2007 — 2010 |
Charles, Andrew Feldman, Jack (co-PI) [⬀] Bozovic, Dolores (co-PI) [⬀] Miao, Jianwei (co-PI) [⬀] Arisaka, Katsushi |
N/AActivity Code Description: No activity code was retrieved: click on the grant title for more information |
Mri: Development of a High-Speed Confocal Microscope For 4d Live-Cell Imaging @ University of California-Los Angeles
This award is for the development of a single-photon-sensitive confocal microscope, capable of true 4D (four dimensional: x, y, z, t) live imaging at video rate. It is based on an Image Intensified CMOS sensor (ICMOS) and a high-speed confocal scanner, which are designed to meet the following specifications: 1) High-speed (< 1 ms/frame), mega-pixel imaging with single-photon sensitivity. This is the same sensitivity as an EMCCD (Electron Multiplying CCD) but at one hundred times faster frame rate. 2) High-speed (< 1 ms/frame) confocal scanning for a single x-y focal plane. In addition, capability to scan in depth (z) up to 100 microns in 30 msec, resulting in a ''true 4D movie'' at video frame rate. 3) High-speed (< 10 ms/frame) FLIM (Fluorescence Lifetime Imaging Microscope) with < 100 psec lifetime resolution for a single x-y focal plane. This is a similar lifetime resolution as conventional scanning confocal microscopes but with a frame rate that is one hundred times faster. It will enable a ''true 4D FLIM movie'' at video frame rate. 4) Video-rate (~30 ms/frame) FLIM with true spectrum analysis for a single x-y focal plane. This new microscope may revolutionize the way millisecond time-scale phenomena are visualized in all biological systems, spanning from single molecules, single cells, and neural networks (such as the brain), to in vivo imaging of tissue in animals.
In addition to the scientific benefit of this new microscope, this award will contribute to multi-disciplinary education of students, at both the graduate and undergraduate level, at the forefront of biology, chemistry, physics, and engineering.
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2012 — 2014 |
Otis, Thomas [⬀] Arisaka, Katsushi |
N/AActivity Code Description: No activity code was retrieved: click on the grant title for more information |
Idbr: Development of a Streak Microscope For Measurement of Fast Multineuronal Signals @ University of California-Los Angeles
IDBR: Development of a streak microscope for measurement of fast multineuronal signals.
One of the greatest challenges in neuroscience is to understand how patterns of electrical activity in networks of neurons underlie brain function. Although the fundamental electrical signals (action potentials) and the basic hardware elements that generate these signals (neurons) are well known, the temporal (~ 1 kHz) and spatial (~10 micrometer) scales in which this signaling occurs prohibit parallel measurements from more than a small number of neurons. In recent years, new optical methods for tracking neural activity have been developed and offer great promise for overcoming some of these technical barriers. However, available microscopy methods and instrumentation are incapable of recording ensemble neural activity with sufficient spatial and temporal resolution. This proposal aims to address this issue directly by constructing a novel microscope optimized to record ensemble neuronal activity with temporal precision (> 8KHz) appropriate to resolve action potential activity in single neurons. The microscope¡¦s design is inspired by star trails observed in long exposure images of the night sky and is thus termed a fluorescent trails microscope (FTM). The FTM will be optimized to perform prolonged optical measurements of spatially distributed signals with submillisecond resolution. Rather than scanning a laser beam, the microscope will utilize a computer addressable diffractive element (a spatial light modulator or SLM) to generate continuous, patterned illumination of user-selected regions of interest. Images will be swept across a CCD or CMOS sensor in synch with the frame rate, allowing for the increased temporal resolution. This flexible design should be adaptable for use in a variety of experimental preparations including in vivo brain imaging and other biomedical applications involving time resolved fluorescence photometry. The proposed activity will foster specific cross-disciplinary, interactions and pre-college educational activities. Once fully developed, this technology will be used by a wide range of scientists at UCLA and beyond who are interested in measuring neuronal network activity as well as studying other biological phenomenon with collective properties. Construction of the instrument will involve recruitment of students in Physics from a nascent "Neurophysics Program" at UCLA who will be engaged in the interpretation of the resulting large quantity of experimental data requiring expertise in systems neurobiology and statistical physics. The project will provide a platform for students in multiple graduate disciplines (i.e., Physics, Neurobiology, Mathematics) to expand their knowledge base beyond the traditional boundaries of their respective fields as well as motivate campus wide interaction and collaboration. Lastly, there will be an opportunity for high school and undergraduate students to tour the laboratory and learn about the science behind the project, as well directly engage in the research, with a particular emphasis on recruiting those from underserved communities.
