Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Download Make Life Visible
Document related concepts
Molecular neuroscience wikipedia , lookup
Neural engineering wikipedia , lookup
Brain morphometry wikipedia , lookup
Feature detection (nervous system) wikipedia , lookup
Biochemistry of Alzheimer's disease wikipedia , lookup
Subventricular zone wikipedia , lookup
Development of the nervous system wikipedia , lookup
Psychoneuroimmunology wikipedia , lookup
Metastability in the brain wikipedia , lookup
Haemodynamic response wikipedia , lookup
Optogenetics wikipedia , lookup
Neuroanatomy wikipedia , lookup
Clinical neurochemistry wikipedia , lookup
Channelrhodopsin wikipedia , lookup
Transcript
Make Life Visible THE 12th UEHARA INTERNATIONAL SYMPOSIUM 2017 Make Life Visible PROGRAM ＆ ABSTRACTS June 12 - 14, 2017 Hyatt Regency Tokyo Sponsored by THE UEHARA MEMORIAL FOUNDATION 1 Make Life Visible We are happy to announce the international symposium “Make Life Visible” sponsored by the Uehara Memorial Foundation, will be held from June 12 to 14, 2017. In recent years, marked advances in imaging technology have enabled the visualization of phenomena formerly believed to be completely impossible. These technologies have made major contributions to the elucidation of the pathology of diseases as well as to their diagnosis and therapy. Adding further promise for future development are imaging tools in the broad sense, such as optics and optogenetics - the revolutionary use of light to control cells and organisms. From molecular imaging to clinical images, the Japanese are world leaders in basic and clinical research of visualization. We strive to foster innovative, creative, advanced research that gives full play to imaging technology in the broad sense, while exploring cross-disciplinary areas in which individual research fields interact, and pursuing the development of new techniques where they fuse together. The 9th Specific Research Project, “Make Life Visible” was established by the Uehara Memorial Foundation as a 3-year research project to support such research. In this Special Project, three areas (Session 1-3) were targeted from basic research to clinical application. Nineteen Japanese researchers were selected, and research was begun in 2015. Session 1. Visualizing and Controlling Molecules for Life Session 2. Imaging Disease Mechanisms Session 3. Imaging-based Diagnosis and Therapy In this international symposium, we will build on the outcomes of the 9th Special Project, with presentations focusing on the cutting-edge findings of visualization technologies by the 19 Japanese Special Project members as well as 10 leading researchers invited from overseas. We look forward to this symposium being a forum for the presentation of the latest research outcomes, future prospects, and new strategies in visualization technology, from basic research to the clinical front lines (diagnosis and treatment). Yoshiaki TOYAMA, M.D., Ph.D. Chair of the Organizing Committee (Vice-President, Keio; Professor emeritus, Keio University) 2 Make Life Visible ORGANIZING COMMITTEE Honorary Chair: Shigetada Nakanishi (Kyoto Univ.) Chair: Vice-Chair: Members: Yoshiaki Toyama (Keio Univ.) Atsushi Miyawaki (RIKEN BSI) Masaya Nakamura (Keio Univ.) Masahiro Jinzaki (Keio Univ.) Kazuo Funabiki (IBRI Laboratory) Yasunori Urano (The Univ. of Tokyo) Yasuhisa Fujibayashi (NIRS) ADVISORY BOARD Chairperson: Akira Uehara (President, The Uehara Memorial Foundation; President and CEO, Taisho Pharmaceutical Holdings Co., Ltd.) Members of the Board Shigeru Uehara (Chairperson of Councilor, The Uehara Memorial Foundation; President, Taisho Pharmaceutical Co., Ltd.) Akira Ohira (Councilor, The Uehara Memorial Foundation; President, Taisho Toyama Pharmaceutical Co., Ltd.) Ken Uehara (Managing Director; The Uehara Memorial Foundation; Vice-President, Taisho Pharmaceutical Co.Ltd.) 3 Make Life Visible General Information 1. Venue for Symposium The symposium will be held in the “Century Room” on the B1 floor of the Hyatt Regency Tokyo. 2-7-2 Nishi-Shinjuku, Shinjuku-Ku, Tokyo, Japan 160-0023 Phone: +81-3-3348-1234 Fax: +81-3-3344-5575 2. Symposium Desk The Symposium Desk for registration will be open from 8: 00 on Monday, June 15 in front of the “Century Room” and remain open during the Symposium. General and travel information will be also given at the Symposium Desk. 3. Name Plates Please wear your name plate at all times during the Symposium. In case you misplace your name plate, please contact the Symposium Desk. Colors of the name plate indicate: Committee Members and Speakers ………………………………Red Participants ……………………………………………………… Blue Members of board of the Foundation and Interests .……………… White Secretariat …………………………………….………………… XXX 4. Symposium Office Symposium Office will be located in the “Yoshino” room on the B1 floor of the Hotel, and take charge of custody of messages and lost articles, and others. 5. Official Language The official language of the Symposium is English. Simultaneous translation is not provided. 6. Notice Cameras, video cameras, tape recorders or any other recording devices are not allowed at any session. 7. Publication The proceedings of the Symposium will be published after the meeting from Springer Japan K.K. 8. Correspondence or Inquiries after the Symposium All correspondence or inquiries concerning the Symposium should be sent to: Secretariat of the Symposium The Uehara Memorial Foundation Takada 3-26-3, Toshima-ku, Tokyo, Japan 171-0033 Phone: +81-3-3985-3500, 8400, Fax : +81-3-3982-5613 E-mail: [email protected] 4 Make Life Visible SCHEDULE DAY 1 June 12 (Mon) 9:00 - 9:05 9:05 - 9:10 9:10 - 9:30 9:30 - 10:20 10:20 - 18:35 Welcome Address Opening Remarks Opening Lecture Keynote Address (Session II) Session II: Imaging Disease Mechanisms Session III :Imaging-based Diagnosis and Therapy Session I : Visualizing and Controlling Molecules for Life 18:45 - 20:00 DAY 2 June 13 (Tue) 9:00 - 12:05 13:05 - 13:55 13:55 - 17:20 MIXER ( For ALL participants, B1 Floor Room “CRYSTAL” ) 18:10 - 20:00 DAY 3 June 14 (Wed) 9:00 - 9:50 9:50 - 12:15 FORMAL RECEPTION ( INVITATION ONLY) 12:15 - 12:20 Session II: Imaging Disease Mechanisms Keynote Address (Session I) Session I : Visualizing and Controlling Molecules for Life Session III :Imaging-based Diagnosis and Therapy Keynote Address (Session III) Session III :Imaging-based Diagnosis and Therapy Session I : Visualizing and Controlling Molecules for Life Closing Remarks 5 Make Life Visible SPEAKERS & LECTURE TITLES Session 1: Visualizing and Controlling Molecules for Life Title S1-1 (Invited) S1-2 S1-3 (Invited) Lihong V. WANG Itaru IMAYOSHI Scott E. FRASER California Institute of Technology, USA Kyoto University University of Southern California, USA Date Photoacoustic Tomography: Deep Tissue Imaging by Ultrasonically Beating Optical Diffusion Regulatory Mechanism of Neural Stem Cells Revealed by Optical Manipulation of Gene Expressions Day1 Eavesdropping on Biological Processes with Multi-Dimensional Molecular Imaging Apical microtubules define the function of epithelial cell sheets consisting of non-ciliated or S1-4 Sachiko TSUKITA Osaka University Keynote Karl Deisseroth Stanford University, USA Illuminating the brain S1-5 Tomomi KIYOMITSU Nagoya University Optogenetic assemblies of cortical force-generating complexes during mitosis S1-6 Kazuya KIKUCHI Osaka University in vivo Imaging Probes with Tunable Chemical Switches S1-7 Kazuo FUNABIKI multi-ciliated cells Day2 S1-8 (Invited) S1-9 S1-10 Evan W. MILLER Sotaro UEMURA Atsushi MIYAWAKI Institute of Biomedical Research and Innovation University of California, Berkeley, USA Circuit-dependent striatal PKA and ERK signaling underlying action selection Electrophysiology, Unplugged: New Chemical Tools to Image Voltage The University of Tokyo Single Cell Analysis of Stimulated Immune Cells with Real-time Selection RIKEN Brain Science Comprehensive approaches Institute using luminescence to studies of cellular functions Day3 Session 2: Imaging Disease Mechanisms Title Date Northwestern University Keynote A. Vania Apkarian Feinberg School of Make Chronic Pain Visible Medicine, USA S2-1 Masaya NAKAMURA Keio University S2-2 Nicholas SMITH Osaka University S2-3 James T PEARSON Mikiyasu SHRAI Research Institute of National Cerebral and Cardiovascular Center S2-4 Ulrich H. VON Harvard Medical School, (Invited) ANDRIAN USA S2-5 Masashi Yanagisawa Tsukuba University S2-6 (Invited) Cortical plasticity after spinal cord injury using resting-state functional magnetic resonance imaging Multimodal Label-free imaging to assess compositional and morphological changes in cells Day1 during immune activation Investigating in vivo myocardial and coronary molecular pathophysiology in mice with X-ray radiation imaging approaches Visualizing the Immune Response to Infections Imaging Sleep and Wakefulness Stanford University and Mark J. Schnitzer Howard Hughes Medical Optical imaging of large-scale neural codes and voltage dynamics in behaving animals Day2 Institute, USA S2-7 Motomasa TANAKA S2-8 Shigeo OKABE RIKEN Brain Science Abnormal local translation in dendrites impairs cognitive functions in neuropsychiatric Institute disorders The University of Tokyo Imaging synapse formation and remodeling in vitro and in vivo Session 3: Imaging-based Diagnosis and Therapy Title S3-1 (Invited) S3-2 S3-3 (Invited) Masaru Ishii Hisataka Kobayashi NeuroSpin, CEA Saclay Center, FRANCE Osaka University National Cancer Institute, NIH, USA RIKEN Center for Life Intravital multiphoton imaging revealing cellular dynamics in vivo Theranostic Near Infrared Photoimmunotherapy for Cancer Yasuyoshi WATANABE S3-5 Yasuteru URANO The University of Tokyo Geoffrey D. Rubin Duke University, USA Coronary Heart Disease Diagnosis: Engineering Triumphs, Economic Barriers Kenji KABASHIMA Kyoto University Live imaging of the skin immune responses (Invited) S3-7 Science Technologies S3-8 Masahiro JINZAKI Keio University Keynote Sanjiv Sam Gambhir Stanford University, USA S3-9 Yasuhisa FUJIBAYASHI National Institute of Radiological Sciences Date How MRI makes the Brain Visible S3-4 S3-6 6 Denis Le Bihan Day1 Novel and integrated imaging on Chronic Fatigue Novel fluorescent probes for rapid tumor imaging and fast glutathione dynamics Day2 Development of a horizontal CT and its application to musculoskeletal disease The Future of Precision Health & Integrated Diagnostics Day3 Imaging and therapy against hypoxic tumors with 64Cu-ATSM Make Life Visible SYMPOSIUM VENUE Hyatt Regency Tokyo 2-7-2 Nishi-Shinjuku, Shinjuku-Ku, Tokyo, Japan, 160-0023 Tel: +81 3 3348 1234, Fax: +81 3 3344 5575, [email protected] Acsess MAP: Airport Limousine Bus for Narita International Airport (NRT) (2 hours) ・ Narita Airport to hotel - 26 trips daily from 7:05 am to 10:30 pm ・ Hotel to Narita Airport - 16 trips daily from 5:50 am to 5:35 pm ・ One-way adult fare - JPY 3100 ・ Airport Limousine Bus for NRT Time Table(PDF) Airport Limousine Bus for Haneda Domestic and International Airport (HND) (1 hour 20 minutes) ・ Haneda Airport to hotel - 21 trips daily from 5:45 am to 11:10 pm ・ Hotel to Haneda Airport - 20 trips daily from 4:25 am to 11:35 pm ・ One-way adult fare - JPY 1230 ・ Airport Limousine Bus for HND Time Table(PDF) For inquiries, please contact the Hotel Concierge Desk：Tel: +81 3 3348 1234/Fax: +81 3 3344 5575. 