JoVE Neuroscience
Reproducible Mouse Sciatic Nerve Crush and Subsequent Assessment of Regeneration by Whole Mount Muscle Analysis
Center for Neural Repair and Rehabilitation, Temple University
In this report we describe a method to crush mouse sciatic nerve. This method uses readily available hemostatic forceps and easily and reproducibly produces complete sciatic nerve crush. In addition, we describe a method to prepare muscle whole mounts suitable for analysis of nerve regeneration after sciatic nerve crush.
JoVE Neuroscience
Chromatin Immunoprecipitation from Dorsal Root Ganglia Tissue following Axonal Injury
1Laboratory for NeuroRegeneration and Repair, Department of Neurology, Hertie Institute for Clinical Brain Research, University of Tuebingen, 2Graduate School for Cellular and Molecular Neuroscience, University of Tuebingen
We present a method for chromatin immunoprecipitation from dorsal root ganglia tissue following axonal injury. The approach can be used to identify specific transcription factor binding sites and epigenetic modification of histone and DNA important for the regeneration of injured axons in both the peripheral and central nervous system.
JoVE Neuroscience
Axoplasm Isolation from Rat Sciatic Nerve
Department of Biological Chemistry, Weizmann Institute of Science
We demonstrate a protocol for axoplasm isolation from adult rat sciatic nerve based on dissection of nerve fascicles and incubation in hypotonic medium to release myelin and lyse non-axonal structures, followed by extraction of the remaining axon-enriched material.
JoVE Clinical and Translational Medicine
Chronic Constriction of the Sciatic Nerve and Pain Hypersensitivity Testing in Rats
School of Medical Sciences, University of New South Wales
Due to the simplicity of surgery and the robust behavioural outcome, chronic constriction of the sciatic nerve is one of the pre-eminent animal models of neuropathic pain. Within 24 hrs following surgery, pain hypersensitivity is established and can be quantitatively measured using a von Frey aesthesiometer (mechanical test) and plantar analgesia meter (thermal test).
JoVE Clinical and Translational Medicine
The Spared Nerve Injury (SNI) Model of Induced Mechanical Allodynia in Mice
1The Lundbeck Foundation Research Center MIND, Department of Biomedicine, Aarhus University, 2Department of Pharmacology and Pharmacotherapy, Faculty of Pharmaceutical Sciences, University of Copenhagen
The Spared Nerve Injury animal model is described here as a mouse model of peripheral neuropathic pain following partial denervation of the sciatic nerve by lesioning the tibial and common peroneal nerve branches, leaving the remaining sural nerve intact. Behavioral modification resulting from mechanical allodynia is quantified by von Frey filaments.
JoVE Neuroscience
An Optic Nerve Crush Injury Murine Model to Study Retinal Ganglion Cell Survival
1National Eye Institute, NIH, 2Ophthalmology Department, The Second Hospital of Harbin Medical University
This protocol shows how to retrogradely label retinal ganglion cells, and how to subsequently make an optic nerve crush injury in order to analyze retinal ganglion cell survival and apoptosis. It is an experimental disease model for different types of optic neuropathy, including glaucoma.
JoVE General
Procedures for Rat in situ Skeletal Muscle Contractile Properties
Faculty of Kinesiology, University of Calgary
This video demonstrates the surgical preparation and procedures needed to study the contractile responses of the rat medial gastrocnemius muscle preparation in situ. This preparation allows measurement of skeletal muscle contractile properties under physiological conditions. The animal is anesthetized and the muscle is separated from surrounding tissue at its distal end. The Achilles tendon is attached to a force transducer, allowing measurement of the muscle’s contractile response at 37 degrees C with an intact circulation.
JoVE General
Evaluation of Muscle Function of the Extensor Digitorum Longus Muscle Ex vivo and Tibialis Anterior Muscle In situ in Mice
Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri
Changes in limb muscle contractile and passive mechanical properties are important biomarkers for muscle diseases. This manuscript describes physiological assays to measure these properties in the murine extensor digitorum longus and tibialis anterior muscles.
JoVE General
Modeling Biological Membranes with Circuit Boards and Measuring Electrical Signals in Axons: Student Laboratory Exercises
1Department of Biology, University of Kentucky, 2Department of Physiology, University of Toronto
This is a demonstration of how biological membranes can be understood using electrical models. We also demonstrate procedures for recording action potentials from the ventral nerve cord of the crayfish for student orientated laboratories.
