双光子显微镜用于在体监测小鼠视交叉上核Ca2+动态变化的方法学研究

摘要

Objective To explore a novel surgical method of skull base exposure and record calcium activity of SCN in vivo combining two-photon fluorescence imaging.Methods The surgery of skull base exposure was per formed on the anesthetized mice and the calcium signals of SCN were real-time monitored by two-photon fluorescence microscopy after calcium indicator injected.Results A set of operative methods to expose the SCN at skull base of mice were successfully established.After surgery,the calcium signals of SCN were clearly monitored with two-photon microscopy.The fluorescence signals of calcium did not quench for a long time along with the detecting time and laser intensity,and the intracellular calcium imagings in vivo were stably displayed.The activity routine of spontaneous calcium responses was different in recording neurons.Conclusion After the surgery of exposing skull base in mouse,the real-time detection of calciumsignals of neurons in SCN are successfully realized with two-photon microscopy.%目的 通过颅底暴露手术联合双光子荧光成像技术实现在体记录视交叉上核(SCN)神经元钙离子活动信号并研究其活动规律的新型方法.方法 对麻醉小鼠实行颅底暴露手术,通过双光子荧光显微镜对已注射钙指示剂的活体小鼠SCN核团钙信号进行实时动态测定.结果 成功建立一套小鼠颅底暴露SCN核团手术方法,术后应用双光子显微镜清晰地检测到SCN核团钙离子荧光信号,也能够实现稳定的在体细胞钙离子成像,并发现不同自发反应细胞中钙离子活动规律差异.结论 通过颅底暴露手术,利用双光子显微镜成功实现了实时监测SCN区域神经元钙离子信号.