7 Make Life Visible Symposium Site Symposium: MIXER: B1 Room CENTURY B1 Room CRYSTAL (Day1, 9:00 ~ Day3, 12:30) (Day1, 18:45 ~ 20:00 ) B1 Floor of the Hyatt Regency Tokyo CENTURY 8 Make Life Visible Symposium Program 9 Make Life Visible Outline of Program Make Life Visible ※Keyn o t e Adress: 50min Invited Lecture: 40min Lecture: 30min Session Ⅰ:Visualizing and Controlling Molecules for Life 生命現象の可視化・操作 Session Ⅱ:Imaging Disease Mechanisms 病態の可視化 Session Ⅲ:Imaging-based Diagnosis and Therapy 可視化技術の診断・治療応用 Time June 12 (Mon) 9:00 9:00 Welcome Address Akira Uehara 9:05 Opening Remarks S.Nakanishi 9:10-9:30 Opening Lecture Yoshiaki Toyama 9:30-10:20 SessionⅡKeynote A. Vania Apkarian 10:00 10:20-10:50 Masaya Nakamura ② June 13 (Tues) 9:00-9:40 Ulrich H. von Andrian 9:40-10:10 Masashi Yanagisawa 12:00 ② ② ③ 9:50-10:20 Yasuhisa Fujibayashi 10:20-10:35 Coffee Break 10:25-11:05 Mark J. Schnitzer 11:00 11:35-12:05 James T Pearson (Mikiyasu Shirai) 12:05-13:05 Lunch 9:00-9:50 SessionⅢ Keynote Sanjiv Sam Gambhir 10:10-10:25 Coffee Break 10:50-11:05 Coffee Break 11:05-11:35 Nicholas Isaac Smith June 14 (Wed) ② 11:05- 11:35 Motomasa Tanaka ① 11:15 - 11:45 Sotaro Uemura 11:35-12:05 Shigeo Okabe 12:05-13:05 10:35 - 11:15 Evan W. Miller 11:45 - 12:15 Atsushi Miyawaki Lunch 12:15 Closing Remarks Y.Toyama END 13:30 13:00 13:05-13:45 Denis Le Bihan 14:00 ③ 13:45-14:15 Masaru Ishii 13:55-14:25 Tomomi Kiyomitsu 14:15-14:55 Hisataka Kobayashi 15:00 16:00 ① 14:25-14:55 Kazuya Kikuchi 14:55-15:25 Yasuyoshi Watanabe 14:55-15:25 Kazuo Funabiki 15:25-15:55 Yasuteru Urano 15:25-15:40 Coffee Break 15:40 - 16:20 Geoffrey D. Rubin 15:55-16:15 Coffee Break 16:15-16:55 Lihong V. Wang 17:00 13:05-13:55 SessionⅠKeynote Karl Deisseroth 16:55-17:25 Itaru Imayoshi 17:25-18:05 Scott E. Fraser ① ③ 16:20 - 16:50 Kenji Kabashima 16:50 - 17:20 Masahiro Jinzaki End: 17:20 （photography 講演者記念写真） 18:00 18:05-18:35 Sachiko Tsukita 18:10~ Formal Reception ( INVITATION Only) 18:45~ Mixer ( for ALL participants) 19:00 10 End: 20:00 ① SessionⅠ ② SessionⅡ ③ SessionⅢ Make Life Visible KEYNOTE ADDRESS Day 1 Day 2 Day 3 SESSION II KEYNOTE 9:30 - 10:20 A. Vania Apkarian SESSION I KEYNOTE 13:05 - 13:55 Karl Deisseroth SRSSION III KEYNOTE 9:00 - 9:50 Sanjiv Sam Gambhir 11 Make Life Visible SESSION I: KEYNOTE ADDRESS Illuminating the brain Karl Deisseroth D.H. Chen Professor of Bioengineering and of Psychiatry and Behavioral Sciences, Stanford University Deisseroth will discuss the development of optogenetics (a technology for controlling millisecond-scale activity patterns in specific cell types using microbial opsin genes and fiberoptic-based neural interfaces, all in freely-behaving animals including adult mammals). He will also speak on the development of hydrogel-tissue composites (e.g. his CLARITY method for creating composites of biological molecules in tissue covalently linked to acrylamide-based hydrogels, allowing removal of unlinked tissue elements to create transparency and accessibility to macromolecular labels; the resulting new structure allows high-resolution optical access to structural and molecular detail within intact tissues without disassembly). He will also discuss the application of these methods to discover the neural cell types and connections that cause adaptive and maladaptive behaviors. 12 Make Life Visible SESSION II: KEYNOTE ADDRESS Make Chronic Pain Visible A. Vania Apkarian Department of Physiology, Northwestern University Feinberg School of Medicine, Chicago, Illinois, 60610, USA Complementary human and animal studies now provide convincing evidence regarding brain mechanisms involved in the development of chronic pain. Multi-modal human brain imaging studies now show that that brain properties determine risk for development of chronic pain, and also brain anatomy and functional responses and connectivity carve the chronic pain state. Consistent and complimentary evidence is now also available in rodent models of chronic pain, where brain reorganization is now unraveled for receptor gene expression, cellular excitability, and synaptic efficacy, as well as whole-brain functional connectivity, all of which are beginning to unravel novel targets for drug development. I will review these concepts with especial emphasis on mechanistic concepts and clinical implications of these exciting new advances in the field. The figure below illustrates the overall view regarding transition to chronic pain: 13 Make Life Visible SESSION III: KEYNOTE ADDRESS The Future of Precision Health & Integrated Diagnostics Sanjiv Sam Gambhir MD, PhD Stanford University Most of the world’s health care systems are focused on patients after they present with disease, and not before. While precision medicine uses personalized information to more effectively treat disease, the emerging field of precision health is situated to help assess disease risks, perform customized disease monitoring, and facilitate disease prevention and earlier disease detection. Currently an individual’s health is evaluated only a few times a year if at all, making it difficult to gather the amount of information needed to implement precision health. The emergence of continuous health monitoring devices with combined in vitro and in vivo (integrated) diagnostics, worn on the body and used in the home, will enable a clearer picture of human health and disease. However, challenges lie ahead in developing and validating novel monitoring technologies, and in optimizing data analytics to extract meaningful and actionable conclusions from continuous health data. This presentation will show some of the emerging technologies for diagnostics with a focus on cancer and the challenges to making precision health a reality in the decades to come. 14 Make Life Visible Session I DAY 1 DAY 2 DAY 3 SI-1. 16:15 - 16:55 Lihong V. WANG KEYNOTE 13:05-13:55 Karl Deisseroth S1-8 10:35 - 11:15 Evan W. MILLER S1-2 16:55-17:25 Itaru IMAYOSHI S1-5 13:55-14:25 Tomomi KIYOMITSU S1-9 11:15 - 11:45 Sotaro UEMURA S1-3 17:25-18:05 Scott E. FRASER S1-6 14:25-14:55 Kazuya KIKUCHI S1-10 11:45 - 12:15 Atsushi MIYAWAKI S1-4 18:05-18:35 Sachiko TSUKITA S1-7 14:55-15:25 Kazuo FUNABIKI 15 Make Life Visible SI-1 Photoacoustic Tomography: Deep Tissue Imaging by Ultrasonically Beating Optical Diffusion Lihong V. WANG California Institute of Technology, Pasadena, California, USA Photoacoustic tomography has been developed for in vivo functional, metabolic, molecular, and histologic imaging by physically combining optical and ultrasonic waves. Broad applications include early-cancer detection and brain imaging. High-resolution optical imaging—such as confocal microscopy, two-photon microscopy, and optical coherence tomography—is limited to superficial imaging within the optical diffusion limit (~1 mm in the skin) of the surface of scattering tissue. By synergistically combining light and sound, photoacoustic tomography provides deep penetration at high ultrasonic resolution and high optical contrast. In photoacoustic computed tomography, a pulsed broad laser beam illuminates the biological tissue to generate a small but rapid temperature rise, which leads to emission of ultrasonic waves due to thermoelastic expansion. The unscattered pulsed ultrasonic waves are then detected by ultrasonic transducers. High-resolution tomographic images of optical contrast are then formed through image reconstruction. Endogenous optical contrast can be used to quantify the concentration of total hemoglobin, the oxygen saturation of hemoglobin, and the concentration of melanin. Exogenous optical contrast can be used to provide molecular imaging and reporter gene imaging as well as glucose-uptake imaging. In photoacoustic microscopy, a pulsed laser beam is delivered into the biological tissue to generate ultrasonic waves, which are then detected with a focused ultrasonic transducer to form a depth resolved 1D image. Raster scanning yields 3D high-resolution tomographic images. Super-depths beyond the optical diffusion limit have been reached with high spatial resolution. The following image of a mouse brain was acquired in vivo with intact skull using optical-resolution photoacoustic microscopy. The annual conference on photoacoustic tomography has become the largest in SPIE’s 20,000-attendee Photonics West since 2010. Wavefront engineering and compressed ultrafast photography will be touched upon. Selected publications: 1. Nature Biotechnology 21, 803 (2003). 2. Nature Photonics 5, 154 (2011). 3. Science 335, 1458 (2012). 4. Nature Methods 13, 67 (2016). 16 Make Life Visible SI-2 Regulatory Mechanism of Neural Stem Cells Revealed by Optical Manipulation of Gene Expressions Itaru IMAYOSHI Graduate School of Biostudies, Kyoto University, Kyoto, Japan The mammalian brain consists of a complex ensemble of neurons and glial cells. Their production during development and remodeling is tightly controlled by various regulatory mechanisms in neural stem cells. Among such regulations, basic helix-loop-helix (bHLH) factors have key functions in the self-renewal, multipotency, and fate determination of neural stem cells. Here, we highlight the importance of the expression dynamics of bHLH factors in these processes. We propose the multipotent state correlates with oscillatory expression of several bHLH factors, whereas the differentiated state correlates with sustained expression of a single bHLH factor. We also developed a new optogenetic method that can manipulate gene expressions in neural stem cells by light. We used this technology to manipulate the growth and fate-determination of neural stem cells. I also introduce various applications of light-induced control of gene expressions in broad fields of biology. Multi-potency and fate-determination of neural stem cells by bHLH factors. Manipulation of gene expressions by blue light. Recent Publications: Imayoshi, I. and Kageyama, R. (2014) bHLH Factors in Self-Renewal, Multipotency, and Fate Choice of Neural Progenitor Cells. Neuron 82: 9-23. Imayoshi, I., Isomura, A., Harima, Y., Kawaguchi, K., Kori, H., Miyachi, H., Fujiwara, T.K., Ishidate, F. and Kageyama, R. (2013) Oscillatory control of factors determining multipotency and fate in mouse neural progenitors. Science (Research Article) 342: 1203-1208. Imayoshi, I., Sakamoto, M., Ohtsuka, T., Takao, K., Miyakawa, T., Yamaguchi, M., Mori, K., Ikeda, T., Itohara, S. and Kageyama, R. (2008) Roles of continuous neurogenesis in the structural and functional integrity of the adult forebrain. Nature Neuroscience 11: 1153-1161. 