内源性大麻素系统在荷瘤应激诱发抑郁中的作用
发布时间：2018-09-08 13:19
【摘要】：内源性大麻素系统(endocannabinoid system,ECS)与应激适应、情绪调控等精神功能存在密切关系。内源性大麻素(endocannabinoids,eCBs)花生四烯酸乙醇胺(anandamide,AEA)和花生四烯酸甘油酯(2-arachydonyl glycerol,2-AG),在神经冲动下诱导合成、释放,激动突触前膜的CB1受体,调节突触可塑性,维持神经功能稳态。本课题组前期通过对荷瘤小鼠行为学,海马基因表达及脑代谢组学的研究,发现外周荷瘤可诱发小鼠的抑郁表现,并导致大脑中与情绪调控相关的结构发生基因水平及代谢水平的改变。研究内源性大麻素系统在荷瘤应激中的变化,可为肿瘤诱发抑郁的生物学机制提供实验依据。本研究主要包括:1.采用替代分析物策略建立LC-MS/MS方法,用稳定同位素标记的AEA-d4和2-AG-d5替代AEA和2-AG,同时对小鼠血浆和大脑前额叶、海马及下丘脑的AEA及2-AG定量测定,其线性范围分别为0.325~32.5ng/mL和11~5500ng/mL。2.采用Western Blot技术对小鼠大脑上述各脑区的CB1受体以及大麻素降解酶即脂肪酸胺水解酶(fatty acid amide hydrolase,FAAH)的表达进行分析。3.采用EIA技术测定小鼠皮质酮水平。研究5-HT再摄取抑制剂类(selective serotonin reuptake inhibitors,SSRIs)抗抑郁药盐酸氟西汀及新型抗抑郁药阿戈美拉汀对荷瘤鼠皮质酮水平的影响;研究FAAH抑制剂URB597及CB1受体激动剂Win55,212-2对荷瘤小鼠皮质酮变化的影响。结果表明,本研究建立的替代分析物LC-MS/MS方法,解决了传统LC-MS/MS定量方法的内源性干扰,操作简捷,结果可靠。荷瘤小鼠前额叶、海马及下丘脑的AEA含量显著降低,而2-AG含量仅在前额叶显著降低,在海马及下丘脑未见明显变化,血浆的AEA及2-AG含量均无明显变化。荷瘤小鼠CB1受体及FAAH的表达仅在海马有显著增加,在前额叶及下丘脑无明显变化。荷瘤小鼠血浆较正常者升高,两种不同抗抑郁药均可降低荷瘤鼠升高的血浆皮质酮。荷瘤小鼠前额叶、海马及下丘脑的皮质酮均较正常组显著升高,抑制FAAH活性可有效降低荷瘤鼠的血浆以及上述不同脑区皮质酮的升高。CB1激动剂对荷瘤鼠血浆及不同脑区的皮质酮水平没有显著影响。综上,荷瘤可导致内源性大麻素系统发生改变,内源性大麻素系统可能在荷瘤应激及进一步诱发抑郁中发挥重要作用。抑制大麻素降解对改善荷瘤诱发的应激反应具改善作用。
[Abstract]:Endogenous cannabinoid system (endocannabinoid system,ECS) is closely related to stress adaptation, emotional regulation and other mental functions. Endogenous cannabinoid (endocannabinoids,eCBs) arachidonic acid ethanolamine (anandamide,AEA) and arachidonic acid glycerol ester (2-arachydonyl glycerol,2-AG) are induced to synthesize release excite the CB1 receptor of presynaptic membrane regulate synaptic plasticity and maintain neural function homeostasis. Through the study of behavior, hippocampal gene expression and brain metabolism in tumor-bearing mice, our group found that peripheral tumor could induce depression in mice. It also leads to changes in gene level and metabolism level in structures related to emotional regulation. To study the changes of endogenous cannabinoid system in tumor-bearing stress may provide experimental evidence for the biological mechanism of tumor-induced depression. This research mainly includes: 1. The LC-MS/MS method was established by the substitution analysis strategy, and the stable isotope labeled AEA-d4 and 2-AG-d5 were used to replace AEA and 2-AG.The linear ranges of AEA and 2-AG in plasma and prefrontal lobe, hippocampus and hypothalamus of mice were determined by 0.325~32.5ng/mL and 115500ng / ml 路L ~ (-2), respectively. Western Blot technique was used to analyze the expression of CB1 receptor and nip degrading enzyme (fatty acid amide hydrolase,FAAH) in the above brain regions of mice. The level of corticosterone in mice was determined by EIA technique. To study the effects of fluoxetine hydrochloride, a new antidepressant, 5-HT reuptake inhibitor (selective serotonin reuptake inhibitors,SSRIs, and a new antidepressant, acomelatin, on corticosterone levels in tumor-bearing mice, and to study the effects of FAAH inhibitor URB597 and CB1 receptor agonist Win55212-2 on the changes of corticosterone in tumor-bearing mice. The results showed that the LC-MS/MS method, which was established in this study, solved the endogenous interference of the traditional LC-MS/MS quantitative method, and the operation was simple and the results were reliable. The content of AEA in prefrontal lobe, hippocampus and hypothalamus decreased significantly in mice bearing tumor, but the content of 2-AG decreased only in prefrontal lobe, but not in hippocampus and hypothalamus. The contents of AEA and 2-AG in plasma did not change significantly. The expression of CB1 receptor and FAAH was increased only in hippocampus, but not in prefrontal lobe and hypothalamus. The plasma levels of corticosterone in tumor-bearing mice were higher than those in normal mice, and both antidepressants and antidepressants could decrease plasma corticosterone levels in tumor-bearing mice. Corticosterone in prefrontal lobe, hippocampus and hypothalamus of tumor-bearing mice was significantly higher than that of normal group. Inhibition of FAAH activity could effectively reduce the plasma levels of corticosterone in tumor-bearing mice and the elevation of corticosterone in different brain regions. CB1 agonists had no significant effect on the levels of corticosterone in plasma and different brain regions of tumor-bearing mice. The endogenous cannabinoid system may play an important role in tumor-bearing stress and further induced depression. Inhibition of cannabinoid degradation can improve tumor-induced stress response.
