Apelin在大鼠中枢痛觉调制中的作用及其机制的实验研究

发布时间：2018-05-18 17:43

本文选题：侧脑室注射 + 痛觉过敏　；参考：《泰山医学院》2007年硕士论文

【摘要】： 目的以辐射热刺激甩尾法为疼痛模型，旨在探讨apelin在大鼠中枢痛觉调制中的作用及可能的作用机制。方法 1．痛阈测定：雄性SD大鼠，以辐射热引起大鼠甩尾，以大鼠甩尾反应的潜伏期(tail-flick latency，TFL)作为痛反应指标。分别向侧脑室微量注射生理盐水(NS)、吗啡(Morphine)和apelin，观察注药后60 min内大鼠痛阈的变化情况。 2．脑内nNOS表达的检测：雄性SD大鼠，，随机分为NS组、apelin组，采用免疫荧光方法在激光共聚焦显微镜下观察给药后30 min大鼠尾核、海马及皮层nNOS表达的变化。 3．血清和脑内NO、总NOS测定：雄性SD大鼠，分别向大鼠侧脑室微量注射NS和apelin，给药后30 min断头取血取脑，应用硝酸还原酶法测定血清及脑内NO和总NOS含量的变化。 4．血浆和脑内cAMP和cGMP测定：雄性SD大鼠，分别向大鼠侧脑室微量注射NS和apelin，给药后30 min断头取血取脑，应用放射免疫方法测定大鼠血浆及脑内cAMP和cGMP含量的变化。结果 1．侧脑室微量注射apelin后10～30 min大鼠痛阈与NS对照组相比显著降低，差异有统计学意义(P＜0.01)，30 min后大鼠痛阈逐渐回升，至60 min时大鼠痛阈仍低正常水平。 2．侧脑室注射apelin后，激光共聚焦显微镜观察到在关注脑区nNOS表达均发生变化。FITC标记的nNOS呈现绿色荧光颗粒分布于细胞胞浆中，为典型的细胞质分布；同时PI染色，细胞核为红色荧光。整个图像细胞形态完整，细胞核与细胞质的荧光色彩标识清晰可见。并且尾核及海马nNOS神经元较NS对照组表达明显降低，差异有统计学意义(P＜0.05～0.01)。 3．侧脑室微量注射apelin后，大鼠血清、尾核及海马中NO和NOS含量明显降低，与NS对照组比较，差异有统计学意义(P＜0.05～0.01)。 4．侧脑室微量注射apelin后，大鼠血浆、尾核、海马中cAMP含量升高，cGMP含量降低，与NS对照组比较，差异有统计学意义(P＜0.05～0.01)。结论 1．侧脑室微量注射apelin显著降低大鼠痛阈，使大鼠痛觉敏感化。 2．Apelin的这种痛觉敏感化作用至少部分是通过尾核、海马等核团的调控作用实现的。 3．侧脑室微量注射apelin明显提高大鼠尾核、海马及血浆中cAMP水平，降低大鼠尾核、海马中NO和NOS及血清中NO的含量，同时使大鼠尾核、海马及血浆中cGMP含量降低，提示apelin的痛觉敏感化作用可能与apelin-cAMP及apelin-NO-cGMP途径相关。本研究从行为学、组织和分子水平探讨了apelin在大鼠中枢痛觉调制中的作用及其机制，为中枢神经系统内痛和镇痛的研究提供了新的实验资料，对于进一步探讨痛觉调制的机制以及新药物的开发都有着重要的意义。
[Abstract]:Objective to investigate the role and possible mechanism of apelin in central pain modulation in rats. Method 1. Pain threshold measurement: male SD rats were induced tail flick by radiation heat, and tail flick latency TFL of tail flick reaction was used as the index of pain response. The changes of pain threshold were observed within 60 min after injection of NS, Morphine and apelin into lateral ventricle. 2. Detection of nNOS expression in brain: male SD rats were randomly divided into NS group and apelin group. The changes of nNOS expression in caudate nucleus, hippocampus and cortex of rats 30 min after administration were observed by immunofluorescence under confocal laser microscope. 3. Determination of no and total NOS in serum and brain: male Sprague-Dawley rats were microinjected with NS and apelin into lateral ventricle respectively. The blood and brain were taken 30 min after administration of the drug, and the changes of no and total NOS in serum and brain were measured by nitrate reductase method. 4. Determination of cAMP and cGMP in plasma and brain: male Sprague-Dawley rats were microinjected with NS and apelin into the lateral ventricle respectively. The blood samples were taken 30 min after administration of the drug. The contents of cAMP and cGMP in plasma and brain were measured by radioimmunoassay. Result 1. The pain threshold of rats at 1030 min after microinjection of apelin into the lateral ventricle was significantly lower than that of the NS control group (P < 0.01). After 30 min, the pain threshold of the rats gradually increased, and the pain threshold of the rats was still lower than that of the control group at 60 min. 2. After intracerebroventricular injection of apelin, it was observed by laser confocal microscopy that the expression of nNOS in the cerebral area of concern changed. FITC-labeled nNOS was distributed in the cytoplasm of the cells, which was typical cytoplasmic distribution and Pi staining. The nucleus is red fluorescent. The morphology of the whole image cell was complete, and the fluorescent color identification of the nucleus and cytoplasm was clearly visible. The expression of nNOS in caudate nucleus and hippocampus was significantly lower than that in NS control group (P < 0.05). 3. The contents of no and NOS in serum, caudate nucleus and hippocampus of rats were significantly decreased after microinjection of apelin into lateral ventricle, and the difference was statistically significant compared with NS control group (P < 0.05). 4. After microinjection of apelin into the lateral ventricle, the content of cAMP in plasma, caudate nucleus and hippocampus of rats increased and decreased, compared with NS control group, the difference was statistically significant (P < 0.05). Conclusion 1. Microinjection of apelin into lateral ventricle significantly reduced the pain threshold and sensitized the pain in rats. The pain sensitivity of 2.Apelin is at least partly mediated by the regulation of caudate and hippocampal nuclei. 3. Intracerebroventricular microinjection of apelin significantly increased the level of cAMP in caudate nucleus, hippocampus and plasma, decreased the content of no and NOS in caudate nucleus, hippocampus and serum, and decreased the content of cGMP in caudate nucleus, hippocampus and plasma of rats. The results suggest that the pain sensitivity of apelin may be related to the apelin-cAMP and apelin-NO-cGMP pathway. This study explored the role and mechanism of apelin in central pain modulation in rats at the behavioral, tissue and molecular levels, and provided new experimental data for the study of pain and analgesia in the central nervous system. It is of great significance to further explore the mechanism of pain modulation and the development of new drugs.
【学位授予单位】：泰山医学院
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R338

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