發(fā)布時(shí)間：2018-12-25 13:01

【摘要】：目的:研究膽囊收縮素CCK-8(cholecystokinin-8;CCK-8)對(duì)小鼠背根神經(jīng)節(jié)(dorsal root ganglion,DRG)神經(jīng)元A-型鉀通道電流及興奮性作用和機(jī)制。方法:急性分離小鼠背根神經(jīng)節(jié)(DRG)神經(jīng)元,利用分子生物學(xué)方法鑒定其膜上CCK2受體的表達(dá)。利用全細(xì)胞膜片鉗技術(shù),研究CCK-8對(duì)小直徑DRG神經(jīng)元A-型鉀通道電流及興奮性的作用,并應(yīng)用藥理學(xué)方法研究其信號(hào)通路作用機(jī)制,最后在行為學(xué)的角度驗(yàn)證CCK-8的致痛作用。結(jié)果:本研究發(fā)現(xiàn):1)在蛋白水平上,利用western blot和免疫熒光技術(shù),小鼠背根神經(jīng)節(jié)上有CCK2受體表達(dá);2)電生理結(jié)果顯示,CCK-8對(duì)DRG神經(jīng)元上的A-型鉀通道電流的抑制作用具有劑量依賴關(guān)系。100 nM CCK-8對(duì)A-型鉀通道電流的抑制率約為25.9%;3)CCK-8對(duì)A-型鉀通道電流的抑制作用能夠被LY225910(CCK2R特異性阻斷劑)、GDP-β-S(特異性G蛋白活性抑制劑)、PTX(百日咳毒素)所阻斷。磷脂酰肌醇激酶(PI3K)的選擇性阻斷劑LY294002預(yù)先孵育背根神經(jīng)節(jié)細(xì)胞后,可以阻斷CCK-8對(duì)A-型鉀通道的抑制作用,而孵育蛋白激酶A(PKA)的選擇性阻斷劑PKI6-22、磷脂酶C(PLC)選擇性阻斷劑U73122和蛋白激酶C(PKC)的選擇性阻斷劑GF109203X、Chelerythrine chloride、RO31-8220,則無(wú)此效果。4)CCK-8顯著增加小直徑背根神經(jīng)節(jié)細(xì)胞的放電頻率,此作用可被4-AP阻斷。結(jié)論:CCK-8是通過(guò)作用于小鼠小直徑背根神經(jīng)節(jié)神經(jīng)元上的CCK2R來(lái)抑制A-型鉀通道的;其對(duì)A-型鉀通道電流的抑制作用是通過(guò)PTX敏感的磷脂酰肌醇激酶(PI3K)啟動(dòng)的信號(hào)通路;CCK-8可增加背根神經(jīng)節(jié)神經(jīng)元細(xì)胞膜的興奮性,進(jìn)而參與疼痛調(diào)節(jié)。
[Abstract]:Aim: to study the effect and mechanism of cholecystokinin CCK-8 (cholecystokinin-8;CCK-8) on A-type potassium channel current and excitability of (dorsal root ganglion,DRG neurons in mouse dorsal root ganglion (DRG). Methods: (DRG) neurons were isolated from mouse dorsal root ganglion (DRG) and the expression of CCK2 receptor on the membrane was identified by molecular biology. The effects of CCK-8 on the current and excitability of A- type potassium channels in small diameter DRG neurons were studied by whole-cell patch clamp technique, and the mechanism of its signaling pathway was studied by pharmacological method. Finally, the pain effect of CCK-8 was verified from the behavioral point of view. Results: 1) at the protein level, CCK2 receptors were expressed in the dorsal root ganglion of mice using western blot and immunofluorescence techniques. 2) the electrophysiological results showed that the inhibitory effect of CCK-8 on the current of A- type potassium channel in DRG neurons was dose-dependent, and the inhibitory rate of 100 nM CCK-8 on the current of A- type potassium channel was about 25.9%. 3) the inhibitory effect of CCK-8 on A-type potassium channel current was blocked by LY225910 (CCK2R specific blocker) and GDP- 尾 -S (specific G protein activity inhibitor), PTX (pertussis toxin). LY294002, a selective inhibitor of phosphatidylinositol kinase (PI3K), preincubated dorsal root ganglion cells (DRG), which blocked the inhibitory effect of CCK-8 on the A- type potassium channel, while PKI6-22, a selective inhibitor of protein kinase A (PKA), was incubated. Phospholipase C (PLC) selective blocker U73122 and protein kinase C (PKC) selective blocker GF109203X,Chelerythrine chloride,RO31-8220, had no such effect. 4) CCK-8 significantly increased the discharge frequency of small diameter dorsal root ganglion cells. This effect can be blocked by 4-AP. Conclusion: CCK-8 inhibits A-type potassium channels by acting on CCK2R in small diameter dorsal root ganglion neurons of mice. The inhibitory effect of A- type potassium channel current was initiated by PTX sensitive phosphatidylinositol kinase (PI3K), and CCK-8 increased the excitability of dorsal root ganglion cell membrane and involved in pain regulation.
【學(xué)位授予單位】：蘇州大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2016
【分類號(hào)】：R402