JoVE Clinical and Translational Medicine
Orthotopic Xenografting of Human Luciferase-Tagged Malignant Peripheral Nerve Sheath Tumor Cells for in vivo Testing of Candidate Therapeutic Agents
1Department of Pathology, University of Alabama at Birmingham - UAB, 2Department of Radiology, University of Alabama at Birmingham - UAB, 3Department of Cell Biology and Neurobiology, University of Alabama at Birmingham - UAB
A method for reliably grafting luciferase-tagged human malignant peripheral nerve sheath tumor cells into the sciatic nerve of immunodeficient mice is described. The use of bioluminescence imaging to demonstrate proper establishment of tumor grafts and criteria for random segregation of animals into study groups are also discussed.
JoVE General
Combining Peripheral Nerve Grafting and Matrix Modulation to Repair the Injured Rat Spinal Cord
Department of Neurobiology and Anatomy, Drexel University College of Medicine
Traumatic injury to the spinal cord disrupts communication with the brain. To restore lost connectivity we utilize a peripheral nerve graft to provide a substratum for regenerating fibers in combination with neurotrophic factors and matrix-modulating enzymes to remove inhibitory molecules to promote long distance growth.
JoVE Neuroscience
Simultaneous Intracellular Recording of a Lumbar Motoneuron and the Force Produced by its Motor Unit in the Adult Mouse In vivo
Department of Physiology, Northwestern University Feinberg School of Medicine
This new method permits the simultaneous intracellular recording of a single adult mouse motoneuron and the measurement of the force produced by its muscle fibers. The combined investigation of the electrical and mechanical properties of motor units in normal and genetically modified animals is a breakthrough for the study of the neuromuscular system.
JoVE General
Using the optokinetic response to study visual function of zebrafish
Optokinetic response has been widely used to assess the visual functions of larval zebrafish. Nevertheless, the standard protocol for larval fish is not yet readily applicable in adults1-5. Here, we introduce how to measure the OKR of adult zebrafish using a new protocol which is established in our lab.
JoVE Neuroscience
Isolation and Culture of Neural Crest Stem Cells from Human Hair Follicles
Department of Pathology and Laboratory Medicine, School of Medicine, University of Pennsylvania
This article presents a robust protocol for isolation and culture of neural crest stem cells from human hair follicles.
JoVE Bioengineering
Fabrication and Application of Rose Bengal-chitosan Films in Laser Tissue Repair
1Bioelectronics and Neuroscience (BENS) research group, University of Western Sydney, NSW Australia, 2Australian School of Advanced Medicine, Macquarie University, NSW Australia, 3School of Medicine, University of Siena, Italy
Sutures are usually needed to repair tissue during surgical procedures. However, their application can be problematic as they are invasive and may damage tissue. The fabrication and application methods of a novel tissue adhesive are here reported. This adhesive film is laser-activated and does not require the use of sutures.
JoVE General
Analysis of Schwann-astrocyte Interactions Using In Vitro Assays
Cambridge Centre for Brain Repair, Department of Clinical Neurosciences, University of Cambridge
This article intends to describe in stepwise fashion the commonly used in vitro assays used in studying Schwann cell-asrtocyte interactions.
JoVE Neuroscience
Live Imaging of Dorsal Root Axons after Rhizotomy
1Temple University, Shriners Hospitals Pediatric Research Center and Department of Anatomy and Cell Biology, 2Medical Research Service, Department of Veterans Affairs Hospital, 3Department of Neurobiology and Anatomy, Drexel University College of Medicine, 4Shriners Hospitals Pediatric Research Center and Department of Anatomy and Cell Biology, Temple University School of Medicine
An in vivo imaging protocol to monitor primary sensory axons following dorsal root crush is described. The procedures utilize wide-field fluorescence microscopy and thy1-YFP transgenic mice, and permit repeated imaging of axon regeneration over 4 cm in the PNS and axon interactions with the interface of the CNS.
JoVE Clinical and Translational Medicine
Biomarkers in an Animal Model for Revealing Neural, Hematologic, and Behavioral Correlates of PTSD
1Department of Psychiatry, Center for the Study of Traumatic Stress, Uniformed Services University of the Health Sciences, Bethesda, Maryland, 2Department of Gene and Protein Biomarkers, GenProMarkers, Inc.
We describe a rat model of post traumatic stress disorder (PTSD) that reveals the persistent alterations in neuroendocrine function and the delayed long-term, exaggerated fear response, characteristic of PTSD patients. The animal model and methods described here are useful for correlating biomarkers in brain nuclei, which are mechanistic but cannot be measured in patients, with biomarkers in peripheral white blood cells, which can.