17 Make Life Visible SI-3 Eavesdropping on Biological Processes with Multi-Dimensional Molecular Imaging Scott E. FRASER Translational Imaging Center, Departments of Molecular and Computational Biology, and Biomedical Engineering, University of Southern California, Los Angeles, CA, USA A major challenge in fields ranging from cell and developmental biology to neurobiology is to draw upon the growing body of high-throughput molecular data to better understand the underlying cellular and molecular mechanisms. Molecular imaging offers powerful tools to obtain key data, but all imaging approaches are challenged by tradeoffs between speed, resolution, sensitivity, accuracy and photon budget. Many modern imaging techniques optimize only one or two of these factors, resulting in instrumentation that is incapable of obtaining accurate data without perturbing the systems under study. To advance the application of molecular imaging to studies within intact, living systems, we construct two-photon light-sheet microscopes, combining the deep penetration of two-photon microscopy and the speed of light sheet microscopy to generate images with more than ten-fold improvement in speed and sensitivity. The use of IR light to create the light sheet results in thin sheet that is dramatically less scattered by passing through tissue, permitting imaging with sufficient speed and resolution to generate unambiguous tracing of cells and signals in intact systems. In parallel with improved intravital imaging, we have refined our ability to quantitatively image multiple labels in the same specimen. In live specimens, we have optimized approaches for creating multiple “test points” in gene regulatory networks so we can collect data on the molecular dynamics that underlie the developmental dynamics of the embryo. These new approaches improve the accuracy and efficiency of gene trapping so that we can quickly generate “test points” in any gene product so that we can eavesdrop on the normal function of the system, using multispectral imaging, lifetime imaging and fluctuation analysis. Improvements in each analytical imaging approach have tailored these approaches to intravital imaging, where the signal-to-noise ratio is often limited. Studying many biological processes (e.g. neuronal activity or heart motions), require true 4D imaging, where events are followed over time across 3D space rather than via reconstructions from stacks of 2D images. Extended 3D capability has been demonstrated with Light Field Microscopy (LFM), where a lenslet array permits a single 2D image to capture an extended light field from the sample space. Computational reconstruction is used to convert the 2D light field images into true 3D images of the sample, trading some x,y spatial resolution for dramatically improved depth of field and depth resolution. We have optimized LFM by combining it with Light Sheet Illuminations strategies, creating Light Field – Selective Volume Illumination Microscopy (LF-SVIM). LF-SVIM illuminates only the volume of interest, significantly reducing the background, providing higher contrast and generating more accurate light field image reconstructions. Our implementation allows concurrent recording of the light-field-reconstructed volumetric image, together with a high-speed, high-resolution, single optical-section of the same sample. The combined generation of higher-resolution/lower-speed, and lower-resolution/higher-speed images facilitates the collection of anatomical and molecular data. 18 Make Life Visible SI-4 Apical microtubules define the function of epithelial cell sheets consisting of non-ciliated or multi-ciliated cells Sachiko TSUKITA Graduate School of Frontier Biosciences and Graduate School of Medicine, Osaka University, Japan. Epithelial cells adhere to each other by tight junctions (TJs) to form cell sheets, which is a critical step in epithelial barrier creation and tissue morphogenesis. We recently discovered that a network of microtubules exists just below the apical membrane of the epithelial cell sheet. This network is organized under the control of the TJs and regulates epithelial barrier function and morphogenesis concomitantly and in conjunction with the TJs; this system is defined as the “TJ-apical complex”. Multiciliated cells (MCCs) drive fluid transport through coordinated ciliary beating, the direction of which is established by the basal body (BB) orientation. Airway MCCs have hundreds of BBs, which are uniformly oriented and linearly aligned by an unknown mechanism. To examine the mechanism for BB alignment, we developed a long-term, high-resolution, live-imaging method, and observed GFP-centrin2-labeled BBs in mouse tracheal MCCs in vitro. The differentiating BB arrays adopted four stereotyped patterns, from a clustering “Floret” pattern to the linear “Alignment”. This alignment process was coordinated with the BB orientations, revealed by double-immunostaining for BBs and their asymmetrically associated basal feet (BFs). The BB alignment was perturbed by disrupting the apical microtubules with nocodazole or by a BF-depleting Odf2 mutation. Finally, we constructed a theoretical model based on our observations. Our findings indicate TJ-apical complex that the apical cytoskeleton provides self-organizing machinery for tracheal MCCs by acting as a viscoelastic material to linearly align the BBs. The general role of the “TJ-apical complex” in both non-ciliated and multi-ciliated epithelial cells will be discussed, with special reference to epithelial morphogenesis and function. Epithelial cells 19 Make Life Visible SI-5 Optogenetic assemblies of cortical force-generating complexes during mitosis Tomomi KIYOMITSU Graduate school of science, Nagoya University, Aichi, Japan Correct positioning of the mitotic spindle is critical for cell fate determination, tissue organization, and development, because spindle position and orientation determine the characteristics of daughter cells by regulating their size, position and the distribution of polarized factors (see Figures below). In animal cells, the position and orientation of the mitotic spindle are defined by cortical pulling forces exerted on astral microtubules emanating from spindle poles. Accumulating evidence indicates that a minus end-directed microtubule-based motor called cytoplasmic dynein (hereafter referred to as dynein) is recruited to specific cortical regions to generate cortical pulling forces in various cell types. However, the sufficiency of and detailed mechanisms underlying dynein-dependent force generation remain largely unknown because of the difficulties associated with in vitro reconstitution of cortical dynein complexes and formation of the mitotic spindle and cortical structure. Here, we developed a light-inducible system for targeting cortical dynein receptors or regulators to locally illuminated regions in the mitotic cell cortex to assemble dynein complexes in a spatially and temporally controlled manner. Our results indicate that NuMA–dynein complex is sufficient to move and orient the mitotic spindle in human cells. In addition, we will discuss how myosin-dependent cortical contractile forces regulate spindle position by altering cellular boundaries during mitosis. Figure: Diagram showing the regulation of daughter cell characteristics through spindle position and orientation (adapted from Kiyomitsu, Trends in Cell Biology 2015. Partially modified.) 20 Make Life Visible S1-6 In vivo Imaging Probes with Tunable Chemical Switches Kazuya KIKUCHI Graduate School of Engineering, Osaka University, Suita City, Osaka, Japan One of the great challenges in the post-genome era is to clarify the biological significance of intracellular molecules directly in living cells. If we can visualize a molecule in action, it is possible to acquire biological information, which is unavailable if we deal with cell homogenates. One possible approach is to design and synthesize chemical probes that can convert biological information to chemical output. In this talk, molecular design strategies for MR and fluorescence imaging probes are introduced. MRI (Magnetic Resonance Imaging) is an imaging technique using nuclear magnetic resonance phenomenon. MRI has been clinically used since it yields highly spatial resolution images of deep regions in living animal bodies. 19F MRI is suitable for monitoring particular signals concerning biological phenomena because 19F MRI shows little endogenous background signals. We have also developed the 19F MRI probes to detect protease activity and gene expression on the basis of paramagnetic resonance enhancement (PRE) effect. However, 19F MRI probes have faced two challenges. First, 19F MRI has the low sensitivity. Second, the suppression of molecular mobility induced by the increase in molecular size shortens the transverse relaxation time (T2), which is a crucial factor affecting the MRI contrast, resulting in attenuation of the MRI signals. To solve these challenges, we developed a novel 19F MRI contrast agent, fluorine accumulated silica nanoparticle for MRI contrast enhancement (FLAME), which is composed of a perfluorocarbon core and a robust silica shell. FLAME has advantages such as high sensitivity, stability, modification of the surface, and biocompatibility. The activatable derivative of FLAME will also be introduced Intravital imaging by two-photon excitation microscopy (TPEM) has been widely utilized to visualize cell functions. However, small molecular probes (SMPs) commonly used for cell imaging cannot be simply applied to intravital imaging because of the challenge of delivering them into target tissues, as well as their undesirable physicochemical properties for TPEM imaging. Here, we designed and developed a functional SMP with an active-targeting moiety, higher photostability, and fluorescence switch, and imaged target cell activity by injecting the SMP into living animals. The SMPs are based on BODIPY structure which is optimized for photostability and for fluorescence wavelenghth overlap for multicolor imaging. The combination of the rationally designed SMP with a fluorescent protein as a reporter of cell localization enabled quantitation of osteoclast activity and time-lapse imaging of its in vivo function associated with changes in cell deformation and membrane fluctuations. 