小鼠胰腺腺泡细胞钙振荡的激光共聚焦成像研究方法

目的：建立一种简易高效的成年小鼠胰腺腺泡细胞钙振荡的激光共聚焦成像研究方法。方法：取成年昆明小鼠胰脏,用胶原酶法急性分离得胰腺腺泡细胞,加荧光染料fluo-4-AM标记胞内钙;以乙酰胆碱（ACh）为兴奋剂作用于胰腺腺泡细胞,激光扫描共聚焦显微镜发射488 nm激光并同步、实时、动态地记录此过程中胰腺腺泡细胞产生的钙振荡。结果：1一定浓度的ACh（如100 nmol/L）可稳定地激发胰腺腺泡细胞产生典型钙振荡,且此钙振荡可被阿托品完全阻断;2胞浆内不同部位的钙振荡有强弱不同,但呈同步变化节律;而不同细胞间的钙振荡强弱和节律往往不同;3钙振荡的幅度和节律与Ach具有剂量依赖效应。结论：小鼠胰腺腺泡细胞钙振荡的激光共聚焦成像研究方法简单易行、直观形象、高效、灵活,可作为常规方法使用,具有良好的应用前景和推广价值。

Laser scanning confocal microscopic imaging for Ca^2 ＋ oscillations of pancreatic acinar cells in mice

Objective： To establish a simple but effective method of laser scanning eonfoeal microscopic imaging for Ca^2＋ oseillations of pancreatic acinar ceils in adult mice. Methods： Pancreatic acinar ceils from adult Kunming mice were isolated acutely with collagenase, and then loaded with fluo-4-AM, a Ca^2＋ indicator. A laser scanning confocal microscope armed with 488 nm laser was employed to record the dynamic fluorescent signals in-time and synchronously while acetylcholine （ACh） was added in the pancreatic acinar cells. Results： ① The classic pancreatic acinar cell Ca^2＋ oscillations were induced by a certain concentration of ACh （100 nmol/L） successfully and steadily, which could be blocked by atropine completely. ② Plasmic Ca^2＋ oscillations from different parts of one acinar cell were usually with different amplitudes and almost the same frequencies. But both of amplitudes and frequencies were different among different cells. ③The acinar cell Ca^2＋ oscillations were induced by ACh in a concentration-dependent manner. Conclusion： The laser scanning confocal microscopic imaging for adult mouse pancreatic acinar cell Ca^2＋ oscillations was established successfully. The features of being easy to use, direct to see lively, high efficiency and good flexibility make it a popular tool for researchers to choose.