan Province (No.2019JJ50428), General Project of Hunan Provincial Education Department (No.18C0371, No.18C0392), Key Project of Hunan Provincial Science & Technology Department (No.2018DK2014), Innovation Project for Graduate Students in Hunan Province (No.CX20200763)

摘要: 目的 观察复方柴金解郁方对抑郁症失眠大鼠海马、下丘脑钙调蛋白激酶Ⅱ(CaMKⅡ)和环磷腺苷反应元件结合蛋白(CREB)表达的影响,探讨复方柴金解郁方改善大鼠抑郁症失眠的作用机制。方法 将SD大鼠随机分为6组,即正常组,模型组,文拉法辛(13.5 μg/g)+地西泮组(0.9 μg/g),复方柴金解郁方高(10.8 g/kg)、中(5.4 g/kg)、低(2.7 g/kg)剂量组,采用慢性不可预见性温和应激(CUMS)联合睡眠剥夺复制抑郁症失眠大鼠模型。造模及给药干预35 d后,采用强迫游泳、戊巴比妥钠实验检测大鼠抑郁及失眠样行为,采用HE染色检测大鼠下丘脑、海马的病理变化,采用ELISA法检测大鼠血清和下丘脑中促肾上腺皮质激素释放激素(CRH)、促肾上腺皮质激素(ACTH)、皮质酮(CORT)的含量,采用HPLC-ECD法检测大鼠下丘脑和海马中去甲肾上腺素(NE)、5-羟色胺(5-HT)、多巴胺(DA)含量,采用Western blot法和实时荧光定量PCR法检测大鼠下丘脑和海马中CaMKⅡ、CREB蛋白及其基因表达。结果 与正常组比较,模型组大鼠强迫游泳实验中不动时间延长(P<0.01),睡眠潜伏期延长、睡眠时长缩短(P<0.01)。与模型组比较,复方柴金解郁方中剂量组强迫游泳实验中不动时间缩短(P<0.01),睡眠潜伏期缩短、睡眠时长延长(P<0.05)。文拉法辛+地西泮组、复方柴金解郁方组下丘脑和海马神经元细胞间隙缩小,排列紊乱和胞体空泡现象减少;复方柴金解郁方高、中剂量组大鼠血清和下丘脑中CRH、ACTH、CORT含量减少(P<0.01);文拉法辛+地西泮组和复方柴金解郁方高、中剂量组大鼠下丘脑和海马中NE、5-HT、DA含量升高(P<0.01);文拉法辛+地西泮组和复方柴金解郁方中剂量组大鼠下丘脑和海马中CaMKⅡ、CREB蛋白及其基因表达增加(P<0.05或P<0.01)。结论 复方柴金解郁方能缓解抑郁症失眠大鼠抑郁样行为、改善大鼠睡眠,其机制与减少下丘脑-垂体-肾上腺轴的过度分泌以及稳定单胺类神经递质表达、上调CaMKⅡ和CREB蛋白及其基因的表达有关。

关键词: 复方柴金解郁方, 抑郁症失眠, 下丘脑, 海马, 钙调蛋白激酶Ⅱ, 环磷腺苷反应元件结合蛋白, 大鼠

Abstract: Objective To observe the effect of Compound Chaijin Jieyu(Bupleurum & Turmeric Root Tuber Depression-Relieving) Formula (CCJJYF) on the expression of Calmodulin-Dependent Protein Kinase Ⅱ (CaMKⅡ) and cAMP-response-element-binding protein (CREB) in hypothalamus and hippocampus of rats with depression and insomnia, and to explore the mechanism of CCJJYF in relieving depression and insomnia. Methods 72 SD rats were randomly divided into normal group, model group,venlafaxine (13.5 μg/g)+diazepam (0.9 μg/g) group, high-dose (10.8 g/kg) CCJJYF group, mid-dose (5.4 g/kg) CCJJYF group and low-dose (2.7 g/kg) CCJJYF group (n=12). Chronic unpredictable mild stress (CUMS) combined with sleep deprivation was used to establish rat models of depression and insomnia. After 35 days of modeling and drug intervention, forced swim test (FST) and pentobarbital sodium test were used to detect depression and insomnia-like behaviors in rats. HE staining was used to detect the pathological changes in hypothalamus and hippocampus. The levels of CRH, ACTH and CORT in serum and hypothalamus were detected by ELISA. In addition, the levels of NE, 5-HT and DA in hypothalamus and hippocampus were detected by HPLC-ECD. Western blot and RT-qPCR were used to detect the protein level of CaMKⅡ and CREB and their gene expression in the hypothalamus and hippocampus. Results Compared with the normal group, the immobility time of rats in the model group was increased in the FST (P<0.01); besides, their sleep latency was prolonged, and their sleep duration was shortened (P<0.01). Compared with the model group, the immobility time in the mid-dose CCJJYF group was reduced in FST (P<0.01), their sleep latency was shortened, and their sleep duration increased (P<0.05). After the intervention of venlafaxine+diazepam and CCJJYF, the gap between hypothalamic and hippocampal neurons was reduced, the incidence of arrangement disorder and cell vacuoles was reduced.The levels of CRH, ACTH and CORT in serum and hypothalamus in the high-dose and mid-dose CCJJYF groups were reduced (P<0.01). The levels of NE, 5-HT and DA in the hypothalamus and hippocampus in the venlafaxine+diazepam group and the high-dose and mid-dose CCJJYF groups were increased (P<0.01). CaMKⅡ and CREB proteins and their gene expression in the hypothalamus and hippocampus in the venlafaxine+diazepam group and the mid-dose CCJJYF group increased (P<0.05 or P<0.01). Conclusion CCJJYF seems to alleviate depression-like behaviors in rats with depression and insomnia and improve their sleep quality. The mechanism may be related to reducing excessive secretion of HPA axis, stabilizing the expression of monoamine neurotransmitters and up-regulating CaMKⅡ and CREB protein and their gene expression.

Key words: Compound Chaijin Jieyu Formula(CCJJYF), depression and insomnia, hippocampus, hypothalamus, CaMKⅡ, CREB, rats

中图分类号: 

  • R285.5