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2014 — 2016 |
Otis, Thomas (co-PI) [⬀] Bozovic, Dolores (co-PI) [⬀] Bentolila, Laurent Hallem, Elissa (co-PI) [⬀] Arisaka, Katsushi |
N/AActivity Code Description: No activity code was retrieved: click on the grant title for more information |
Idbr Type a: Development of a Line Confocal Bessel Beam Platform For High-Speed High-Volume 3d Imaging in Vivo @ University of California-Los Angeles
This NSF IDBR award is made to Prof. Katsushi Arisaka and collaborators at the University of California, Los Angeles, to develop a Bessel beam line confocal microscope. The goal of this project is to develop a system enabling the recording of three-dimensional cell structure in vivo, in real-time, with exceptional penetrating depth, and with minimal damage to the targeted sample. The proposed system will advance scientific understanding by facilitating cellular observation and systems level biologic analysis. Significantly, this system will enable the unprecedented high-speed recording of cell dynamics at super-resolution, in a temporal range permitting observation of developmental phenomena.
The broader impact is a cost-effective, easily configurable and user-friendly 3D imaging system for use by scientists towards the in vivo structural characterization of dynamic biological samples over a timespan of days. This collaboration will generate an available and reproducible microscope that significantly advances obtainable information concerning embryo development, dynamic neural network function, cellular differentiation and regulation, while enabling high-volume 3D cell imaging. This project directly integrates educational and research goals through incorporation of microscope development into a novel, interdisciplinary laboratory course developed by Dr. Arisaka at UCLA. Thus, development and construction of the system will serve as an educational platform directly fostering student learning. Moreover, the system will be housed in the Advanced Light Microscope Facility at the California NanoSystems Institute (CNSI), resulting in widespread availability to the extended scientific community.
This award is being made jointly by two Programs- (1) Instrument Development for Biological Research, in the Division of Biological Infrastructure (Biological Sciences Directorate), and (2) Biomedical Engineering, in the Division of Chemical, Bioengineering, Environmental and Transport Systems (Engineering Directorate).
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2017 — 2018 |
Arisaka, Katsushi Bentolila, Laurent |
R21Activity Code Description: To encourage the development of new research activities in categorical program areas. (Support generally is restricted in level of support and in time.) |
Trans-Sheet Illumination Microscopy (Transim) For Decoding Whole Brain Activity At Submillisecond Temporal Resolution @ University of California Los Angeles
Today?s knowledge of large-scale neural networks is advancing along two orthogonal directions. Spatial (static, structural) connectomic understanding is achieved through optical and electron microscopy; yielding high spatial resolution with limited or no temporal information. Conversely, electrophysiological methods provide an exceptional temporal understanding of millisecond-order neurodynamic activities in vivo, with restrictions placed on spatial information. Unfortunately, these approaches have historically been rather mutually exclusive and incompatible with each other. This proposal is precisely aimed at breaking the methodological barrier between spatial and temporal observation, through innovative 3D optical scanning concepts which rival the temporal resolution of electrophysiology. The resulting system is ideally suited to imaging genetically expressed voltage- sensitive fluorescent markers. The proposed optical microscope is named TranSIM: Trans-Sheet Illumination Microscope. It is designed to observe brain-wide neurodynamics in model organisms (up to 1 mm3) with ~1 ?m resolution in space, and sub- millisecond resolution in time. Unlike a conventional sheet illumination microscope, which illuminates a single x-y focal plane of the detection objective (on the z-axis), the conceived design forms a light sheet in the transverse direction along the z-axis (y-z plane). This sheet of light is then rapidly scanned along the x-axis as it is imaged. Multiple thin z-slices are then collected simultaneously by spatially multiplexing next-generation large-format sCMOS sensors. Regions of Interest, each with slight depth off-set from the focal plane, will map to a segmented part of the sensor, creating an eight-image focal volume in the time of a single frame. As a result, a thick brain volume can be covered by scanning the focal (x-y) plane (~1 mm2) in one direction (in x) at the frame rate of the sCMOS. The fastest sCMOS cameras are currently being jointly developed and optimized for this purpose. A line confocal readout will be implemented electrically by a rolling shutter mode of the sCMOS sensor to minimize contamination of scattering light. With proper Regions of Interest, a large volume (800 x 80 x 1000 ?m3) can be scanned at 780 volumes / second; 100 times faster than today?s fastest 3D optical microscopy systems. A smaller volume (100 x 80 x 1000 ?m3) can be scanned at an unprecedented rate of 6240 volumes / second, reaching the sub-millisecond temporal resolution of electrophysiological sampling. The illuminative sheet can be formed through the same detective objective for enhanced geometrical flexibility, or by an orthogonal objective (as in conventional horizontal sheet illumination) to minimize phototoxicity. In the case of a multi-view based on four-objective geometry, a large open volume (40 x 20 x 8 mm3) exists between lenses, to observe model organisms (like Zebrafish) under visual and optogenetic stimulation. Such a volume also provides smaller model organisms (C. elegans and Drosophila larvae) with ample space to navigate freely in 3D, while whole brain neural activity is monitored and controlled under various external and internal (optogenetic) stimulations.
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