实时监测GHRP对心肌细胞内[Ca^(2+)]i和NO调控的双标记方法研究  

To Explore the Double-labeling Method of Monitoring the GHRP Regulatory Function on [Ca^(2+)]i and NO on Real Time in Cardiomyocytes Under LSCM

作　　者：李梅秀[1] 王艳茹[2] 曹济民[3] 田国忠[1] 赵玲辉[4] 

机构地区：[1]黑龙江省佳木斯大学基础医学院,154007 [2]北京大学分子医学研究所 [3]中国协和医科大学 [4]哈尔滨医科大学

出　　处：《医学研究杂志》2007年第8期22-25,共4页Journal of Medical Research

基　　金：黑龙江省卫生厅重点科研项目(2005-350);黑龙江省自然科学基金资助项目D2006-24与YZSCX2007-0077HLJ资助项目

摘　　要：目的探索实时监测GHRP对心肌细胞内[Ca2+]i和NO调控的双标记方法,为研究GHRP对心脏直接保护效应机制提供新方法。方法用改良的Langendorff恒流灌注仪系统和酶解法急性分离SD成年大鼠心肌细胞,在LSCM下分别以Ca2+探针Rhod-2/AM和NO探针DAF-FM/DA对心肌细胞内Ca2+和NO进行双标记并观察GHRP即时刺激对两者的调控影响。结果在LSCM下心肌细胞内Ca2+呈红色荧光,NO呈绿色荧光,两者双标记后呈现黄绿色荧光,GHRP使红色荧光出现瞬时增强后又迟缓地回降,绿色荧光强度没有明显变化,双标记下GHRP对两者的调控同单标记结果。结论LSCM下实验中设计的双标记方法能实时监测成年大鼠心肌细胞内Ca2+和NO存在及GHRP对两者同一时相调控作用,引起[Ca2+]i两个时相的变化,而对NO信号系统未见明显影响。Objective To explore the double - labeling method of monitoring the GHRP regulatory function on [ Ca^2＋ ] i and NO in cardiomyocytes of rats on real time under LSCM. Methods The reformed constant - flow Langendorff system and enzyme - dissociated was used to isolate cardiomyocytes. [ Ca^2＋ ] i and NO in the cardiomyocytes of SD rats were double - labeled by their molecular probe Rhod - 2/AM and DAF - FM/DA ,respectively to monitor the regulatory function of GHRP on [ Ca^2＋ ] i and NO on real time by LSCM. Results Ca^2＋ signal showed a red fluorescence and NO showed a green fluorescence while the overlapping of the two signals showed a yellow - green fluorescence by this system ,and the similar effect presents in both double -labeled state and the single labeled one： GHRP induced a transient[ Ca^2＋ ] i increase then followed by a plateau phase while there was not significant change in NO signal system after GHRP stimulation under the LSCM in the cardiomyocytes of rats. Conclusions After having established the double - labeling method we monitored the GHRP regulatory function on [ Ca^2＋ ] i and NO on real time in cardiomyocytes of rats under LSCM causing the [ Ca^2＋ ] i biphasic increase while no significant change in NO signal system .

关 键 词：心肌细胞 细胞内游离钙离子浓度 NO 双标记 GHRP LSCM 

分 类 号：R282.71[医药卫生—中药学]