本文選題：膽堿能受體 + 腎上腺素能受體　； 參考：《吉林大學(xué)》2008年碩士論文

【摘要】： 本實(shí)驗(yàn)以純化的大鼠腦毒蕈堿乙酰膽堿m2受體(mAChR2)和β2腎上腺素能受體膜蛋白(β2-AR)為研究對(duì)象,觀察氨甲酰膽堿、阿托品和肝素分別對(duì)m2受體磷酸化過程的影響以及特布他林和肝素對(duì)β2-AR膜蛋白磷酸化過程的影響,探討激活劑或抑制劑影響的G蛋白偶聯(lián)受體激酶2 (GRK2)介導(dǎo)的膽堿能m2受體和β2-AR磷酸化的調(diào)節(jié)機(jī)制。探討興奮劑是如何使m2受體和β2-AR受體活化,引發(fā)涉及受體磷酸化的級(jí)聯(lián)反應(yīng),進(jìn)而影響蛋白激酶催化受體的磷酸化。分析這些受體的調(diào)節(jié)過程中涉及一些特異過程如受體的脫敏,內(nèi)化和下調(diào)的起始機(jī)制。通過體外實(shí)驗(yàn)結(jié)果顯示氨甲酰膽堿明顯增強(qiáng)m2受體的磷酸化,而阿托品或肝素(GRK2抑制劑)完全阻斷了m2受體的磷酸化。m2受體的磷酸化是依賴激活劑如氨甲酰膽堿作用發(fā)生的,呈明顯的劑量依賴關(guān)系;由GRK2介導(dǎo)的β2-AR的激活劑特布他林以及肝素對(duì)β2-AR的磷酸化的檢測(cè)結(jié)果顯示,特布他林增強(qiáng)腎上腺素能受體的磷酸化,而肝素可完全阻斷β2-AR的磷酸化;氨甲酰膽堿不能影響腎上腺素能受體的磷酸化,特布他林對(duì)mAChR2磷酸化也無明顯影響。這些結(jié)果證實(shí)氨甲酰膽堿對(duì)mAChR2,特布他林對(duì)β2-AR磷酸化的增強(qiáng)作用是選擇性的作用結(jié)果。進(jìn)一步采用HEL299細(xì)胞磷酸化實(shí)驗(yàn),結(jié)果顯示氨甲酰膽堿可以增強(qiáng)HEL299細(xì)胞m2受體的磷酸化,阿托品可以抑制HEL299細(xì)胞m2受體的磷酸化但不如體外實(shí)驗(yàn)作用明顯,揭示體內(nèi)m2受體的磷酸化過程可能受激動(dòng)劑和抑制劑共同調(diào)節(jié)。 體外和體內(nèi)實(shí)驗(yàn)證實(shí)了氨甲酰膽堿、特布他林和阿托品等一些m2受體和β2-AR的激活劑及抑制劑對(duì)M2受體和β2-AR磷酸化的調(diào)節(jié)作用。提示激活劑增強(qiáng)m2受體和β2-AR的磷酸化可能是這些受體活化一個(gè)重要部分,由此開始加速受體敏感性下調(diào),受體耐受性增強(qiáng)。
[Abstract]:In this experiment, the effects of carbachol, atropine and heparin on the phosphorylation of M2 receptor and the effect of terbutaline and heparin on the phosphorylation of beta 2-AR membrane protein were investigated with the purified acetylcholine acetylcholine M2 receptor (mAChR2) and beta 2 adrenergic receptor membrane protein (beta 2-AR). The regulation mechanism of G protein coupled receptor kinase 2 (GRK2) mediated cholinergic M2 receptor and beta 2-AR phosphorylation. It is discussed how stimulant activates the M2 receptor and beta 2-AR receptor, triggering cascade reaction involving receptor phosphorylation and further affecting the phosphorylation of protein kinase catalytic receptor. Some specific processes, such as the desensitization, internalization and downregulation of receptors, show that methylcholine significantly enhances the phosphorylation of M2 receptors, while atropine or heparin (GRK2 inhibitor) completely blocks the phosphorylation of the phosphorylated.M2 receptor of the M2 receptor, which is dependent on the action of activator such as carbachol. The dose dependence; the results of GRK2 mediated beta 2-AR activator terbutaline and heparin phosphorylation of beta 2-AR showed that terbutaline enhanced the phosphorylation of adrenergic receptors, while heparin could completely block the phosphorylation of beta 2-AR; carbachol did not affect the phosphorylation of adrenergic receptors, terbutaline against mAChR 2 phosphorylation has no obvious effect. These results confirm that carbacholine is selective to the enhanced effect of mAChR2 and terbutaline on the phosphorylation of beta 2-AR. Further use of HEL299 cell phosphorylation experiments shows that carbachol can enhance the phosphorylation of M2 receptor in HEL299 cells, and atropine can inhibit M2 receptor in HEL299 cells. The phosphorylation of the body is not as effective as that in vitro. It reveals that the phosphorylation of M2 receptors may be regulated by agonists and inhibitors.
In vitro and in vivo experiments have confirmed the regulatory effects of some M2 receptors and beta 2-AR activators and inhibitors on the phosphorylation of M2 receptors and beta 2-AR, which suggest that activators enhance the phosphorylation of M2 receptors and beta 2-AR, which may be a critical part of these receptors, which begin to accelerate the sensitivity of the receptor. Intolerance, enhanced receptor tolerance.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2008
【分類號(hào)】：R341

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