JoVE Neuroscience
An Optimized Procedure for Fluorescence-activated Cell Sorting (FACS) Isolation of Autonomic Neural Progenitors from Visceral Organs of Fetal Mice
Division of Genetic Medicine, Department of Medicine, Vanderbilt University School of Medicine
An optimized procedure to purify neural crest-derived neuronal progenitors from fetal mouse tissues is described. This method takes advantage of expression from fluorescent reporter alleles to isolate discrete populations by fluorescence-activated cell sorting (FACS). The technique can be applied to isolate neuronal subpopulations throughout development or from adult tissues.
JoVE General
Microdissection of Zebrafish Embryonic Eye Tissues
Department of Biological Sciences, Purdue University
This article describes an approach to microdissect zebrafish retinas with and without retinal pigment epithelium attached, from one to three days postfertilization embryos.
JoVE Neuroscience
Analyzing Murine Schwann Cell Development Along Growing Axons
1Department of Molecular Embryology, Institute of Anatomy and Cell Biology, University of Freiburg, 2Department of Neuroanatomy, University of Heidelberg, 3FRIAS, University of Freiburg
Here we describe a Schwann cell (SC) migration assay in which SCs are able to develop along extending axons.
JoVE Clinical and Translational Medicine
Intraspinal Cell Transplantation for Targeting Cervical Ventral Horn in Amyotrophic Lateral Sclerosis and Traumatic Spinal Cord Injury
Department of Neuroscience, Thomas Jefferson University Medical College
Neural precursor transplantation is a promising strategy for protecting and/or replacing lost/dysfunctional cervical phrenic motor neurons in spinal cord injury (SCI) and the motor neuron disorder, amyotrophic laterals sclerosis (ALS). We provide a protocol for cell delivery to cervical spinal cord ventral horn in rodent models of ALS and SCI.
JoVE Clinical and Translational Medicine
Pseudofracture: An Acute Peripheral Tissue Trauma Model
1Department of Surgery, University of Pittsburgh, 2Department of Orthopedic Surgery, University of Aachen Medical Center
Pseudofracture, a reproducible murine model of sterile musculoskeletal trauma, allows for evaluation of late term post-traumatic immune responses. This article describes the procedural execution of the model step by step, including the potential for experimental model combinations to permit study of multiple trauma.
JoVE General
Thin Sectioning of Slice Preparations for Immunohistochemistry
1Department of Medicine, Yonsei University College of Medicine, Severance Hospital, 2Department of Psychiatry, Harvard Medical School
The present method allows reproducible cryostat sectioning of small, difficult-to-manage, tissue pieces, such as biopsies and brain slices. We utilize a simple aluminum freezing stage to facilitate handling of tissue and a standard cryostat to routinely produce 5-10 micron serial sections from 400 micron thick brain slices.
JoVE Clinical and Translational Medicine
Surgical Management of Meatal Stenosis with Meatoplasty
Department of Urology, Pediatric Urology, Johns Hopkins School of Medicine
Meatoplasty, surgical management of meatal stenosis.
JoVE Neuroscience
An In Vitro Preparation for Eliciting and Recording Feeding Motor Programs with Physiological Movements in Aplysia californica
1Department of Biology, Case Western Reserve University, 2Department of Neurosciences, Case Western Reserve University, 3Department of Biomedical Engineering, Case Western Reserve University
We describe a technique to extracellularly record and stimulate from nerves, muscles, and individual identified neurons in vitro while eliciting and observing different types of feeding behaviors in the feeding apparatus of Aplysia.
JoVE Neuroscience
Selective Tracing of Auditory Fibers in the Avian Embryonic Vestibulocochlear Nerve
Department of Neurobiology and Behavior, University of California, Irvine
Here we describe a microdissection technique followed by fluorescent dye injection into the acoustic ganglion of early chick embryos for selective tracing of auditory axon fibers in the nerve and hindbrain.
JoVE Neuroscience
Electrode Fabrication and Implantation in Aplysia californica for Multi-channel Neural and Muscular Recordings in Intact, Freely Behaving Animals
1Biology, Case Western Reserve University, 2Neurosciences, Case Western Reserve University, 3Biomedical Engineering, Case Western Reserve University
A technique is described for implanting four in vivo electrodes to monitor the neuromuscular control of feeding behavior in Aplysia californica.