【相似文獻(xiàn)】

相關(guān)期刊論文 前10條

1 章小莉;汪琳;徐龍;鄒麗;;白術(shù)對(duì)人妊娠子宮平滑肌細(xì)胞膜鈣依賴鉀通道電流的影響[J];中國(guó)婦幼保健;2009年03期

2 張國(guó)紅;耿仙;張璇;楊冀杰;張海林;;雙苯氟嗪對(duì)爪蟾卵母細(xì)胞上表達(dá)的hERG鉀通道電流的影響[J];中國(guó)藥理學(xué)通報(bào);2009年01期

3 張國(guó)紅;耿仙;趙志英;王娜;貝俊杰;張海林;;雙苯氟嗪對(duì)表達(dá)于爪蟾卵母細(xì)胞上的KCNQ1/KCNE1鉀通道電流的影響[J];中國(guó)藥理學(xué)與毒理學(xué)雜志;2007年01期

4 徐龍,余保平,陳明鍇,董衛(wèi)國(guó),羅和生;黃體酮對(duì)豚鼠結(jié)腸平滑肌及細(xì)胞膜鈣依賴的鉀通道電流的影響[J];中國(guó)藥理學(xué)通報(bào);2003年09期

5 賈慶忠,賈占峰,張英俊,劉伯一,張海林;KCNQ2/3鉀通道電流特征及M_1受體的調(diào)節(jié)作用[J];中國(guó)藥理學(xué)通報(bào);2005年10期

6 倪錦,劉洋,張貞雄,唐顯玲,劉進(jìn);異丙酚對(duì)人腸系膜小動(dòng)脈平滑肌細(xì)胞膜外向鉀通道電流的影響[J];廣州醫(yī)學(xué)院學(xué)報(bào);2004年01期

7 林吉進(jìn);劉樹楷;鄭方芳;馬青艷;余宏;任莉;沈心遠(yuǎn);;蛋白酪氨酸磷酸酶非受體型12負(fù)性調(diào)控心臟HERG鉀通道電流[J];南方醫(yī)科大學(xué)學(xué)報(bào);2013年12期

8 宿燕崗;心肌細(xì)胞感染病毒后鈣、鉀通道電流及其表達(dá)改變和�；撬岬淖饔�(摘要)[J];復(fù)旦學(xué)報(bào)(醫(yī)學(xué)版);2003年05期

9 公英子;丁維光;吳捷;松浦博;徎US子;伊藤英j9;李學(xué)奇;堀江稔;;二十碳四烯酸對(duì)轉(zhuǎn)基因中國(guó)倉(cāng)鼠卵母細(xì)胞hKv1.5鉀通道電流的作用[J];中國(guó)心臟起搏與心電生理雜志;2008年03期

10 丁超;傅向華;何振山;陳會(huì)校;薛玲;;辛伐他汀對(duì)血脂正常兔心肌缺血再灌注后瞬間外向鉀通道電流的影響[J];中國(guó)病理生理雜志;2009年02期

相關(guān)會(huì)議論文 前2條

1 丁超;傅向華;趙玉英;王冬梅;張莉;薛玲;;山莨菪堿對(duì)兔缺血再灌注后心室肌細(xì)胞瞬間外向鉀通道電流的影響[A];第十三次全國(guó)心血管病學(xué)術(shù)會(huì)議論文集[C];2011年

2 王騰;唐其柱;江洪;李建軍;黃從新;李庚山;;丹參注射液對(duì)缺血預(yù)處理引起心室肌細(xì)胞動(dòng)作電位和ATP敏感性鉀通道電流變化的影響[A];中華醫(yī)學(xué)會(huì)心血管病分會(huì)第八次全國(guó)心血管病學(xué)術(shù)會(huì)議匯編[C];2004年

相關(guān)碩士學(xué)位論文 前3條

1 趙先洋;膽囊收縮素CCK-8對(duì)小鼠背根神經(jīng)元A-型鉀通道電流作用及機(jī)制研究[D];蘇州大學(xué);2016年

2 鄭守超;雌二醇抑制人成骨樣細(xì)胞MG-63外向電壓門控鉀通道電流[D];華中科技大學(xué);2013年

3 楊麗;低頻電磁場(chǎng)對(duì)細(xì)胞膜電特性的影響[D];山西大學(xué);2010年

，

本文編號(hào)：2391163