【学位授予单位】：上海交通大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R749.4
本文编号：2230636
[Abstract]:Endogenous cannabinoid system (endocannabinoid system,ECS) is closely related to stress adaptation, emotional regulation and other mental functions. Endogenous cannabinoid (endocannabinoids,eCBs) arachidonic acid ethanolamine (anandamide,AEA) and arachidonic acid glycerol ester (2-arachydonyl glycerol,2-AG) are induced to synthesize release excite the CB1 receptor of presynaptic membrane regulate synaptic plasticity and maintain neural function homeostasis. Through the study of behavior, hippocampal gene expression and brain metabolism in tumor-bearing mice, our group found that peripheral tumor could induce depression in mice. It also leads to changes in gene level and metabolism level in structures related to emotional regulation. To study the changes of endogenous cannabinoid system in tumor-bearing stress may provide experimental evidence for the biological mechanism of tumor-induced depression. This research mainly includes: 1. The LC-MS/MS method was established by the substitution analysis strategy, and the stable isotope labeled AEA-d4 and 2-AG-d5 were used to replace AEA and 2-AG.The linear ranges of AEA and 2-AG in plasma and prefrontal lobe, hippocampus and hypothalamus of mice were determined by 0.325~32.5ng/mL and 115500ng / ml 路L ~ (-2), respectively. Western Blot technique was used to analyze the expression of CB1 receptor and nip degrading enzyme (fatty acid amide hydrolase,FAAH) in the above brain regions of mice. The level of corticosterone in mice was determined by EIA technique. To study the effects of fluoxetine hydrochloride, a new antidepressant, 5-HT reuptake inhibitor (selective serotonin reuptake inhibitors,SSRIs, and a new antidepressant, acomelatin, on corticosterone levels in tumor-bearing mice, and to study the effects of FAAH inhibitor URB597 and CB1 receptor agonist Win55212-2 on the changes of corticosterone in tumor-bearing mice. The results showed that the LC-MS/MS method, which was established in this study, solved the endogenous interference of the traditional LC-MS/MS quantitative method, and the operation was simple and the results were reliable. The content of AEA in prefrontal lobe, hippocampus and hypothalamus decreased significantly in mice bearing tumor, but the content of 2-AG decreased only in prefrontal lobe, but not in hippocampus and hypothalamus. The contents of AEA and 2-AG in plasma did not change significantly. The expression of CB1 receptor and FAAH was increased only in hippocampus, but not in prefrontal lobe and hypothalamus. The plasma levels of corticosterone in tumor-bearing mice were higher than those in normal mice, and both antidepressants and antidepressants could decrease plasma corticosterone levels in tumor-bearing mice. Corticosterone in prefrontal lobe, hippocampus and hypothalamus of tumor-bearing mice was significantly higher than that of normal group. Inhibition of FAAH activity could effectively reduce the plasma levels of corticosterone in tumor-bearing mice and the elevation of corticosterone in different brain regions. CB1 agonists had no significant effect on the levels of corticosterone in plasma and different brain regions of tumor-bearing mice. The endogenous cannabinoid system may play an important role in tumor-bearing stress and further induced depression. Inhibition of cannabinoid degradation can improve tumor-induced stress response.
【学位授予单位】：上海交通大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R749.4
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1 Daniel J.Liput;Eleftheria Tsakalozou;Dana C.Hammell;Kalpana S.Paudel;Kimberly Nixon;Audra L.Stinchcomb;;Quantification of anandamide,oleoylethanolamide and palmitoylethanolamide in rodent brain tissue using high performance liquid chromatographyelectrospray mass spectroscopy[J];Journal of Pharmaceutical Analysis;2014年04期
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