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University of Yamanashi

9 ARTICLES PUBLISHED IN JoVE

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Biology

Measuring Near Plasma Membrane and Global Intracellular Calcium Dynamics in Astrocytes
Eiji Shigetomi 1, Baljit S. Khakh 1
1Departments of Physiology and Neurobiology, David Geffen School of Medicine, University of California, Los Angeles

We describe how to measure near membrane and global intracellular calcium dynamics in cultured astrocytes using total internal reflection and epifluorescence microscopy.

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Medicine

Production of Apolipoprotein C-III Knockout Rabbits using Zinc Finger Nucleases
Dongshan Yang 1, Jifeng Zhang 1, Jie Xu 1, Tianqing Zhu 1, Yanbo Fan 1, Jianglin Fan 2, Y. Eugene Chen 1
1Center for Advanced Models for Translational Sciences and Therapeutics, Department of Internal Medicine, University of Michigan Medical Center, 2Department of Molecular Pathology, University of Yamanashi

Recent development in gene targeting tools makes production of knockout (KO) rabbits possible. In the present work, we generated five Apolipoprotein (Apo) C-III KO rabbits using Zinc Finger Nucleases (ZFN). This work demonstrated that ZFN is a highly efficient method to produce KO rabbits.

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Neuroscience

Observation of the Ciliary Movement of Choroid Plexus Epithelial Cells Ex Vivo
Takafumi Inoue 1, Keishi Narita 2, Yuta Nonami 1, Hideki Nakamura 1, Sen Takeda 2
1Department of Life Science and Medical Bioscience, Faculty of Science and Engineering, Waseda University, 2Department of Anatomy and Cell Biology, Interdisciplinary Graduate School of Medicine & Engineering, University of Yamanashi

In this study, a detailed light microscopic technique was optimized for real-time observation and analysis of the motion of CPEC cilia ex vivo together with an electron microscopic method for ultrastructural analysis.

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Developmental Biology

Zygotic Fluorescence Recovery After Photo-bleaching Analysis for Chromatin Looseness That Allows Full-term Development
Masatoshi Ooga 1,3, Satoshi Funaya 2, Fugaku Aoki 2, Teruhiko Wakayama 1,3
1Faculty of Life and Environmental Sciences, Department of Biotechnology, University of Yamanashi, 2Department of Integrated Bioscience, Graduate School of Frontier Sciences, University of Tokyo, 3Advanced Biotechnology Center, University of Yamanashi

Chromatin looseness appears to be involved in the developmental potential of blastomeres. However, it is not known whether chromatin looseness can be used as a reliable index for the developmental potential for embryos. Here, an experimental system in which chromatin looseness-evaluated zygotes can develop to full term has been described.

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Chemistry

Preparation of Polyoxometalate-based Photo-responsive Membranes for the Photo-activation of Manganese Oxide Catalysts
Akira Yamaguchi 1,5, Toshihiro Takashima 2, Kazuhito Hashimoto 1,6, Ryuhei Nakamura 3,4
1Department of Applied Chemistry, The University of Tokyo, 2Clean Energy Research Center, University of Yamanashi, 3Biofunctional Catalyst Research Team, RIKEN Center for Sustainable Resource Science, 4Earth-Life Science Institute (ELSI), Tokyo Institute of Technology, 5Department of Materials Science and Engineering, Tokyo Institute of Technology, 6National Institute for Materials Science

Here, we present a protocol to prepare charge transfer chromophores based on a polyoxometalate/polymer composite membrane.

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Biochemistry

Sample Preparation for Probe Electrospray Ionization Mass Spectrometry
Sen Takeda 1, Kentaro Yoshimura 1, Hiroshi Tanihata 2
1Department of Anatomy and Cell Biology, University of Yamanashi School of Medicine, 2Shimadzu Corporation

This article introduces sample preparation methods for a unique real-time analytical method based on the ambient mass spectrometry. This method lets us perform real-time analysis of the biological molecules in vivo without any special pretreatments.

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Biochemistry

Isolation and Analysis of Plasma Lipoproteins by Ultracentrifugation
Manabu Niimi 1, Haizhao Yan 1, Yajie Chen 1, Yao Wang 2, Jianglin Fan 1,2
1Department of Molecular Pathology, Interdisciplinary Graduate School of Medicine, University of Yamanashi, 2School of Biotechnology and Health Sciences, Wuyi University

Several methods have been used for analyzing plasma lipoproteins; however, ultracentrifugation is still one of the most popular and reliable methods. Here, we describe a method regarding how to isolate lipoproteins from plasma using sequential density ultracentrifugation and how to analyze the apolipoproteins for both diagnostic and research purposes.

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Neuroscience

Transplantation of Human Induced Pluripotent Stem Cell-Derived Microglia in Immunocompetent Mice Brain via Non-Invasive Transnasal Route
Bijay Parajuli 1,2, Youichi Shinozaki 1,2, Eiji Shigetomi 1,2, Schuichi Koizumi 1,2
1Department of Neuropharmacology, Interdisciplinary Graduate School of Medicine, University of Yamanashi, 2GLIA Center, University of Yamanashi

The protocol presented here allows the transplantation of induced pluripotent stem cell-derived human microglia (iPSMG) into the brain via a transnasal route in immunocompetent mice. The method for the preparation and transnasal transplantation of cells and the administration of cytokine mixture for the maintenance of iPSMG is shown.

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Neuroscience

In Vivo Wide-Field and Two-Photon Calcium Imaging from a Mouse Using a Large Cranial Window
Satoshi Manita 1, Eiji Shigetomi 2,3, Haruhiko Bito 4, Schuichi Koizumi 2,3, Kazuo Kitamura 1
1Department of Neurophysiology, Faculty of Medicine, University of Yamanashi, 2Department of Neuropharmacology, Faculty of Medicine, University of Yamanashi, 3Yamanashi GLIA center, Interdisciplinary Graduate School of Medicine, University of Yamanashi, 4Department of Neurochemistry, Graduate School of Medicine, The University of Tokyo

The present protocol describes making a large (6 x 3 mm2) cranial window using food wrap, transparent silicone, and cover glass. This cranial window allows in vivo wide-field and two-photon calcium imaging experiments in the same mouse.

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