21 Make Life Visible S1-7 Circuit-dependent striatal PKA and ERK signaling underlying action selection. Kazuo FUNABIKI Institute of Biomedical Research and Innovation, Foundation for Biomedical Research and Innovation, Kobe, Japan The striatum is a key neural substrate that controls selection of reward-seeking and aversive behaviors. Striatal transmission is mediated by 2 distinct neuronal subpopulations consisting of D1 and D2 receptor-expressing medium spiny neurons (dMSNs, iMSNs) and is regulated by protein kinase A (PKA) and extracellular signal-regulated kinase (ERK) cascades. However, the in vivo roles and signaling mechanisms of dMSNs and iMSNs remain largely elusive, because of the lack of techniques to monitor such kinase activities in behaving animals. In this presentaion, we report that the activities of PKA and ERK are coordinately and reciprocally regulated in dMSNs and iMSNs of the dorsal striatum by rewarding and aversive stimuli. This finding was made by developing a novel in vivo method in which Förster resonance energy transfer (FRET) biosensors of PKA and ERK(Kamioka et al, Cell Struct Funct, 2012) were specifically expressed in either dMSNs (D1-PKA and D1-ERK) or iMSNs (D2-PKA and D2-ERK), with fluorescence changes optically recorded by micro-endoscopy in freely moving mice (Goto et al, PNAS, 2015). Cocaine administration rapidly and continuously activated both D1-PKA and D1-ERK but markedly suppressed both D2-PKA and D2-ERK. Conversely, prominent activation of D2-PKA and D2-ERK together with strong suppression of D1-PKA and D1-ERK was evoked in response to electric foot shocks. To further explore the dynamic modulation of PKA and ERK in MSNs underlying naturally occurring action selection behavior, we similarly monitored PKA and ERK during mating behavior of male mice. Importantly, D1-PKA and D1-ERK were activated during the mating reaction of male mice; but D1-PKA was promptly inactivated upon ejaculation. In contrast, D2-PKA and D2-ERK were elevated when male mice became indifferent to mating. Manipulation of cAMP levels by DREADDs successfully induced PKA response of either dMSNs or iMSNs and concomitantly affected mating behaviors, indicating the causal relationship between PKA activity of dMSNs and iMSNs in the dorsal striatum and the mating reactions in male mice. Thus, a dynamic regulatory shift of PKA and ERK between dMSNs and iMSNs underlies the rapid selection of reward-directed and aversive behaviors. We also applied these techniques to measure the temporal dynamics of PKA response in the formation of aversive memory in the core part of nucleus accumbens(NAc). We found that PKA activities of iMSNs at NAc occurred not instantaneously after footshock but in a delayed and progressive manner(Yamaguchi et al, PNAS, 2015). We believe that the above methodologies allow us to study regulatory mechanisms of neural circuits involved in a wide range of animal behaviors. 22 Make Life Visible SI-8 Electrophysiology, Unplugged: New Chemical Tools to Image Voltage Evan W. MILLER Departments of Chemistry, Molecular & Cell Biology and Helen Wills Neuroscience Institute, University of California, Berkeley, United States of America Fast changes in membrane potential drive the unique physiology of excitable cells like neurons. Efforts to monitor dynamic changes in membrane potential in living systems typically rely on either highly invasive electrode and patch clamp techniques or on indirect imaging methods like calcium imaging. Direct imaging of voltage changes with voltage-sensitive indicators remains an outstanding challenge in neurobiology. I will present our efforts to design, synthesize, and apply new voltage-sensitive fluorescent dyes that use photoinduced electron transfer (PeT) as a voltage-sensing trigger to achieve fast and sensitive voltage imaging in a variety of biological contexts. 23 Make Life Visible SI-9 Single Cell Analysis of Stimulated Immune Cells with Real-time Selection Sotaro UEMURA Dept of Biol Sci, Grad Sch of Sci, The University of Tokyo, Tokyo, Japan Life is maintained at the organism level and at the organ level by the skillful orchestration of cell to cell communication and regulation. Cells can communicate with surrounding cells and even over long distances mainly through soluble factors such as hormones or cytokines. Traditionally, these secreted factors have been purified from the supernatant of a large number of cultured cells and analyzed by gel electrophoresis or immunoassays. However, it is impossible to understand when and how secretion occurs on a single cell level with this conventional approach. Y. Shirasaki et al have developed new technology to monitor single cell secretion dynamics on nanofabricated chips by merging conventional sandwich immunoassays with TIRF (Total Internal Reflection Fluorescence). Together with this advancement in real-time single cell secretion imaging, we are also upgrading pre-existing techniques such as live cell imaging and microfluidics to elucidate secretion mechanisms. Here, we developed a new tool for a parallel measurement platform for real-time single-cell secretion imaging with a multi-reservoir integrated nano litter-well array chip. It enables us to compare secretion responses on the multi specimens with various stimuli simultaneously. We could successfully monitor IL-13 periodical secretion dynamics from IL-33 stimulated mouse Type 2 innate lymphoid (ILC2). Furthermore, we applied this system to human Type 2 innate lymphoid (ILC2) cells, rarely obtained from 20 mL of peripheral blood (~1x103 cells) of human donor, and succeeded in monitoring secretion response. Interestingly, both results showed strong cell-cell heterogeneity in secretion dynamics. To clarify what factors generate heterogeneity, next we combined our real-time secretion imaging system and selectively picking technique for further analysis of picked specific cell. We also focus on the picking timing due to mRNA instability in time. In contrast to cells picked regardless of secretion timing, cells picked 30 min after the observation of the beginning of secretion showed homogeneous cytokine mRNA levels. Our results suggest that the cell-to-cell heterogeneity mainly originates from the fluctuation in responding time. 24 Single cell secretion measurements Y. Shirasaki et al, Scientific Reports 4: 4736 (2014) Make Life Visible S1-10 Comprehensive approaches sing luminescence to studies of cellular functions Atsushi MIYAWAKI Brain Science Institute, and Center for Advanced Photonics, RIKEN, Japan In a signal transduction diagram, arrows are generally used to link molecules to show enzymatic reactions and intermolecular interactions. To obtain an exhaustive understanding of a signal transduction system, however, the diagram must contain three axes in space and the time base, because all events are regulated ingeniously in space and time. The scale over time and space is ignored in biochemical approaches in which electrophoresis is applied to a specimen prepared by grinding millions of cells. A farseeing article entitled “Fluorescence Imaging Creates a Window on the Cell” was written by Dr. Roger Y. Tsien in 1994 (appearing in Chemical & Engineering News). Dr. Tsien, the pioneer of fluorescence imaging methods and recipient of the 2008 Nobel Prize in Chemistry, passed away on August 24, 2016. He was 64. Dr. Tsien advocated employing the so-called real-time and single-cell imaging technique to fully appreciate cell-to-cell heterogeneity. He also had steadfastly pursued the creation of a reliable gate that would enable researchers to better understand the “feelings” of individual cells. Over the past two decades, various genetically encoded probes have been generated principally using fluorescent proteins. I will discuss how the probes have advanced our understanding of the spatio-temporal regulation of biological functions, such as cell-cycle progression, autophagy, and metabolism, inside cells, neurons, embryos, and brains. I will also speculate on how these approaches will continue to improve due to the various features of luminescent (fluorescent) proteins that serve as the interface between light and life. Newly emerging genetically encoded tools will surely stimulate the imagination of many life scientists, and this is expected to spark an upsurge in the demand for them. As a result, light microscopes will inevitably be equipped with special hardware and software functions to optimize their use. In this regard, a significant evolution in microscopy will be necessary if luminescent (fluorescent) protein technologies are to enjoy widespread use. 25 Make Life Visible Session II DAY 1 DAY 2 KEYNOTE 9:30-10:20 A. Vania Apkarian S2-4 9:00-9:40 S2-1 10:20-10:50 Masaya NAKAMURA S2-5 9:40-10:10 Masashi Yanagisawa S2-2 11:05-11:35 Nicholas SMITH S2-6 10:25-11:05 Mark J. Schnitzer, S2-3 11:35-12:05 James T PEARSON & Mikiyasu SHRAI S2-7 10:05- 10:25 Motomasa TANAKA Ulrich H. VON ANDRIAN S2-8 11:35-12:05 Shigeo OKABE 26 DAY 3 Make Life Visible S2-1 Cortical plasticity after spinal cord injury using resting-state functional magnetic resonance imaging Masaya NAKAMURA Department of Orthopaedic Surgery, Keio University School of Medicine, Tokyo, Japan It is recently reported that after spinal cord injury (SCI), neuronal connectivity changes occur not only in the spinal cord, but also in the brain. However, there has been no report investigating the changes of the neuronal functional communication among the several regions of the cerebral cortex after SCI .Therefore, we examined the changes in neuronal functional connectivity of the brain after SCI by taking the resting state-functional MRI (rs-fMRI). C57BL6 female mice were subjected to rs-fMRI without anesthesia. After careful acclimation to environmental stress in taking MRI, rs-fMRI was performed and the data of different mice brain were standardized with the stereotaxic MRI brain template. By classifying the regions of the brain based on the Allen mouse brain atlas, the neuronal functional connectivity was analyzed among the specific regions. Next, complete transection or contusion SCI was induced at Th10 level in these mice. rs-fMRI was taken at 1,3,7 and 14 weeks after injury and the changes in neuronal functional connectivity of their brain were visualized using SPM12 software and CONN toolbox, and these data were analyzed using graph theory. First, we succeeded in detecting the normal neuronal functional connectivity in the brain of the awake mice through rs-fMRI. In the comparative analyses before and after the transection SCI, the changes in functional connectivity was observed between the primary and secondary motor cortex. Moreover, by analyzing the brain connectivity after the contusion SCI, we detected the changes in the neuronal functional connectivity in accordance with the motor function recovery. Based on the graph theory, quantitative analyses of the whole brain structural community revealed the decrease in the density of the whole brain network and the changes in the pattern of the components of the specific brain community structure after SCI. In the current study, we demonstrated the feasibility to examine neuronal functional connectivity in the brain of awake mice using rs-fMRI. We also showed the relative changes in the neuronal functional connectivity in the brain after SCI and identified the regions that were strongly related to the functional recovery after SCI. In addition, we detected that the networks changed in the brain after SCI by the analyses using a graph theory. These networks were divided into small communities and the density of these networks decreases immediately after SCI, then individual regions change the importance of their network, suggesting that brain networks were reorganized and changed the efficiency of the entire network after SCI. 27 Make Life Visible S2-2 Multimodal Label-free imaging to assess compositional and morphological changes in cells during immune activation Nicholas SMITH Immunology Frontier Research Center, Osaka University, Osaka, Japan When activated by pathogen-based or other triggers, immune cells respond in a wide variety of ways, which involve multiple pathways. The discovery of, and ability to inhibit, or enhance these pathways is a cornerstone of immunological