双光子显微镜检测活体小鼠脑内微循环技术的建立

In vivo imaging of blood flow using two-photon laser scanning fluorescent microscopy

目的:利用正置双光子显微镜系统和荧光探针标记技术,观察活体小鼠脑内血管的三维立体分布,建立测量单根毛细血管血流速度的新方法。方法:麻醉小鼠,制作活体小鼠颅骨开窗样本,尾静脉注射血浆标记物Texas-Red dextran,利用双光子显微镜z序列扫描检测脑内微循环系统的三维分布;利用线扫描测量毛细血管的血流速度。结果:通过双光子显微镜可以探测到脑内500μm处的血管分布和走向,图像清晰且信噪比高;通过计算单位时间内红细胞的运动距离测得毛细血管(直径≤6μm)的血流速度为(0.59±0.12)mm/s。结论:利用双光子显微镜观察脑内微循环系统技术平台初步建立,为基础研究和医药应用提供了在体实验依据。

Objective： To observe the three-dimensional distribution of vessels, and to establish a new method for measurement of blood flow velocity in mice cerebral cortex using two-photon laser scanning microscopy and fluorescence probe labeling technique. Methods： The mouse was made cranial window surgery and injected Texas-Red through tail vein after anesthetized. The three-dimensional imaging of vessel was obtained through z-stack scanning, and blood flow velocity was quantified through line scanning. Results： We could detect vascular distri- bution for more than 500 tan depth using two-photon microscopy. The velocity of blood flow was （0.59 ±0.12） mm/s in capillary. Conclu- sion： The method for observing the brain blood flow by two-photon microscopy was established, which could achieve quantification of single vascular blood flow velocity and provide experimental evidence for basic research and medical applications.