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目的:研究大鼠脑、小肠肌间神经丛神经元和血浆内Ghrelin的表达,探讨Ghrelin在水浸加束缚应激性胃溃疡中的作用及机制．方法:选择健康♂Wiater大鼠76只,随机分为 6组:水浸加束缚组(10只);侧脑室注射Ghrelin 组(24只);皮下注射L-NAME+侧脑室注射 Ghrelin组(8只)和相应的3个对照组(正常大鼠组18只,侧脑室注射生理盐水组8只,皮下注射 L-NAME+侧脑室注射生理盐水组8只)．采用放射免疫分析、荧光免疫组化双染和神经生理学等实验方法,观察脑、小肠肌间神经丛和血浆内Ghrelin的表达,探讨Ghrelin对大鼠束缚加水浸诱导的应激性胃溃疡的影响及机制．结果:在正常大鼠小肠肌间神经丛内和原代培养的肠肌间神经丛神经元均可见有Ghrelin 样免疫反应阳性物(Ghrelin-IR)的表达,且 Ghrelin-IR与胆碱乙酰转移酶(CHAT)共同表达于同一神经元内,但Ghrelin-IR不与一氧化碳合酶(NOS)和消化道感觉性神经元内特有的钙结合蛋白(Calbindin,Calb)共存．在大鼠应激性胃溃疡产生的同时,其血浆内Ghrelin- IR的含量明显减少(198．3±29．6 ng/L vs 141．7 ±26．5 ng／L．P=0．026)．而下丘脑、延脑、垂体和肌间神经丛神经元内Ghrelin-IR的含量明显升高(分别为96．2±18．1 pg／mg vs 153．2 ±11．6 pg／mg．P=0．006;89．8±16．5 pg／mg vs 144．4±13．9 pg／mg,P=0．007;108．3± 11．9 pg／mg vs 198．2±23．3 pg／mg,P=0．002; 48．8±12．8 pg／mg vs 86．2±21．5 pg／mg．P= 0．02);侧脑室注射Ghrelin大鼠应激性胃溃疡的产生明显减少,且呈明显的量效依赖关系(溃疡指数:生理盐水,86．7±6．2;50 ng Ghrelin,79．3±10．7 P=2．18;500 ng Ghrelin, 61．3±11．7,P=0．04;5 000 ng Ghrelin,35．6 ±10．8,P=0．005),经sc一氧化氮合酶抑制剂 L-NAME后,Ghrelin的胃黏膜细胞保护作用消失．结论:Ghrelin与ChAT共存表达于肠肌间神经丛胆碱能神经元;应激性胃溃疡发生时,中枢神经系统和血浆内Ghrelin的表达发生变化; 中枢Ghrelin对胃黏膜细胞具有保护作用,且呈明显的量效依赖关系．
Objective: To investigate the expression of Ghrelin in the brain and small intestine myenteric plexus neurons and plasma, and to explore the role and mechanism of Ghrelin in water stress plus gastric ulcer. Methods: Sixty-six healthy rats were randomly divided into 6 groups: water immersion plus restraint group (10 rats); intracerebroventricular injection of Ghrelin group (24 rats); subcutaneous injection of L-NAME + intracerebroventricular injection of Ghrelin group ) And the corresponding three control groups (normal rats 18, intraventricular injection of normal saline 8, subcutaneous injection of L-NAME + intraventricular injection of normal saline 8). The expression of Ghrelin in the brain, small intestine myenteric plexus and plasma were observed by radioimmunoassay, fluorescence double staining and neurophysiological methods to investigate the effect of Ghrelin on gastric ulcer induced by binding and water immersion in rats And mechanism. RESULTS: Ghrelin-IR expression was observed in the myenteric plexus of primary rat and primary cultured myenteric plexus neurons. Ghrelin-IR was associated with choline acetyltransferase (CHAT) co-expressed in the same neuron, but Ghrelin-IR does not coexist with carbon monoxide synthase (NOS) and calbindin (Calbindin, Calb) that is characteristic of gastrointestinal sensory neurons. In the same time, the level of Ghrelin-IR in plasma was significantly decreased in rats with gastric ulcer (198.3 ± 29.6 ng / L vs 141.7 ± 26.5 ng / L. P = 0.026 ). The levels of Ghrelin-IR in hypothalamus, medulla oblongata, pituitary and myenteric plexus neurons were significantly increased (96.2 ± 18.1 pg / mg vs 153.2 ± 11.6 pg / mg.P = 0.006; 89.8 ± 16.5 pg / mg vs 144.4 ± 13.9 pg / mg, P = 0.007; 108.3 ± 11.9 pg / mg vs 198.2 ± 23. 3 pg / mg, P = 0.002; 48.8 ± 12.8 pg / mg vs 86.2 ± 21.5 pg / mg.P = 0.02); intracerebroventricular injection of Ghrelin stress stomach There was a significant dose-dependent decrease in ulcer production (ulcer index: saline, 86.7 ± 6.2; 50 ng Ghrelin, 79.3 ± 10.7 P = 2.18; 500 ng Ghrelin, 61.3 ± 11.7, P = 0.04; 5 000 ng Ghrelin, 35.6 ± 10.8, P = 0.005). After sc-nitric oxide synthase inhibitor L-NAME, Gastric mucosal cell protection disappears. CONCLUSIONS: Ghrelin and ChAT coexist in cholinergic neurons of intestinal myenteric ganglion. The expression of Ghrelin in central nervous system and plasma changes when stress ulcer occurs. Ghrelin plays a protective role in gastric mucosal cells Significant dose-effect dependence.