research. The majority of research carried out in immunology uses either wetlab based techniques such as electrophoresis or ELISA, in combination with cell counting based on surface markers and targeted imaging by fluorescent labelling or gene modification. In our research we use only light, in a label-free microscopy approach to study how immune cells change following activation, as well as use these imaging modes to collect fundamental information about the compositional differences between cells. This has a number of unique features over the more common techniques. By spectroscopic measurement of Raman scattered photons, and by mapping the spatial distribution of Raman spectra, we can generate an image of the molecular distributions in living cells. This information is then used to evaluate the cellular composition and how it changes over time [1]. Without labelling, the light-sample interaction is not affected by staining protocols, but more importantly, the technique carries few assumptions about the preexisting conditions in the cells, and is able to measure fundamental changes in the cells which can occur by pathways not yet isolated. However, with Raman imaging, the data is generated from low numbers of photons, so that while it contains a large amount of information in terms of content, it is relatively slow, taking minutes to produce a cell image. We therefore take the approach of adding an additional imaging mode to provide more information and higher throughput. From a number of different possible modes [2], we have implemented quantitative phase imaging by digital holographic microscopy [3] which can give up to video rate imaging during the collection of the Raman data, and provides a quantitative map of phase shifts in the cell. By comparing the two sets of data, we determined that the phase shifts in the cell are linked to the protein distribution [4]. Using both of these modes simultaneously, we achieve one phase data set that is accurate in spatial location and well-resolved in time but poor in terms of chemical information, while the complementary Raman mode provides the ability to isolate and distinguish chemical components in sub-cellular locations, but with a lower degree of spatio-temporal resolution. Fig. 1. Simultaneous quantitative phase by digital holographic microscopy (a,b) and Raman scattering image (c) of a live mouse embryonic fibroblast (MEF) cell. From Pavillon et al [3]. 28 Make Life Visible These two imaging modes working together allow us to recognize components in the cell that move during the Raman acquisition, but also provide more quantitative data from the cell that allows us to discriminate cell types and cell states, with relatively high-throughput [5]. One significant advantage of using label-free methods to study cellular reactions is that they do not require knowledge of the cellular reaction which is expected to take place. In Raman imaging, the endogenous molecular distribution is measured, so that unexpected or new reactions in the cell may be measured. We are applying these techniques to measure activation of macrophages, discrimination of major types and subtypes of immune cells, as well as looking at the spatial redistribution of cellular components. As an example, when malaria infects a host, the parasite produces a by-product, known as hemozoin, which has a spectral signature that can be recognized and separated from other cellular heme-based compounds. This can be used as a diagnostic of disease [6], and with imaging, we can also visualize not only how the hemozoin is uptaken, but how the cell rearranges its endogenous components following uptake, as shown in Fig. 2., which illustrates the significant changes in composition after uptake [7]. Fig. 2. Label-free imaging of the macrophage uptake of malarial hemozoin. Color channels are linked to spectral (i.e. molecular compositional) components in label-free Raman imaging. From Hobro et al, [7]. [1] M. Okada, N. Smith, A. Palonpon, H. Endo, S. Kawata, M. Sodeoka, K. Fujita, P.N.A.S., 109, 28-32, 2012. [2] N. Pavillon, K. Fujita and N. I. Smith, J. Innov. Opt. Health Sci. 7(5), pp. 1330009-1-22, 2014. [3] N Pavillon, NI Smith, EPJ Techniques and Instrumentation 2 (1), 1-11, 2015 [4] N. Pavillon, A. Hobro, and N. Smith, Biophys. J., 105, 1123-1132, 2013. [5] N. Pavillon and N. I. Smith, J. Biomed. Opt. 20(1), pp. 016007-1-016007-10, 2015. [6] A. J. Hobro, A. Konishi, C. Coban and N. I. Smith, Analyst 138(14), pp. 3927-3933, 2013 [7] A. J. Hobro, N. Pavillon, K. Fujita, M. Ozkan, C. Coban and N. I. Smith, Analyst 140, pp. 2350-2359, 2015. 29 Make Life Visible S2-3 Investigating in vivo myocardial and coronary molecular pathophysiology in mice with X-ray radiation imaging approaches James T PEARSON and Mikiyasu SHRAI National Cerebral & Cardiovascular Center, Osaka, Japan We have developed X-ray imaging approaches utilizing synchrotron radiation to investigate the pathophysiological mechanisms of myocardial and vascular dysfunction in small animal models. These approaches are being utilized to determine the neurohormonal factors that contribute most to the early origins of diabetic heart disease and various forms of heart failure in rats, and more recently in mice. Our X-ray diffraction imaging approach has revealed that contractile dysfunction and impaired muscle relaxation is non-uniform in distribution in diabetic rats, being most profound in the deeper cardiac muscle layers. Further, at the level of the sarcomere, upregulation of protein kinase C and rho-kinase evoke dysregulation of cross-bridge cycling by impairing myosin head extension to the actin binding sites of muscle filaments. Whether such impairment in contractility and relaxation also occurs at the level of the sarcomeric proteins in other cardiac disease states, such as hypertrophic cardiomyopathy, is currently being investigated utilizing a knockin mouse model bearing a human gene mutation in cardiac TnT. At the same time, we are also utilizing fast synchrotron microangiography to reveal the early origins of endothelial and smooth muscle dysfunction in diabetes, hypertension and heart failure. Our recent data show how protein kinase C/rho kinase and other factors such as angiotensin-II and sympathetic overactivation, are the principal factors in impaired regulation of coronary flow in the microvessels and a decline in perfusion of cardiac muscle with type 2 diabetes disease progression in rats. In other studies with obese diabetic (db/db) mice, we can now show directly with high temporal and spatial resolution imaging how high-intensity exercise but not moderate-intensity exercise improves endothelial dysfunction in the coronary microcirculation. Furthermore, utilizing immunohistochemistry and quantitative PCR we can show that improved coronary perfusion and cardiac function in obese mice occurred through the mitigation of inflammation and oxidative stress through upregulation of beneficial microRNA molecules. Finally, we are continuing to progress the development of an improved microfocus X-ray system for real time vascular imaging of anaesthetised rodents without the need for synchrotron radiation. Recent tests with high-speed digital cameras coupled to an image intensifier suggest that small arterioles can be resolved in cerebral, renal and peripheral the resolving ability of the visible third and fourth branches of the coronary arterial vessels in vivo. 30 Make Life Visible S2-4 Visualizing the Immune Response to Infections Ulrich H. VON ANDRIAN Dept. of Microbiology & Immunobiology, Harvard Medical School, Boston, MA, USA The immune system is tasked with detecting and responding to infections anywhere in the body. To accomplish this task requires the coordinated migration of immune cells and highly dynamic interactions of the migrating cells with their environment. Lymph nodes play a central role in this process by acting as local filter stations that prevent the spread of invading microbes and by providing a sophisticated environment to initiate and regulate innate and adaptive immune responses to antigens derived from pathogens, malignant cells and vaccines. To this end, lymph nodes harbor specialized antigen presenting cells and constantly recruit diverse lymphocyte subsets that engage in continuous immune surveillance and mount protective effector and memory responses. Our laboratory has developed intravital microscopy techniques that allow us to identify and track the intra- and extravascular trafficking and dynamic interactions of different immune cell subsets within lymph nodes of anesthetized mice. Using fluorescence imaging strategies, we have traced the dissemination of invading bacteria and viruses via the lymph and analyzed how lymph-borne pathogens are handled upon entering a lymph node. We have characterized how pathogen-derived antigens are presented to T and B lymphocytes and how the in vivo kinetics of antigen recognition impact anti-microbial immunity and the formation and quality of immunological memory. This lecture will provide an overview of our lymph node imaging strategies and summarize key insights that have been gained from their use to dissect the mechanisms and consequences of the multi-facetted immune responses to infections. 31 Make Life Visible S2-5 Imaging Sleep and Wakefulness Masashi Yanagisawa, Takeshi Kanda, and Takehiro Miyazaki International Institute for Integrative Sleep Medicine (WPI-IIIS), University of Tsukuba, JAPAN We spend a third of our life in sleep, yet function and regulation of sleep remains mysterious. Sleep is not a simple rest state. The brain performs a variety of processing even during sleep, which is essential for brain functions such as synaptic scaling and consolidation of memory. Electroencephalogram (EEG), which arises from the cerebral cortex, can objectively classify the internal states of the brain into three stages: wakefulness, non-rapid eye movement (NREM) sleep, and REM sleep. Each of the three stages shows distinctive EEG activity. Toward understanding physiological nature of sleep at the cellular level, it has been of primary interest over the decades to clarify precisely the spatio-temporal organization of the cortical activity modulated by sleep/wake states. However, it is impossible to estimate spatio-temporal activity patterns of individual cells from EEG data. To explore individual neuronal activity in the cerebral cortex that underlies sleep and wakefulness, we used two-photon microscopy combined with transgenic labelling. We performed Ca2+ imaging in layer 2/3 (L2/3) of primary motor cortex (M1) of spontaneously sleeping and awake mice. To identify neuron types in vivo, Vgat-IRES-Cre mice, in which Cre recombinase is exclusively expressed in GABAergic neurons, were crossed with tdTomato reporter mice. Histological analysis of these Vgat-tdTomato mice demonstrated that 99% of GABAergic neurons (inhibitory neurons) in L2/3 of the M1 were labeled with tdTomato. Adeno-associated virus (AAV) vectors encoding a Ca2+ indicator GCaMP were injected into the M1 to visualize spontaneous neuronal activity in vivo. The AAV-injected mice were