University of Yamanashi

9 ARTICLES PUBLISHED IN JoVE

Biology

Measuring Near Plasma Membrane and Global Intracellular Calcium Dynamics in Astrocytes

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.

Medicine

Production of Apolipoprotein C-III Knockout Rabbits using Zinc Finger Nucleases

Dongshan Yang¹, Jifeng Zhang¹, Jie Xu¹, Tianqing Zhu¹, Yanbo Fan¹, Jianglin Fan², Y. Eugene Chen¹

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.

Neuroscience

Observation of the Ciliary Movement of Choroid Plexus Epithelial Cells Ex Vivo

Takafumi Inoue¹, Keishi Narita², Yuta Nonami¹, Hideki Nakamura¹, Sen Takeda²

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.

Developmental Biology

Zygotic Fluorescence Recovery After Photo-bleaching Analysis for Chromatin Looseness That Allows Full-term Development

Masatoshi Ooga^1,3, Satoshi Funaya², Fugaku Aoki², 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.

Chemistry

Preparation of Polyoxometalate-based Photo-responsive Membranes for the Photo-activation of Manganese Oxide Catalysts

Akira Yamaguchi^1,5, Toshihiro Takashima², 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.

Biochemistry

Sample Preparation for Probe Electrospray Ionization Mass Spectrometry

Sen Takeda¹, Kentaro Yoshimura¹, Hiroshi Tanihata²

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.

Biochemistry

Isolation and Analysis of Plasma Lipoproteins by Ultracentrifugation

Manabu Niimi¹, Haizhao Yan¹, Yajie Chen¹, Yao Wang², 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.

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.

Neuroscience

In Vivo Wide-Field and Two-Photon Calcium Imaging from a Mouse Using a Large Cranial Window

Satoshi Manita¹, Eiji Shigetomi^2,3, Haruhiko Bito⁴, Schuichi Koizumi^2,3, Kazuo Kitamura¹

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.