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Doshisha University

14 ARTICLES PUBLISHED IN JoVE

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Neuroscience

Morphological Analysis of Drosophila Larval Peripheral Sensory Neuron Dendrites and Axons Using Genetic Mosaics
M. Rezaul Karim 1,2, Adrian W. Moore 1
1Disease Mechanism Research Core, RIKEN Brain Science Institute, 2Graduate School of Science and Engineering, Saitama University

The dendritic arborization sensory neurons of the Drosophila larval peripheral nervous system are useful models to elucidate both general and neuron class-specific mechanisms of neuron differentiation. We present a practical guide to generate and analyze dendritic arborization neuron genetic mosaics.

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Neuroscience

Immunohistological Labeling of Microtubules in Sensory Neuron Dendrites, Tracheae, and Muscles in the Drosophila Larva Body Wall
Cagri Yalgin 1,2, M. Rezaul Karim 1,2, Adrian W. Moore 1
1Disease Mechanism Research Core, RIKEN Brain Science Institute, 2Graduate School of Science and Engineering, Saitama University

To understand how complex cell shapes, such as neuronal dendrites, are achieved during development, it is important to be able to accurately assay microtubule organization. Here we describe a robust immunohistological labeling method to examine microtubule organization of dendritic arborization neuron sensory dendrites, trachea, muscle, and other Drosophila larva body wall tissues.

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Neuroscience

Whole Mount Immunolabeling of Olfactory Receptor Neurons in the Drosophila Antenna
M. Rezaul Karim 1, Keita Endo 2, Adrian W Moore 3, Hiroaki Taniguchi 1
1Laboratory for Genetic Code, Graduate School of Life and Medical Sciences, Doshisha University, 2Laboratory for Circuit Mechanisms of Sensory Perception, RIKEN Brain Science Institute, 3Disease Mechanism Research Core, RIKEN Brain Science Institute

Herein we describe the process of whole mount immunostaining of Drosophila antennae, which enables us to better understand the molecular mechanisms involved in the diversification of olfactory receptor neurons (ORN)s.

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Developmental Biology

An Efficient Method to Obtain Dedifferentiated Fat Cells
Hiroaki Taniguchi *1, Tomohiko Kazama *1, Kazuhiro Hagikura *1, Chii Yamamoto 1, Minako Kazama 1, Yuki Nagaoka 1, Taro Matsumoto 1
1Division of Cell Regeneration and Transplantation, School of Medicine, Nihon University

We have modified the conditions for DFAT cell generation and provide herein information regarding the use of an improved growth medium for the production of these cells.

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Developmental Biology

Imaging of Cell Shape Alteration and Cell Movement in Drosophila Gastrulation Using DE-cadherin Reporter Transgenic Flies
M. Rezaul Karim 1, Tomohiro Haruta 2, Taro Matsumoto 3, Hiroki Oda 2, Hiroaki Taniguchi 3,4,5
1Biotechnology and Genetic Engineering Department, Jahangirnagar University, 2JT Biohistory Research Hall, 3Division of Cell Regeneration and Transplantation, School of Medicine, Nihon University, 4Graduate School of Life and Medical Sciences, Doshisha University, 5Institute of Genetics and Animal Breeding of the Polish Academy of Sciences

Herein we describe a procedure to capture live images of Drosophila gastrulation. This has enabled us to better understand the apical constriction involved in early development and further analyze mechanisms governing cellular movements during tissue structure modification.

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Biochemistry

A Protein Preparation Method for the High-throughput Identification of Proteins Interacting with a Nuclear Cofactor Using LC-MS/MS Analysis
Megumi Tsuchiya *1, M. Rezaul Karim *2, Taro Matsumoto 3, Hidesato Ogawa 1, Hiroaki Taniguchi 3,4,5
1Graduate School of Frontier Biosciences, Osaka University, 2Department of Biotechnology and Genetic Engineering, Jahangirnagar University, 3Division of Cell Regeneration and Transplantation, School of Medicine, Nihon University, 4Institute of Genetics and Animal Breeding of the Polish Academy of Sciences, 5Graduate School of Life and Medical Sciences, Doshisha University

We have established a method for the purification of coregulatory interaction proteins using the LC-MS/MS system.

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Immunology and Infection

Evaluation of Blood Lactate and Plasma Insulin During High-intensity Exercise by Antecubital Vein Catheterization
Minas Nalbandian 1, Zsolt Radak 2, Masaki Takeda 3
1Graduate School of medicine, Kyoto University, 2Research Institute of Sports Science, University of Physical Education, 3Graduate School of Sports and Health Science, Doshisha University

Here we present a protocol to obtain several blood samples from the antecubital vein during high-intensity interval training. This protocol may be useful for the measurement of blood metabolites and endocrinal markers during exercise.

