L-NNA对酒精中毒家兔脑组织NOS活性及NOS阳性神经元的影响

发布时间：2018-04-23 05:31

本文选题：纳洛酮 + NG-硝基-L-精氨酸　；参考：《山东农业大学》2015年硕士论文

【摘要】：本试验用兔作为实验动物制备慢性酒精中毒模型,一氧化氮合酶抑制剂L-NNA及纳络酮对成功模型进行比较治疗。通过测定不同组慢性酒精中毒家兔额叶、海马、纹状体中NOS的活性、NO水平,观测NOS阳性神经元形态结构及分布来探讨慢性酒精中毒脑损伤与NO的关系以及L-NNA对受损脑组织中NOS活性、NO水平的影响,为慢性酒精中毒脑损伤机制、病理学研究以及临床治疗提供理论依据。通过酒精灌胃的方法建立家兔慢性酒精中毒模型,并通过体重、行为学变化以及病理组织学确定家兔是否已经造成脑组织损伤。酒精灌胃的家兔在6周后体重出现负增长,经病理组织学观察可知:在酒精灌胃8周时,酒精灌胃家兔脑组织出现轻微的神经元缺失,随着灌胃时间的增长,脑组织神经元缺失加重,且有细胞溶解消失,海马区锥体细胞排列紊乱,免疫组织化学实验显示nNOS及iNOS阳性神经元在不同脑区的表达发生一定变化,进一步验证了模型的可靠性。利用硝酸还原酶法和生物化学检测方法来分别检测各组家兔额叶、海马、纹状体内的NOS活性变化及NO水平变化。结果显示:模型组家兔额叶、海马及纹状体中NOS活性一直保持上升趋势,且在各个时间点均显著高于对照组,NO含量变化与其相一致;给予纳洛酮及L-NNA后,治疗组各脑区NOS及NO水平均上升缓慢,给药3 d后均呈下降趋势,14 d后变化趋于平稳,三组治疗组不同脑区NOS活性及NO水平均显著低于模型组。利用SABC免疫组织化学法在光学显微镜下对各组家兔额叶、海马各区以及纹状体的nNOS阳性神经元与i NOS阳性神经元的形态结构进行观测。试验结果表明:各组兔脑组织内nNOS和iNOS阳性神经元的数量与NO的含量及NOS活性在脑组织中的变化趋势基本一致。模型组、纳洛酮组及L-NNA组中不同脑组织内nNOS及iNOS阳性神经元的表达显著高于正常对照组,与酒精模型组相比,三组治疗组脑内NOS阳性神经元的表达均有所降低,其中联合治疗组降低幅度最大。结果表明:酒精模型组中额叶、海马及纹状体中的神经元缺失、死亡伴随相应组织中NO含量的增加以及nNOS和iNOS阳性神经元表达增强,而给予纳洛酮及L-NNA治疗后,治疗组中NO水平、nNOS及iNOS阳性神经元的表达均有所下降,家兔脑组织病理变化减轻,一般状态及体重增长情况得到改善,提示慢性酒精中毒时脑组织的损伤可能是由NO参与介导的,而L-NNA及纳络酮均能抑制酒精中毒时NOS的活性,使额叶、海马及纹状体组织中NO水平降低,改善家兔的一般状态,为临床上慢性酒精中毒的治疗和新药研发提供新的依据和方向。
[Abstract]:The model of chronic alcoholism was established in rabbits. The successful model was treated by nitric oxide synthase inhibitor (L-NNA) and naloxone. The activity of NOS in frontal lobe, hippocampus and striatum of rabbits with chronic alcoholism was measured. The morphological structure and distribution of NOS positive neurons were observed to explore the relationship between brain injury and no in chronic alcoholism and the effect of L-NNA on the level of NOS activity in the injured brain tissue, which was the mechanism of brain injury in chronic alcoholism. Pathological study and clinical treatment provide theoretical basis. The model of chronic alcoholism in rabbits was established by the method of alcohol administration, and the changes of body weight, behavior and histopathology were used to determine whether the rabbit had caused brain injury. The weight of rabbits fed with alcohol showed negative growth after 6 weeks. By histopathological observation, there was a slight neuronal deficiency in the brain tissue of the rabbits fed with alcohol for 8 weeks, and with the increase of the time of gastric perfusion, there was a slight loss of neurons in the brain tissue of the rabbits fed with alcohol for 8 weeks. The loss of neurons in brain tissue was aggravated, cell dissolution disappeared, and the pyramidal cells in hippocampus were disordered. Immunohistochemical experiments showed that the expression of nNOS and iNOS positive neurons in different brain regions changed to a certain extent. The reliability of the model is further verified. The changes of NOS activity and no level in frontal lobe, hippocampus and striatum of rabbits were detected by nitrate reductase method and biochemical method respectively. The results showed that the activity of NOS in frontal lobe, hippocampus and striatum of the model group maintained an increasing trend, and the content of no in the model group was significantly higher than that in the control group at each time point. The levels of NOS and no increased slowly in all brain regions of the treatment group, and showed a decreasing trend after 3 days of administration. The NOS activity and no level in different brain regions of the three treatment groups were significantly lower than those in the model group. The morphology of nNOS positive neurons and I NOS positive neurons in frontal lobe, hippocampus and striatum of rabbits were observed by SABC immunohistochemical method under optical microscope. The results showed that the number of nNOS and iNOS positive neurons in the brain tissue of each group was consistent with the content of no and the activity of NOS in the brain tissue. The expression of nNOS and iNOS positive neurons in different brain tissues in model group, naloxone group and L-NNA group were significantly higher than those in normal control group. Compared with alcohol model group, the expression of NOS positive neurons in brain of three treatment groups were decreased. In the combined treatment group, the decrease was the largest. The results showed that the loss of neurons in frontal lobe, hippocampus and striatum was accompanied by the increase of no content and the expression of nNOS and iNOS positive neurons in the corresponding tissues. After the treatment of naloxone and L-NNA, Naloxone and L-NNA were given. In the treatment group, the expression of nNOS and iNOS positive neurons were decreased, the pathological changes of brain tissue were alleviated, and the general state and weight gain were improved. L-NNA and naloxone could inhibit the activity of NOS, decrease the level of no in frontal lobe, hippocampus and striatum, and improve the general state of rabbits. To provide a new basis and direction for the clinical treatment of chronic alcoholism and the development of new drugs.
【学位授予单位】：山东农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S858.291

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