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National Institute of Neurological Disorders and Stroke

9 ARTICLES PUBLISHED IN JoVE

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Biology

Methods for Patch Clamp Capacitance Recordings from the Calyx
Kenneth Paradiso 1, Wei Wu 1, Ling-Gang Wu 1
1National Institute of Neurological Disorders and Stroke, National Institute of Health

We demonstrate the basic techniques for presynaptic patch clamp recording at the calyx of Held, a mammalian central nervous system nerve terminal.

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Neuroscience

Ex vivo Culturing of Whole, Developing Drosophila Brains
Ranjini Prithviraj 1,2, Svetlana Trunova 1,2, Edward Giniger 1,2
1National Institute of Neurological Disorders and Stroke, 2National Human Genome Research Institute, National Institutes of Health, Bethesda, MD

This article describes a method by which one can mimic in vivo development of the Drosophila mushroom body in an ex vivo culture system.

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Neuroscience

Utilizing 3D Printing Technology to Merge MRI with Histology: A Protocol for Brain Sectioning
Nicholas J Luciano 1, Pascal Sati 1, Govind Nair 1, Joseph R Guy 1, Seung-Kwon Ha 1, Martina Absinta 1, Wen-Yang Chiang 2, Emily C Leibovitch 3, Steven Jacobson 3, Afonso C Silva 2, Daniel S. Reich 1
1Translational Neuroradiology Section, National Institute of Neurological Disorders and Stroke, 2Cerebral Microcirculation Section, National Institute of Neurological Disorders and Stroke, 3Viral Immunology Section, National Institute of Neurological Disorders and Stroke

The overall goal of this protocol is to accurately align magnetic resonance imaging (MRI) image volumes with histology sections via the creation of customized 3D-printed brain holders and slicer boxes.

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Neuroscience

Measuring Synaptic Vesicle Endocytosis in Cultured Hippocampal Neurons
Seth Villarreal 1, Sung Hoon Lee 1,2, Ling-Gang Wu 1
1National Institute of Neurological Disorders and Stroke, 2College of Pharmacy, Chung-ang University

Synaptic vesicle endocytosis is detected by light microscopy of pHluorin fused with synaptic vesicle protein and by electron microscopy of vesicle uptake.

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Neuroscience

Isolation of Adult Spinal Cord Nuclei for Massively Parallel Single-nucleus RNA Sequencing
Kaya J.E. Matson 1, Anupama Sathyamurthy 1, Kory R. Johnson 2, Michael C. Kelly 3,4, Matthew W. Kelley 3, Ariel J. Levine 1
1Spinal Circuits and Plasticity Unit, National Institute of Neurological Disorders and Stroke, 2Bioinformatics Section, Information Technology Program, National Institute of Neurological Disorders and Stroke, 3Laboratory of Cochlear Development, National Institute on Deafness and Other Communication Disorders, 4Single Cell Analysis Facility, Frederick National Laboratory

Here, we present a protocol to rapidly isolate high-quality nuclei from the fresh or frozen tissue for downstream massively parallel RNA sequencing. We include detergent-mechanical and hypotonic-mechanical tissue disruption and cell lysis options, both of which can be used for isolation of nuclei.

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Bioengineering

3D Analysis of Multi-cellular Responses to Chemoattractant Gradients
Tae-Yun Kang 1, David Ellison 2, Sung Hoon Lee 1, Andrew J. Ewald 2,3, Andre Levchenko 1
1Department of Biomedical Engineering and Yale Systems Biology Institute, Yale University, 2Department of Biomedical Engineering, Johns Hopkins University, 3Center for Cell Dynamics and Department of Cell Biology, Johns Hopkins University

We describe a method to construct devices for 3D culture and experimentation with cells and multicellular organoids. This device allows analysis of cellular responses to soluble signals in 3D microenvironments with defined chemoattractant gradients. Organoids are better than single cells at detection of weak noisy inputs.

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Medicine

Invasive Hemodynamic Assessment for the Right Ventricular System and Hypoxia-Induced Pulmonary Arterial Hypertension in Mice
Fuliang Luo 1, Xin Wang 1, Xiaokang Luo 1, Bo Li 1, Desheng Zhu 2, Hansong Sun 3, Yue Tang 1
1Beijing Key Laboratory of Pre-clinical Research and Evaluation for Cardiovascular Implant Materials, State Key Laboratory of Cardiovascular Disease, Fuwai Hospital, National Centre for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College, 2Laboratory Animal Center, Peking University, 3Center of Cardiac Surgery for Adults, State Key Laboratory of Cardiovascular Disease, Fuwai Hospital, National Centre for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College

Here, we present a protocol to perform an invasive hemodynamic assessment of the right ventricle and pulmonary artery in mice using an open-chest surgery approach.

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Medicine

Technical Modification of the Terminal Ureter During Total Transperitoneal Laparoscopic Nephroureterectomy for Upper Urinary Tract Urothelial Carcinoma
Chunlong Fu *1,2, Yaqun Zhang *1, Xin Wang 1, Shengcai Zhu 1, Huimin Hou 1, Shengjie Liu 1, Cheng Pang 1, Xinda Song 1, Jia Chen 1, Jianye Wang 1, Ming Liu 1,2
1(Urology) Beijing Hospital, National Center of Gerontology, The Joint Unit of the Chinese Academy of Medical Sciences, 2Graduate School of Peking Union Medical College, Beijing Hospital

Here, we present a protocol to increase the surgical field of view and reduce the difficulty of total transperitoneal laparoscopic nephroureterectomy surgery by precutting the umbilical ligament before treating the terminal ureter.

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Neuroscience

Confocal Microscopy to Measure Three Modes of Fusion Pore Dynamics in Adrenal Chromaffin Cells
Sue Han *1, Xin Wang *1, Nicholas Cordero 1, Ling-Gang Wu 1
1National Institute of Neurological Disorders and Stroke

This protocol describes a confocal imaging technique to detect three fusion modes in bovine adrenal chromaffin cells. These fusion modes include 1) close-fusion (also called kiss-and-run), involving fusion pore opening and closure, 2) stay-fusion, involving fusion pore opening and maintaining the opened pore, and 3) shrink-fusion, involving fused vesicle shrinkage.

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