instrumented for imaging, and acclimated to experimental conditions for several days. After the acclimation, the mice were able to sleep spontaneously and wake up on a trackball under head-fixed conditions, as assessed by EEG and electromyogram (EMG). The experiment was conducted intermittently for 5 hours. Two-photon imaging and electrophysiological recording were performed simultaneously using a custom-built microscope set-up. Regions of interest (ROIs) were drawn manually on all cell bodies in the two-photon images for analysis of the intensity of fluorescence and the distance between cells. After subtracting the background signal, the fluorescence signal was converted to the rate of change from the baseline (ΔF/F) to detect Ca2+ transients, and to the Z score to analyze the correlation between the cells. Ca2+ transients were confirmed manually after automatically detecting continuous increases in the fluorescence signal. Data from ROIs lacking Ca2+ transients were excluded from the correlation analysis. Spontaneous Ca2+ dynamics in both excitatory and inhibitory neurons was highly active during wakefulness and REM sleep in comparison with NREM sleep. Correlation of Ca2+ activity between neurons was increased not only in NREM sleep but also in REM sleep. The activity correlation was higher among excitatory neurons spatially close to each other during wakefulness and REM sleep, and decreased with distance between neurons. The distance-dependent activity correlation was not prominent in NREM sleep. These results suggest that NREM and REM sleep exert distinct influence on cortical neurons. Contrary to classical prediction from EEG, synchronized neuronal activity occurs not only during NREM sleep but also during REM sleep in the microscopic scale. 32 Make Life Visible S2-6 Optical imaging of large-scale neural codes and voltage dynamics in behaving animals Mark J. Schnitzer, Stanford University and Howard Hughes Medical Institute. A longstanding challenge in neuroscience is to understand how the dynamics of large populations of individual neurons contribute to animal behavior and brain disease. Addressing this challenge has been difficult partly due to lack of appropriate brain imaging technology for visualizing cellular dynamics in awake behaving animals. I will discuss several new optical technologies of this kind. The miniature integrated fluorescence microscope allows one to monitor the dynamics of up to ~1000 individual genetically identified neurons in behaving mice over weeks, allowing time-lapse studies of the neural codes underlying episodic, emotional and reward related memories. Toward elucidating the interactions between brain areas during active behavior, multi-axis optical imaging can record the dynamics of two or more neural ensembles residing in different brain regions. Lastly, genetically encoded voltage indicators are progressing rapidly in their capacities to allow high fidelity detection of neural spikes, accurate estimation of spike timing, and studies of oscillatory voltage dynamics in targeted cell types of awake behaving animals. 33 Make Life Visible S2-7 Abnormal local translation in dendrites impairs cognitive functions in neuropsychiatric disorders Motomasa TANAKA, Ryo ENDO, Noriko TAKASHIMA, Yoko NEKOOKI-MACHIDA, Akira SAWA RIKEN Brain Science Institute, Wako, Japan Department of Psychiatry, Johns Hopkins University School of Medicine, USA Neurodegenerative disorders involving protein aggregation, including frontotemporal lobar degeneration (FTLD) and Huntington’ disease1), are often co-morbid with mental disorders of unknown origin. Although toxic protein aggregates may entrap proteins involved in psychiatric disorders and elicit deleterious effects, candidate mechanisms for this co-aggregation hypothesis remain unknown. Here, we report a novel cytosolic protein aggregate in FTLD brain composed of DISC1, a biological mediator for mental illness and TDP-43, the major protein constituent of insoluble inclusions in FTLD. FTLD model mice with DISC1/TDP-43 co-aggregation in frontal cortex showed hyperactivity and social interaction deficits that were rescued by exogenous DISC1 expression. At the cellular level, fluorescent imaging analyses revealed that the amounts of neuronal RNA granules and new protein synthesis in dendrites are reduced in the neurons that lack DISC1 expression or contain DISC1/TDP-43 co-aggregates. Furthermore, cell biology experiments showed that the deficiency of DISC1 destabilized neuronal RNA granules and impaired local dendritic translation via impairment in translation initiation and peptide chain elongation. These results support our novel hypothesis that co-aggregation of risk factors for psychiatric diseases may compromise local translation in dendrites and mental conditions. [Reference] 1) Tanaka M., Ishizuka K., Nekooki-Machida Y., Endo R., Takashima N., Sasaki H., Komi Y., Gathercole A., Huston E., Ishii K., Hui K.K., Kurosawa M., Kim S.H., Nukina N., Takimoto E., Houslay M.D., and Sawa A. J. Clin. Invest., in press (2017). 34 Make Life Visible S2-8 Imaging synapse formation and remodeling in vitro and in vivo Shigeo OKABE Department of Cellular Neurobiology, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan Formation of synapses and their subsequent remodeling and elimination are highly regulated and provide a basis for proper functions of the mature brain circuits. Redundant connections between neurons may exist in the early phase of neural circuit development, but extrinsic instructive signals regulate their remodeling and elimination. In vivo two-photon excitation microscopy is a useful technique for the quantitative analyses of synapse dynamics in the mouse neocortex. Our imaging analyses confirmed the presence of two phases of synapse dynamics in the layer II/III pyramidal neurons during postnatal development. In the first phase (until postnatal 20 days), dynamics of synapses is high and more than 10% of spine synapses are replaced within 24-48 hours. Dynamics of spine synapses is suppressed remarkably at postnatal 8 weeks. Consistently, the cortical synaptic density increases rapidly in the first phase, while it shows the trend of gradual decrease in the second phase. This transition should be important in both normal development and pathology of cortical neural circuits. Pathophysiology of juvenile-onset psychiatric disorders may be related to impairments in the postnatal circuit remodeling. To test this possibility, we analyzed in vivo dynamics of spine synapses in mouse models of autism spectrum disorders (ASDs). We found up-regulation of spine turnover in multiple ASD mouse models with different genetic backgrounds. Neurons in the somatosensory cortex of ASD mouse models responded poorly to whisker stimulation, suggesting dysfunction of cortical information processing. Selective impairment in spine dynamics and less precise wiring in the neocortical circuits may be a core pathology of juvenile-onset psychiatric disorders. To further characterize dynamics of spine synapses both in the postnatal developmental stage and in pathological conditions, novel strategies for measurement and classification of spines at different functional states should be developed. High resolution imaging technologies, including localization microscopy (PALM and STORM) and structured illumination microscopy (SIM), are promising approaches. These optical techniques allow us to visualize and quantitatively analyze molecular distribution and surface geometry of postsynaptic spines. We applied high resolution techniques to the analyses of spine structure in hippocampal neurons. Quantitative analyses revealed basic structural properties of spines and their alterations in disease-related conditions. High resolution imaging of synapses, together with in vivo analyses of their dynamics, will provide new information in physiology and pathology of neural circuit construction. 35 Make Life Visible Session III DAY 1 DAY 2 DAY 3 S3-1 13:05 -13:45 Denis LE BIHAN S3-6 15:40 - 16:20 Geoffrey D. Rubin KEYNOTE 9:00 - 9:50 Sanjiv Sam Gambhir S3-2 13:45 -14:15 Masaru ISHII S3-7 16:20 - 16:50 Kenji KABASHIMA S3-9 9:50 -10:20 Yasuhisa FUJIBAYASHI S3-3 14:15 -14:55 Hisataka KOBAYASHI S3-8 16:50 - 17:20 Masahiro JINZAKI S3-4 14:55 -15:25 Yasuyoshi WATANABE S3-5 15:25 -15:55 Yasuteru URANO 36 Make Life Visible S3-1 How MRI makes the Brain Visible Denis LE BIHAN NeuroSpin, CEA-Saclay Center, Gif-sur-Yvette, France Human Brain Research Center, Kyoto University, Kyoto, Japan The understanding of the human brain is one of the main scientific challenges of the 21st century: Academic or even philosophical stake: unravel the biological mechanisms of our mental life; Technological stake: replicate cerebral functional features in artificial systems; Medical stake: understand neurological or psychiatric diseases to allow early diagnosis and treatment of patients, especially for aging populations; Societal stake: improve our cognitive performances, education, neuronal bases of cultural and social behaviors, etc. All of those stakes have obviously economical counterparts. In this quest of the human brain neuroimaging has become an inescapable pathway, because it allows getting maps of brain structure and function in situ, non-invasively, in patients or normal volunteers of any age. Magnetic Resonance Imaging (MRI), especially, has become the reference approach over the last three decades to investigate the human brain functional anatomy in vivo because it provides high resolution images without ionizing radiations or the need for tracer injections. MRI allows brain anatomy of individuals to be visualized in 3 dimensions with great details, as well as networks of brain regions activated by high order cognitive functions up to consciousness, together with stunning images of the connections between those areas. Still, images remain at a macroscopic scale (millions of brain cells), while invasive techniques in animals and tissues explore very small ensembles of neurons. This large gap must be bridged to understand how the brain works, as interaction and synergy exist between all brain levels. Hence, an important challenge for neuroimaging is to push its current limits as far as possible to make the brain visible at spatial and temporal scales which may give access to a neural code. This quest for a "neural code" is one of the major challenges of contemporary science, together with the exploration of matter, of the universe. Obviously, this intimate knowledge of normal brain functioning mechanisms will lead to a better understanding of its dysfunctions, neurological or psychiatric pathologies in a broad sense, which will probably open new therapeutic possibilities (such as brain reprogramming). One may gain the necessary increase in the spatial and temporal resolution of the images obtained by MRI by boosting the magnetic field MRI. Ouststanding instruments operating to field of 11.7 teslas or above are emerging. Such instruments will not only allow us to "better" see inside our brain, confirming or invalidating our current assumptions on how it works, but also to generate new assumptions, today impossible to anticipate, to help us decode the functioning of our brain. 