本文選題：視網(wǎng)膜色素上皮細(xì)胞 + SK通道�。� 參考：《延邊大學(xué)》2012年碩士論文

【摘要】：目的：觀察并了解嘌呤受體在視網(wǎng)膜色素上皮細(xì)胞功能的作用。 方法：應(yīng)用FURA/2熒光壽命成像顯微術(shù)檢測ARPE-19細(xì)胞內(nèi)的鈣離子濃度。并應(yīng)用RT-PCR實(shí)驗(yàn)測定BK,SK,IK通道的存在,然后使用內(nèi)向外式膜片實(shí)驗(yàn)進(jìn)一步鑒定BK,SK,IK通道的存在及電流密度和開放概率。最后,通過使用一種網(wǎng)格蛋白或小窩蛋白(caveolin-1)為介質(zhì)的運(yùn)輸機(jī)制,檢測ATP是否通過Gaq減少了細(xì)胞表面的Kir4.1豐富度,并采用western blot蛋白印記分析,來分析小窩蛋白和網(wǎng)格蛋白重鏈。 結(jié)果：實(shí)驗(yàn)結(jié)果表明,ATP的應(yīng)用能夠通過激活嘌呤受體增加RPE細(xì)胞內(nèi)的鈣離子濃度。增加的細(xì)胞內(nèi)鈣離子濃度能夠打開鈣離子激活鉀離子通道。許多組織中的SK通道是在P2Y受體與SK通道和P2Y受體的嘌呤刺激下激活的。P2Y受體與SK通道激活的偶聯(lián),是通過蛋白質(zhì)依賴性磷脂酶c的激活和IP3生產(chǎn)進(jìn)行的。IP3受體操縱儲(chǔ)備物引起的局部鈣離子釋放看來是SK的主要刺激。在這項(xiàng)研究里,能受體的激活不僅能引起B(yǎng)K通道開放概率的增加,還會(huì)引起SK通道開放概率的增加。SK通道的激活導(dǎo)致向外電流,降低細(xì)胞膜興奮性。在此我們還得出,通過一種小窩蛋白內(nèi)吞作用參與的機(jī)制,長期在ATP中培育會(huì)導(dǎo)致內(nèi)向整流鉀離子通道的電流密度降低。 結(jié)論：無論是對鈣離子激活的鉀通道,還是內(nèi)向整流鉀離子鉀離子通道,ATP都扮演一個(gè)很重要的角色。嘌呤激活可以增加SK和BK通道的開放概率,并能通過一種小窩蛋白內(nèi)吞作用參與的機(jī)制降低內(nèi)向整流鉀離子通道的電流密度。
[Abstract]:Objective: to investigate the function of purine receptor in retinal pigment epithelium (RPE) cells. Methods: calcium concentration in ARPE-19 cells was detected by FURA/2 fluorescence lifetime imaging microscopy. The existence of IK channel was determined by RT-PCR experiment, and the existence, current density and open probability of IK channel were further identified by internal and external diaphragm experiments. Finally, by using a transport mechanism of griddle protein or nest protein caveolin-1, we detected whether ATP reduced the abundance of Kir4.1 on cell surface through Gaq, and analyzed the heavy chain of nest protein and griddle protein by western blot protein imprinting. Results: the results showed that ATP could increase calcium concentration in RPE cells by activating purine receptors. Increased intracellular calcium concentration can open the calcium-activated potassium channel. SK channels in many tissues are coupled with SK channels activated by P2Y receptors and by purine stimulation of SK channels and P2Y receptors. The local calcium release induced by the manipulation of the. IP3 receptor reserve through the activation of protein-dependent phospholipase c and the production of IP3 appears to be the main stimulation of SK. In this study, activation of energy receptors not only increased the opening probability of BK channels, but also increased the opening probability of SK channels. The activation of SK channels led to outward currents and decreased membrane excitability. We also found that long-term cultivation in ATP resulted in a decrease in the current density of inward rectifier potassium channels through a mechanism involving endocytosis of fossa proteins. Conclusion: both calcium-activated potassium channel and inward rectifier potassium channel ATP play an important role. Purine activation can increase the opening probability of SK and BK channels, and decrease the current density of inward rectifier potassium channels through a mechanism involved in the endocytosis of nest proteins.
【學(xué)位授予單位】：延邊大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2012
【分類號】：R774.1

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