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Neuroscience

Visualization of Amyloid β Deposits in the Human Brain with Matrix-assisted Laser Desorption/Ionization Imaging Mass Spectrometry
Masaya Ikegawa *1, Takashi Nirasawa *2, Nobuto Kakuda 1, Tomohiro Miyasaka 1, Yuki Kuzuhara 1, Shigeo Murayama 3, Yasuo Ihara 4
1Department of Life and Medical Systems, Doshisha University, 2Bruker Daltonics K.K., 3The Brain Bank for Aging Research, Tokyo Metropolitan Geriatric Hospital and Institute of Gerontology, 4Graduate School of Brain Science, Doshisha University

Molecular imaging with matrix-assisted laser desorption/ionization-based imaging mass spectrometry (MALDI-IMS) allows the simultaneous mapping of multiple analytes in biological samples. Here, we present a protocol for the detection and visualization of amyloid β protein on brain tissues of Alzheimer's disease and cerebral amyloid angiopathy samples using MALDI-IMS.

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Biochemistry

Manipulating Living Cells to Construct Stable 3D Cellular Assembly Without Artificial Scaffold
Takehiro Yamazaki *1, Hiroaki Taniguchi *2, Shoto Tsuji 1, Shiho Sato 1, Takahiro Kenmotsu 1, Kenichi Yoshikawa 1, Koichiro Sadakane 1
1Faculty of Life and Medical Sciences, Doshisha University, 2The Institute of Genetics and Animal Breeding, Polish Academy of Sciences

We demonstrate a novel method for constructing a single-cell-based 3-dimensional (3D) assembly without an artificial scaffold.

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Behavior

Automated, Long-term Behavioral Assay for Cognitive Functions in Multiple Genetic Models of Alzheimer's Disease, Using IntelliCage
Akira Masuda 1,2, Yuki Kobayashi 1, Shigeyoshi Itohara 1
1Laboratory for Behavioral Genetics, Center for Brain Science, RIKEN, 2Organization for Research Initiatives and Development, Doshisha University

This paper describes a protocol for cognitive assessments for genetic models of the Alzheimer's disease using the IntelliCage system, which is a high throughput automated behavioral monitoring system with operant conditioning.

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Developmental Biology

Neurogenesis Using P19 Embryonal Carcinoma Cells
Paweł Leszczyński 1, Magdalena Śmiech 1, Aamir S. Teeli 1, Aleksandra Zołocińska 2, Anna Słysz 2, Zygmunt Pojda 2, Mariusz Pierzchała 3, Hiroaki Taniguchi 1
1Department of Experimental Embryology, Institute of Genetics and Animal Breeding, Polish Academy of Sciences, 2Department of Regenerative Medicine, Maria Skłodowska-Curie Institute - Oncology Center, 3Department of Genomics and Biodiversities, Institute of Genetics and Animal Breeding, Polish Academy of Sciences

The P19 mouse embryonic carcinoma cell line (P19 cell line) is widely used for studying the molecular mechanism of neurogenesis with great simplification compared to in vivo analysis. Here, we present a protocol for retinoic acid-induced neurogenesis in the P19 cell line.

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Behavior

A Task for Assessing the Impact of a Partner on the Speed and Accuracy of Motor Performance in Rats
Yayoi Sekiguchi 1,2, Toshimichi Hata 3
1Graduate School of Psychology, Doshisha University, 2Japan Society for the Promotion of Science, 3Faculty of Psychology, Doshisha University

A procedure to measure the speed and accuracy of rats’ motor performance in a social condition is described. The protocol enables us to investigate the effect of the mere presence of others on speed and accuracy of motor performance in one experiment.

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Biochemistry

Quantitative Microtubule Fractionation Technique to Separate Stable Microtubules, Labile Microtubules, and Free Tubulin in Mouse Tissues
Ayaka Hagita-Tatsumoto 1,2, Tomohiro Miyasaka 1,2,3
1Department of Neuropathology, Faculty of Life and Medical Sciences, Doshisha University, 2Center for Research in Neurodegenerative Diseases, Doshisha University, 3Laboratory of Physiology and Anatomy, School of Pharmacy, Nihon University

Microtubules, which are tubulin polymers, play a crucial role as a cytoskeleton component in eukaryotic cells and are known for their dynamic instability. This study developed a method for fractionating microtubules to separate them into stable microtubules, labile microtubules, and free tubulin to evaluate the stability of microtubules in various mouse tissues.

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Behavior

Utilizing a Reconfigurable Maze System to Enhance the Reproducibility of Spatial Navigation Tests in Rodents
Fumiya Sawatani 1, Yuta Tamatsu 1, Kaoru Ide 1, Hirotsugu Azechi 1, Susumu Takahashi 1
1Laboratory of Cognitive and Behavioral Neuroscience, Graduate School of Brain Science, Doshisha University

The present protocol describes a reconfigurable maze, a unique system for testing spatial navigation and behavioral phenotypes in rodents. The adaptability of this maze system enables the execution of various experiments in a single physical environment. The ease of structural rearrangement generates reliable and reproducible experimental results.

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