37 Make Life Visible S3-2 Intravital multiphoton imaging revealing cellular dynamics in vivo Masaru ISHII Department of Immunology and Cell Biology, Graduate School of Medicine and Frontier Biosciences, Osaka University, Osaka, Japan Intravital multiphoton imaging has especially contributed to the discovery in novel concepts in the field of immunology, where the cells comprising various immune tissues and organs are dynamic. For example, dynamic observations in lymph nodes have revealed the migratory changes in T cells in close contact with antigen-presenting dendritic cells. When T cells encounter antigen-bearing dendritic cells, they form stable contacts lasting for at least several hours for priming, and thereafter regain motility for recirculation. In thymus, the interactions were detected between thymocytes and stromal cells during positive and negative selections. Live imaging has been tested for visualization of immune cell inflammatory reactions in many other tissues, such as the skin, lung, liver, and small intestine, and has revealed critical biological phenomena in each system Bone is a mineralized hard tissue that limits the passage of visible or infrared lasers, and it has long been considered to be extremely difficult to observe intact bone tissues in living animals. We have developed a novel imaging system for visualization inside bone cavities with high spatiotemporal resolution. By utilizing this technique, we can demonstrate that osteoclasts, bone-destroying osteoclasts, and their precursors are dynamically migrating under the control of several chemokines and lipid mediators. By improving bone imaging techniques and originally developing a chemical fluorescent probe, we also successfully visualized bone-resorbing activity of mature osteoclasts lining bone surfaces and identified their real mode of action in situ. Despite its hardness, bone is a dynamic and elastic tissue that undergoes continuous remodeling by bone-resorbing osteoclasts and bone-replenishing osteoblasts. Inflammation and hormonal perturbation lead to the aberrant activation of osteoclasts, resulting in several bone-resorptive disorders, chiefly osteoporosis and rheumatoid arthritis. Thus, osteoclasts have emerged as a good therapeutic target against these diseases, and the intravital imaging of bone tissues would be a good tool for the identification of novel target molecules and the development and evaluation of novel therapeutics (Figure 1). Major progress has been made recently in imaging techniques, and several tools for the visualization of live biological systems in situ have become available. These tools must bring a paradigm shift in the field of various fields of biological sciences, and lead to changes in the treatment of Figure 1. In vivo dynamic behaviors of osteoclast, a several intractable diseases in the bone-’destroying’ macrophage – visualized by bone intravital future. microscopy 38 Make Life Visible S3-3 Theranostic Near Infrared Photoimmunotherapy for Cancer Hisataka KOBAYASHI Molecular Imaging Program, CCR/NCI/NIH, Bethesda, MD, USA Near infrared (NIR) photoimmunotherapy (PIT) is a newly developed, molecularly-targeted cancer photo-theranostic technology based on conjugating a near infrared silica-phthalocyanine dye, IRdye700DX (IR700) to a monoclonal antibody (MAb) thereby targeting specific cell-surface molecules. A first-in-human Phase 1 clinical trial of NIR-PIT with the cetuximab-IR700 (RM1929) targeting EGFR in patients with inoperable head and neck cancer was approved by the US FDA in April 2015, and is now in transition to a Phase 2 trial (https://clinicaltrials.gov/ct2/show/NCT02422979). When exposed to NIR light, the conjugate rapidly induces a highly-selective, necrotic/immunogenic cell death (ICD) only in antigen-positive MAb-IR700-bound cancer cells. ICD occurs as early as 1 minute after exposure to NIR light and results in irreversible morphologic changes on target-expressing cells including cellular swelling, bleb formation, and rupture of vesicles due to membrane damage. Meanwhile, immediately adjacent receptor-negative cells are totally unharmed. Dynamic 3D observation of tumor cells undergoing NIR-PIT along with novel live cell microscopies showed rapidly swelling in treated cells immediately after light exposure suggesting rapid water influx into cells. Cell biological analysis showed that ICD induced by NIR-PIT rapidly maturate immature dendritic cells adjacent to dying cancer cells initiating a host anti-cancer immune response. Additionally, NIR-PIT can enhance nano-drug delivery into the treated tumor bed up to 24-fold compared with untreated tumors, a phenomenon that we have termed the “super-enhanced permeability and retention” (SUPR) effect. Furthermore, NIT-PIT targeting immuno-suppressor cells, such as Treg, in a local tumor, can enhance tumor cell-selective systemic host-immunity leading to significant responses in distant metastatic tumors. In conclusion, due to it highly targeted cancer cell-selective cytotoxicity, NIR-PIT carries few side effects and healing is rapid. NIR-PIT induces ICD on cancer cells that initiates host immunity. Moreover, NIR-PIT can locally deplete Tregs and other immune suppressor cells infiltrating in tumor beds, thus, activating systemic anti-cancer cellular immunity without potential autoimmune adverse effects. Mechanism of NIR-PIT 39 Make Life Visible S3-4 Novel and integrated imaging on Chronic Fatigue Yasuyoshi WATANABE RIKEN Center for Life Science Technologies, Kobe, Japan Chronic fatigue is a serious sign of the state of ahead sick, so that it could be one of crucial targets of pre-emptive medicine. We therefore should develop good ways of evaluation of the degree of and prevention from chronic fatigue. In our questionnaire-based investigation, Japanese statistics of chronic fatigue in 2004 were ca. 39% of population. Such a high percentage of the people really suffer from prolonged fatigue sensation. The economic loss by chronic fatigue in Japanese society was calculated with the statistics, and the loss with the primary reason of chronic fatigue could be over 1,200 billion JPY (~ 10 billion USD) per year. So, we should clarify the mechanisms of chronic fatigue and invent better solution from it. So far, we have investigated the mechanisms of chronic fatigue and Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME). Fatigue is induced by over-work without enough rest. First, fatigue occurs with oxidization or oxygenation of the key components of biological functions such as functional proteins and membrane phospholipids. Then, if enough repair energy is lacking, the oxidized or oxygenated components could not be repaired by Ubiquitin systems nor replaced by de novo newly biosynthesized ones. Accumulation of such unrecoverable intracellular insults (dysfunction of proteins and other machineries) induces cellular dysfunction, which is detected by circulating immune cells. Immune cells which detect such cellular dysfunctions would produce and release cytokines to transmit the information where and to what extent the insults are. Along with this hypothesis, we should prevent the molecular and functional imaging methods for all cellular and molecular events in the vicinity of insults. Our major contribution with the methods developed is: 1) Elucidation of the brain regions and their neurotransmitter systems responsible for fatigue sensation and chronic fatigue; 2) Development of a variety of methods and scales to quantitatively evaluate the extent of fatigue; 3) Development of animal models of different causes of fatigue; 4) Elucidation of molecular/neural mechanisms of fatigue in humans and animals; 5) Invention of various methods or therapies on chronic fatigue and chronic fatigue syndrome; and 6) Development of the foods and methods to overcome fatigue. The outline of the concept and results of imaging will be presented in this symposium. [Ref. 1. Watanabe, Y. et al. eds., Fatigue Science for Human Health, Springer, 2008. 2. Watanabe Y, et al. Biochemical indices of fatigue for anti-fatigue strategies and products. In: Matthews G, et al. (eds), The Handbook of Operator Fatigue. Ashgate Publishing Limited, Great Britain, pp. 209-224, 2012.] 40 Make Life Visible S3-5 Novel fluorescent probes for rapid tumor imaging and fast glutathione dynamics Yasuteru URANO Graduate School of Pharmaceutical Sciences and Graduate School of Medicine, The University of Tokyo, Tokyo, Japan AMED-CREST, Japan Agency for Medical Research and Development, Tokyo, Japan Recently, we have succeeded to develop novel fluorogenic probes for various aminopeptidases based on our newly established rational design strategy with intramolecular spirocyclization. For example, gGlu-glutamyltranspeptidase (GGT), which is well-known to be upregulated in various cancer cells, was developed. In mouse models of disseminated human peritoneal ovarian cancer, activation of gGlu-HMRG occurred within 1 min of topically spraying onto tissue surfaces that are suspected of harboring tumors, with high tumor-to-background contrast1). Encouraged by the promising results described above, we next examined the feasibility of clinical application of gGlu-HMRG during surgical procedures. By topically spraying of gGlu-HMRG onto the surface of freshly excised human breast specimens, tumorous lesions exhibited a time-dependent increase of green fluorescence, and were clearly distinguished from surrounding mammary gland and fat. Breast tumors, even those smaller than 1 mm in size, could be easily discriminated from normal mammary gland tissues with 92% sensitivity and 94% specificity, within 5-15 min of probe application2). Also, we validated gGlu-HMRG could diagnose metastatic lymph nodes in breast cancer with sufficiently high sensitivity (97%), specificity (79%) and negative predictive value (99%) to render it useful for an intraoperative diagnosis of cancer3). These findings confirmed the availability of gGlu-HMRG to detect cancerous lesions in clinical samples, which should be a breakthrough in intraoperative margin assessment during breast-conserving surgery. Furthermore, in order to find out esophageal cancer-specific enzymatic activities, a library of fluorogenic probes for various aminopeptidases were applied one-by-one to tumor and normal biopsy samples from cancer patients. We could find out the enzymatic activity of dipeptidyl peptidase-4 (DPPIV) was upregulated in tumor-positive biopsy samples, but not in tumor-negative ones. Indeed, esophageal cancer in the human resected fresh specimens could be visualized by topically spraying DPPIV-activatable fluorogenic probes within 10 min4). These findings clearly demonstrated that our fluorogenic probes should be a breakthrough in rapid detection of tumors during endoscopic and surgical procedures. Another feature of the alterations in redox state is well known to be associated with a variety of diseases and cellular functions. Among them, glutathione (GSH) is the most abundant redox-related biomolecule, however, existing fluorogenic probes are unsuitable for live imaging of GSH dynamics due to irreversible mechanisms or slow reaction rates. Very recently, we have succeeded to develop first-in-class reversible and rapid fluorescent probes for intracellular GSH by utilizing the concept of intermolecular attack of nucleophiles to specific rhodamines. Our probes exhibited GSH concentration-dependent ratiometric fluorescence changes with reversible and rapid detection capabilities, and live-cell imaging and quantification of GSH with temporal resolution of seconds were successfully achieved5). <Reference> 1)Sci. Transl. Med., 2011, 3, 110ra119. 