Cellular and Molecular Imaging of Synaptic Vesicle Dynamics

我们正在进行一项成像计划，利用最新的超材料超透镜技术建立一个超透镜纳米显微镜，以实现卓越的时间和空间分辨率，并研究脑突触传递和细胞内吞/胞外作用过程中的膜动力学。超透镜由专门设计和微加工的结构元件组成，可以在亚衍射极限空间分辨率下放大近场图像，并实时投影远场高分辨率图像（Yao等，2008,Science, 321:930； Liu等，2007,Science, 315:1686； Fang等，2005,Science, 308:534-7）。与其他成像技术如EM、STED （Hell, 2007, Science, 316: 1153-8）和STORM/PALM （Huang, et al., 2008, Science, 319:810-813; Bates, et al., 2007, Science 317, 1749-53; Hess, et al., 2007, PNAS 104, 17370-5; Betzig, et al, 2006, Science 313:1642）相比，这种形式的光学纳米镜在实时光学成像（视频速率）和纳米空间分辨率方面具有一些独特的优势。在孙成教授（西北大学的校外合作者）和张旭日教授（加州大学伯克利分校）的领导下，超透镜相关技术正在快速发展。

We are undertaking an imaging initiative to build a hyperlens nanoscope, using newly advanced meta-material hyperlens technology, to achieve superior temporal and spatial resolution and to investigate membrane dynamics during brain synaptic transmission and cellular endocytosis/exocytosis in general. The hyperlens consists of specially designed and micro-fabricated structure elements that can magnify near-field images at sub-diffraction-limited spatial resolution and project the high resolution image at far-field in real time (Yao, et al, 2008, Science, 321:930; Liu, et. al, 2007, Science, 315:1686; Fang, et. al, 2005, Science, 308:534-7). This form of optical nanoscope possesses some unique advantages of both real-time optical imaging (at video rate) and nano-metric spatial resolution, in comparison with other imaging techniques such as EM, STED (Hell, 2007, Science, 316: 1153-8), and STORM/PALM (Huang, et al., 2008, Science, 319:810-813; Bates, et. al, 2007, Science 317, 1749-53; Hess, et al, 2007, PNAS 104, 17370-5; Betzig, et. al, 2006, Science 313:1642). Hyperlens-related technology is under rapid development led by the groups of Prof. Cheng Sun (extramural collaborator now at Northwestern University) and Prof. X. Zhang (UC Berkeley). After winning an equipment grant from the trans-NIH imaging initiative, we have been developing our imaging setup at NIH and are working to resolve modeling and nano-manufacturing issues toward improved hyperlens design and construction. In this project year, we have also investigated bovine chromaffin cells (fixed or live, or transfected with various fluorescent protein constructs) as a vesicular trafficking model system, by leveraging our related experience on multimodal instrumentation around atomic force microscopy (AFM) and various optical microscopy platforms. We are still working toward breakthrough investigations of sub-cellular membrane dynamics for both fundamental biology and medicine.