High Speed 3D Volumetric Imaging Microscopy
| Autor: | Chih-Wei Liu, 劉致維 |
|---|---|
| Rok vydání: | 2017 |
| Druh dokumentu: | 學位論文 ; thesis |
| Popis: | 105 Brain, which controls our behaviors, emotions, consciousness, memories and all other vital signs of human, is one of the most important organs in human body. Although the history of studying individual neuron behaviors is more than 100 years, our understanding toward brain function is still very limited. One of the key reasons is that brain function is not dictated by a single neuron, but emerges from the sophisticated connections among neurons, i.e. connectome. Therefore, recently there have been significant amount of researches toward the understanding of connectome. However, most current studies concentrate on “structural” connectome through anatomical approaches, while the development of “functional connectome”, though much more informative, is limited by technical difficulties to map the brain with high enough spatiotemporal resolution throughout the whole volume. In order to study the way how functional connectome works in brain of living animals, the required tool should exhibit spatial resolution less than ~μm, temporal imaging speed as high as ~ms, and imaging size approaching millimeter cubic in volume. Among current techniques, optical microscopy is the best candidate to match these specifications. It can achieve non-invasive detection and provide sub-μm spatial resolution. In addition, optical microscopy is compatible with a variety of probes such as voltage, calcium or metabolic indicators. However, optical microscopy still has its limitation. In deep tissue imaging, conventional wide-field optical microscopy suffers from strong scattering and lack of optical sectioning capability. Although two-photon laser scanning microscopy provides siginificant improvement on the issues, the imaging speed is limited. To acquire 3D volumetric image, it have to scan every pixel laterally on xy plane and then scan axially by moving objective lens or sample. During the past decade, several schemes have been demonstrated recently to boost 3D imaging speed for functional connectome study of brain in vivo, including piezoelectric translator, acousto-optic deflectors, spatial light modulator, light field microscopy, light sheet microscopy, liquid lens, etc. However, most of these techniques cannot enhance imaging speed on both lateral and axial directions simultaneously. In addition, some of these suffer from scattering from tissue because of its wide-field illumination or can only use for transparent samples. To overcome these issues, we have presented a powerful technique that combines high-speed axial scanning lens and multiphoton microscopy to increase scanning speed of 3D volumetric imaging down to sub-second scale. However, in the earlier system, even though the axial scanning speed is more than 100 kHz, the 3D volume imaging speed is limited by the single point scanning on the lateral plane. In this thesis, we developed a 32-beam multifocal system to significantly enhance the lateral scanning speed with 1-2 orders of magnitude. Combined with the high-speed axial scanning lens, the volumetric imaging speed approaches milliseconds scale, that is adequate to probe dynamics of action potential among neurons. In addtion, in order to acquire large amount of signal from fast imaging, we cooperate with SouthPort Co.Ltd. Taiwan to achieve high-speed data streaming and 3D image reconstruction. In this thesis, we present preliminary results to demonstarte the system performance. This powerful and innovative system is expected to greatly facilitate in vivo brain functional connectome studies in the near future. |
| Databáze: | Networked Digital Library of Theses & Dissertations |
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