2)Sci. Rep., 2015, 5, 12080. 3)Sci. Rep., 2016, 6, 27525. 4)Sci. Rep., 2016, 6, 26399. 5)Nat. Chem., in press. 41 Make Life Visible S3-6 Coronary Heart Disease Diagnosis: Engineering Triumphs, Economic Barriers Geoffrey D. Rubin, MD, MBA, FACR Department of Radiology, Duke University, Durham, North Carolina, USA As the leading cause of death amongst men and women throughout the developed world, coronary heart disease is an important and expensive health problem. The fundamental pathophysiological mechanism of this disease is insufficient oxygen delivery to the heart muscle resulting in myocardial ischemia or infarction. Amongst a spectrum of secondary consequences of this tissue injury are dysrhythmia or cardiac failure leading to sudden death. Coronary artery disease (CAD) secondary to atherosclerosis of the coronary arteries is the principal limiter of oxygen delivery to the heart. When pre-existing atherosclerotic plaque ruptures and incites local thrombus formation, coronary arterial occlusion may ensue and cause fatal or debilitating myocardial injury. Restoration of myocardial oxygenation requires coronary revascularization through the placement of a coronary artery stent or bypass grafting. Chest pain is an extremely common cause for patients to seek medical attention, and determining who are the minority of chest pain patients with myocardial ischemia necessitating revascularization is a major activity of many medical centers. As the best predictor of future major cardiac morbidity and mortality from CAD, coronary angiography with fractional flow reserve measurement is the best determinant of which patients should undergo revascularization. Fractional flow reserve (FFR) is expressed as the blood pressure within a coronary artery divided by the blood pressure within the ascending aorta during maximum coronary flow. It requires the positioning of a pressure transducing wire within the coronary artery just distal to a suspect lesion and pressure measurements made following an intracoronary injection of adenosine. Both coronary angiography and FFR procedures are invasive and expensive. Moreover, approximately two thirds of patients with chest pain who undergo coronary angiography do not require revascularization, resulting in tremendous expense and waste. Prior to coronary angiography, most chest pain patients are triaged using noninvasive imaging. The most common noninvasive imaging tests are radionuclide scintigraphy, echocardiography, and computed tomography (CT). The former two “functional” exams are performed during resting and stressed states, while the “anatomical” exam of CT is almost always performed at rest. Recently, computational fluid dynamics have been applied to carefully segmented coronary artery lumina to model FFR measurements. These CT-derived FFR or CT-FFR measurements have been shown to effectively predict significant coronary artery lesions and when performed as a pre-requisite for coronary angiography can reduce the number of coronary angiograms performed by 60%, transforming the frequency of exams that reveal significant CAD from 27% to 69% and reducing the frequency of exams revealing no significant CAD by 84%. This remarkable advance is associated with a substantial reduction in per patient costs and improved quality of life scores when compared to other noninvasive imaging strategies. While evidence derived through scientific investigations position the clinical application of CT-FFR to substantially improve the efficiency of CAD diagnosis and triage to definitive 42 Make Life Visible management, economic barriers are emerging, particularly where the providers of healthcare are not aligned with the payers of healthcare. Within a fee-for-service payment system, reduced coronary angiography volumes result in revenue reductions to providers and cost savings to payers. Because the majority of provider costs associated with coronary angiography are fixed or overhead costs, this payment system places a financial burden on providers that disincentivizes the use of CT-FFR despite its benefits to patients and to the overall cost of care. While technology development can bring profound clinical improvements, economic considerations represent an important dimension when seeking high-value healthcare. 43 Make Life Visible S3-7 Live imaging of the skin immune responses Kenji KABASHIMA Department of Dermatology, Kyoto University Graduate School of Medicine, Kyoto, Japan. Varieties of immune cells orchestrate cutaneous immune responses. To capture such dynamic phenomena, intravital imaging is an important technique and it may provide substantial information that is not available using conventional histological analysis. Multiphoton microscopy enables the direct, three-dimensional, and minimally invasive imaging of biological samples with high spatio-temporal resolution, and it has now become the leading method for in-vivo imaging studies. Using fluorescent dyes and transgenic reporter animals, not only skin structures but also cell- and humor-mediated cutaneous immune responses have been visualized. In this symposium, I will introduce recent findings in cutaneous immune responses and skin structural changes in mice and humans. 44 Make Life Visible S3-8 Development of a horizontal CT and its application to musculoskeletal disease Masahiro JINZAKI Department of Radiology, Keio University School of Medicine, Tokyo, Japan Computed tomography is now the most widely used imaging tool in the clinical field for the diagnosis of various disease. However, CT imaging has been obtained only in a supine position since its development in 1972. The supine position has enabled us to keep the patient stable even when scanning time had previously taken much longer and can easily evaluate the organic disease such as cancer, infectious disease or atherosclerotic disease. Recently, a wider detector scan has become available and the rotation time is faster. As a result, the scanning time for the whole body is very short, approximately 20 seconds, which will enable us to keep the patient stable in the standing position. Furthermore, most of the functional diseases such as swallowing, respiratory function, urinary function, motor function and organ herniation are difficult to be evaluated in the supine position. Thus, we started the project to develop a CT scanner which enables us to obtain images in the standing position in 2012. It took one and a half year to persuade a company to collaborate this project and they agreed in 2014. We did many tasks by trials and errors to develop a device which keep the standing position safe and would prevent the patient from falling. This machine was accomplished in December, 2016. By the introduction of this machine, we will first collect a large amount of the normal cross-sectional data of the whole body in a sitting or standing position, which has not been evaluated so far. By determining the normal range of each anatomical structure, we hope to establish the criteria of many functional diseases. In the field of musculoskeletal disease, for example, the early detection of osteoarthritis in the knee, disk herniation of the supine or moving disorder of various joints will be possible, which has not been able to be detected in the conventional CT. In this presentation, we will show many representative images obtained by this CT scanner. 45 Make Life Visible S3-9 Imaging and therapy against hypoxic tumors with 64Cu-ATSM Yasuhisa FUJIBAYASHI National Institute of Radiological Sciences, National Institutes for Quantum and Radiological Science and Technology, Chiba, Japan Background Hypoxia in tumors contributes to the malignant characteristics such as therapy resistance and metastatic potential. We have developed a novel PET imaging agent, Cu-diacetyl-bis (N4-methylthiosemicarbazone) (Cu-ATSM), which can target tumor hypoxia with over-reduced conditions. Cu-ATSM can be labeled with several Cu radioisotopes, such as 60Cu, 62Cu and 64Cu. In recent years, clinical studies of Cu-ATSM PET has been conducted worldwide. In Japan, our group produced a generator system of 62Cu and multicenter clinical 62Cu-ATSM PET studies have been conducted using our system. These clinical studies have demonstrated that Cu-ATSM uptake is associated with therapeutic resistance, metastatic potential, and poor prognosis in several types of cancer. Among Cu-ATSM tracers, we demonstrated that 64Cu-ATSM can be used not only as a PET imaging agent but also as a therapeutic agent, because 64Cu emits β- particle and Auger electron, which can damage the cells. Therefore, 64Cu-ATSM is a promising theranostic agent to target malignant hypoxic tumors. This project In this project, we developed a method to detect and treat tumors that became malignant by acquiring decreased vascularity and hypoxia, during antiangiogenic treatment, with 64Cu-ATSM. 1. 64Cu-ATSM imaging First, we developed a method to simultaneously visualize vascularity and hypoxia within tumors with a dual-isotope SPECT/PET/CT system. In this method, we used both SPECT and PET tracers, 99mTc-labeled human serum albumin (99mTc-HSA) to detect vascularity and 64Cu-ATSM to detect hypoxia. An in vivo imaging study was conducted with HT-29 tumor-bearing mice. Both 64Cu-ATSM (37 MBq) and 99mTc-HSA (18.5 MBq) were intravenously injected into each mouse at 1 h and 10 min, respectively, before SPECT/PET/CT scanning. As a result, we demonstrated that this method can clearly visualize the distribution of each tracer and that 64Cu-ATSM high-uptake regions barely overlapped with 99mTc-HSA high-uptake regions within HT-29 tumors (Fig. 1). Second, we applied this method to tumors treated with an antiangiogenic agent bevacizumab. The bevacizumab-treated tumors showed reduced vascularity and increased proportion of hypoxic areas in tumors compared with untreated control (Fig. 2). 2. 64Cu-ATSM therapy Next, we examined efficacy of 64Cu-ATSM therapy with a mouse model with bevacizumab-treated HT-29 tumors. 64Cu-ATSM effectively inhibited tumor growth in bevacizumab-treated HT-29 tumors. 64Cu-ATSM therapy could be a novel approach as an add-on to antiangiogenic therapy. 46 Make Life Visible 47 Make Life Visible The Secretariat of the Symposium THE UEHARA MEMORIAL FOUNDATION Takada 3-26-3, Toshima-ku, Tokyo, 171-0033, JAPAN Phone: +81-3-3985-3500,8400, Fax: +81-3-3982-5613 E-mail:[email protected] http://www.ueharazaidan.or.jp 〒171-0033 東京都豊島区高田 3 丁目 26 番 3 号 TEL 03(3985)3500,8400 FAX 03(3982)5613 48
