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標題: | 超高速雙光子顯微鏡之電控及其應用 Digital architecture implementation of Ultra-high speed Two-photon fluorescence microscopy |
作者: | Han-Wee Chong 張漢維 |
指導教授: | 孫啟光(Chi-Kuang Sun) |
關鍵字: | 雙光子熒光顯微鏡,超高速,生醫影像,動作電位, 2PFM,Ultra-high speed,Voltage imaging,ASAPs,Action Potentials, |
出版年 : | 2020 |
學位: | 碩士 |
摘要: | 神經元之間通過動作電位交互作用,其所耗之時間往往發生在毫秒這個級距內。因此,我們需要可以提供高時間解析度的光學顯微鏡,用以研究神經元細胞之間的神經迴路。為此,我們需要具有上千赫茲掃描功能的雙光子熒光顯微鏡(2PFM),用以監控動作電位來描述神經元之間的電壓變化。此外,利用Accelerated Sensor of Action Potentials (ASAPs)與2PFM的結合,可以從大量清醒細胞中進行無創、高靈敏度和長時間的光學記錄;這將為理解神經系統如何在神經迴路和神經訊息編碼和處理訊息鋪平道路。本篇論文的重點將描述兩千赫茲分辨率的電壓影像。
本文中,為了進行原理驗證研究,我們首先演示我們的超高速2PFM數位電控成果實現及其特性。其次,我們也分享了毫秒級精度的ASAPs雙光子電壓影像。在這裡,利用我們擁有2k幀率顯微鏡的優勢,我們將其用以觀測清醒小鼠大腦中的神經元細胞。最後,以2k幀率的ASAPs結果,我們已經能夠獲得其動作電位。高時間解析度不僅提供了動作電位的更多細節,而且還提供了更高精準度。 使用這個平台,可以觀測到體內的動作電位。而且這超高時間解析度的研究,至今沒有學者公開發表過。我們的系統非常穩定、強大、完整、以及具有極大的成本效益。這平台為研究動物行為打下了強大的可能性。將來,不僅是神經元迴路的病理分析,這平台還可用以不同的生醫影像研究。 The neuronal communication through action potential and neurotransmitter release occurs on millisecond time scale. Thus, we need optical microscopy tools which can provide high temporal resolution to investigate activity of neuron cells and consequently, neuronal circuits. For this, we need ultrahigh speed sub-resolution optical imaging tools viz. two-photon fluoresce microscopy (2PFM) with kilohertz frequency scanning capability to monitor the action potential signal, depicting the voltage signaling in neurons. Consequently, combining the imaging of voltage sensor, Accelerated Sensor of Action Potentials (ASAPs) with high-speed 2PFM, allows non-invasive, ultrasensitive and chronic simultaneous optical recording from large populations of awake cells; which will pave the way in understanding the how nervous systems encode and process information at circuit and single-cell levels. However, the technological development in this regard is still to report the voltage imaging at kilohertz sub-resolution. In this thesis, for a proof-of-principle study, we, for the first time, will demonstrate our digital architecture implementation of Ultra-high speed 2PFM and its characteristics. Second, we will demonstrate ASAPs-based two-photon voltage imaging with millisecond-timescale precision. Here, taking the advantages of our microscopy with nearly 2k frame rate high-speed scanning capability, we will apply it on neuron cells observation in awake mice brain, particularly for visual cortical neurons. Finally, taking the 2k frame rate ASAPs results, we have been able to obtain its action potentials. The high temporal resolution of neuron observation not only provides more details of action potential, but also higher precision of action potential recording. With this platform, in vivo measurement for action potential could be thus observed, with time-resolution not yet reported. Our system is significantly robust, stable, compact, and cost-effective with comparable spatial resolution. This platform opens the possibility for detailed explorations of network dynamics in the context of behavior in animals. In the future, not only pathological features on neuronal circuit, the developed imaging platform will be further utilized for varied other investigations of biomedical importance. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/15383 |
DOI: | 10.6342/NTU202100397 |
全文授權: | 未授權 |
顯示於系所單位: | 光電工程學研究所 |
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U0001-0202202116254000.pdf 目前未授權公開取用 | 18.47 MB | Adobe PDF |
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