【作者】 曹国红；

【导师】 艾洪滨； 崔希云；

【作者基本信息】 山东师范大学 ， 动物学， 2007， 硕士

【摘要】 大鼠束缚-浸水应激(restraint water-immersion stress,RWIS)模型被广泛用来研究应激性胃粘膜损伤的发生机制及其临床治疗药物的筛选等。众多实验结果表明:束缚-浸水应激(RWIS)状态下胃运动机能亢进、胃酸分泌增多主要是由于支配胃的副交感神经活动加强所致。这与其它多种类型的应激所致“下丘脑-腺垂体-肾上腺皮质轴”和“交感神经-肾上腺髓质系统”活动加强的现象相反;在束缚-浸水应激模型中却是副交感神经的活动加强,而这些副交感神经来自延髓的迷走背核和疑核。迷走神经背核(DMV)、疑核(NA)、孤束核(NTS)作为副交感神经的初级中枢对胃运动均具有调控作用,但是兴奋作用还是抑制作用,文献报道颇不一致。为了搞清楚这三个核团对胃运动到底起什么样的调控作用,本实验室前面的工作分别电刺激或化学刺激这三个核团,结果发现这三个核团受刺激后对胃运动均起抑制作用。那么,大鼠束缚-浸水应激状态下这些核团是兴奋还是抑制?脊髓灰质后角、前角、侧角的神经元是兴奋还是抑制?本文用即刻早期基因c-fos表达的Fos蛋白来作为神经元激活的标志,研究了束缚-浸水应激不同时间段的大鼠脊髓和脑干中各个神经核团的神经元的兴奋情况,联系大鼠的胃溃疡程度,进一步探讨脊髓与脑干各核团的兴奋与RWIS之间的关系。实验分为2部分。实验研究I:我们将实验大鼠随机分为五组,各组大鼠束缚-浸水应激时间分别为0 min、30 min、60 min、120 min、180 min,第一组作为对照组。取出后过量戊巴比妥钠麻醉,心脏灌流,取脊髓和脑干,后固定过夜,冰冻切片,片厚25μm。切片做Fos免疫组化。结果发现在脊髓前、后角和脑干多个核团(NTS, DMV, AP, SubP, LC, PBN, CGO, LRt-PC,等)中,Fos样免疫反应阳性细胞都有极显著的表达,但其表达的个数与强度不完全相同,且随着RWIS时间的延长,Fos表达呈现一定的规律。总体来说,应激60 min时大鼠脑干和脊髓中Fos表达达到顶峰,但具体到每个核团,又稍有差别。以后随时间延长,动物产生适应,Fos表达减少。实验研究Ⅱ:切断大鼠膈肌下迷走神经后RWIS 180 min。发现切断膈肌下迷走神经后再RWIS,大鼠脑干各核团中Fos表达明显减少,而脊髓的表达有所增强。胃粘膜部几乎没有溃疡点。本研究的结果表明:RWIS不同时间段大鼠脊髓前角、后角和脑干许多核团都有不同程度的Fos表达;RWIS 30-60 min时脊髓前角、后角和脑干大部分核团的Fos表达达到高峰。以后逐渐下降;发现切断膈肌下迷走神经后再RWIS 180 min,大鼠脑干各核团中Fos表达明显减少,而脊髓的表达有所增强。更多还原

【Abstract】 The rat under restraint water-immersion stress(RWIS) is widely used to study the stress-induced gastric ulcer and to find the medicine that can cure this disease. The results of many experiments indicated that the RWIS-induced gastric ulcer was mostly due to the hyperactivity of parasympathetic that control the stomach while other stresses induced the enhancement of hypothalamo-pituitary-adrenocortical axis (HPA axis) and sympathetic-adrenomedullar (SA). Parasympathetic nerve innervating the stomach is the vagal nerve which are largely from dorsal motor nucleus of the vagus (DMV) and partly from nucleus ambiguus (NA).As the center of parasympathetic preganglionic, NA, DMV and nucleus of solitary tract (NTS) all regulate gastric motility. However, there are no unanimous standpoints about the excited or inhibitory effects of NA, DMV and NTS on the gastric motility by now. To clarify the effects of the three nucleus on gastric motility, our research group has electrically stimulated them respectively and found that they all inhibited gastric motility. Whether excitement of neuron body or excitement of passed fibres conduced to these inhibiting results was unclear. So, in this study, we observed the Fos expression in each neuro-neucleus of the spinal cords and brainstems of rats following RWIS under different times, to discuss the relation between this stress and the excitement of neuro-neucleus in the spinal cords and brainstems of rats.The study included two parts, the experimentⅠa nd the experimentⅡ.The experimentⅠ: the rats were randomly devided into five experimental groups, four rats in each group. In conscious rats, the stress was controlled by employing the RWIS by 0min, 30min, 60min, 120min and 180min respectively. After heart perfusion, the spinal cord and brainstem were removed, and post-fixed overnight, then coronally sliced into 25μm thick of each piece. We used c-Fos immunohistochemistry to characterize neuronal activation of spinal cord and brainstem. There were very few signals for Fos-like immunoreactivities in the spinal cord and brainstem of unstressed rats. But RWIS could induced remarkable c-Fos expression in many parts(NTS, DMV, AP, SubP, LC, PBN, CGO, LRT-PC, et al) of spinal cord and brainstem, furthermore, the number of c-Fos positive nucleus and the intensity of c-Fos expression were different among each parts of the spinal cord and brainstem. Resumptively, the levels of c-Fos expression increased during the beginning time of RWIS and reached a maximum at 60min of stress. But to each nerval nucleus, it is not completely consistent. Fos expression decreased following a long stress time.The experimentⅡ: after bilateral vagotomy beneath diaphragm, rats suffered RWIS for 180min. Then the expression of c-Fos was detected by immunohistochemistry. We found that the Fos expression cut down in brainstem of the rats but boost up in spinal cord of the same rats.The results of this study indicate that RWIS could induced remarkable c-Fos expression in spinal anterior horn and posterior horn and brainstem, the levels of c-Fos expression increased during the beginning time of RWIS and reached a maximum at about 60min of stress. But after bilateral vagotomy beneath diaphragm, Fos expression cut down in brainstem but boost up in spinal cord of rats that suffered RWIS for 180min.更多还原