JoVE Clinical and Translational Medicine
Utilizing Transcranial Magnetic Stimulation to Study the Human Neuromuscular System
Transcranial magnetic stimulation (TMS) is a non-invasive tool to gain insight on the physiology and function of the human nervous system. Here, we present our TMS techniques to study cortical excitability of the upper limb and lumbar musculature.
JoVE Neuroscience
Extracellularly Identifying Motor Neurons for a Muscle Motor Pool in Aplysia californica
1Department of Biology, Case Western Reserve University, 2Department of Neurosciences, Case Western Reserve University, 3Department of Biomedical Engineering, Case Western Reserve University
In animals with large identified neurons (e.g. mollusks), analysis of motor pools is done using intracellular techniques1,2,3,4. Recently, we developed a technique to extracellularly stimulate and record individual neurons in Aplysia californica5. We now describe a protocol for using this technique to uniquely identify and characterize motor neurons within a motor pool.
JoVE General
Isolation and Transplantation of Hematopoietic Stem Cells (HSCs)
JoVE General
Homing of Hematopoietic Cells to the Bone Marrow
Center for Regenerative Medicine, MGH - Massachusetts General Hospital
This article describes a protocol used to study the homing of hematopoietic cells to their niches in the bone marrow.
JoVE General
Intraperitoneal Injection into Adult Zebrafish
1Department of Organismal Biology and Anatomy, The University of Chicago, 2Committee on Molecular Metabolism and Nutrition, The University of Chicago, 3Department of Medicine, The University of Chicago
We demonstrate intraperitoneal injection into adult zebrafish. We use a 10 μl NanoFil microsyringe controlled by a Micro4 controller and UltraMicroPump III. This demonstration includes the use of cold water as an anesthetic.
JoVE Clinical and Translational Medicine
MALDI Imaging Mass Spectrometry of Neuropeptides in Parkinson's Disease
1Department of Pharmaceutical Biosciences, Uppsala University, 2Department of Chemical and Biological Engineering, Chalmers University of Technology
Dopamine replacement pharmacotherapy using L-DOPA is the most commonly used symptomatic treatment of Parkinson’s disease, but is accompanied by side effects including involuntary abnormal movements, termed dyskinesia 1. Here, a protocol for MALDI imaging mass spectrometry is presented that detects changes in rat brain neuropeptide levels related to dyskinesia.
JoVE Clinical and Translational Medicine
A Contusive Model of Unilateral Cervical Spinal Cord Injury Using the Infinite Horizon Impactor
1International Collaboration on Repair Discoveries (ICORD), University of British Columbia, 2Department of Orthopaedics, University of British Columbia
A reliable and repeatable way to produce a cervical unilateral spinal cord injury using the Infinite Horizon impactor is described. The method takes advantage of a custom designed frame and clamp to stabilize the spine. The standardized procedure and biomechanical injury parameters result in sufficient and sustained injuries.
JoVE Neuroscience
Nerve Excitability Assessment in Chemotherapy-induced Neurotoxicity
1Prince of Wales Clinical School, University of New South Wales, 2Neuroscience Research Australia, University of New South Wales, 3School of Medical Sciences, University of New South Wales
This abstract describes a novel method to assess the development of neurotoxicity in patients receiving chemotherapy treatment. While conventional assessment methods are limited in their ability to detect early changes in nerve function, nerve excitability techniques provide early identification of patients at risk of severe neurotoxicity and insight into pathophysiology.
JoVE General
Electroeluting DNA Fragments
This procedure allows the purification of DNA fragments with high yield.
JoVE Neuroscience
Optic Nerve Transection: A Model of Adult Neuron Apoptosis in the Central Nervous System
Department of Surgery, University of Toronto
Optic Nerve transection is a widely used model of adult CNS injury. Ninety percent of retinal ganglion cells (RGCs) whose axons are completely transected (axotomy) die within 14 days after axotomy. This model is easily amenable to experimental manipulations and highly reproducible.
JoVE Clinical and Translational Medicine
Controlled Cervical Laceration Injury in Mice
1Norton Neuroscience Institute, Norton Healthcare, 2Spinal Cord and Brain Injury Research Group, Stark Neurosciences Research Institute, Department of Neurological Surgery and Goodman and Campbell Brain and Spine, Medical Neuroscience Graduate Program, and Department of Anatomy and Cell Biology, Indiana University School of Medicine
A novel technique to create a reproducible in vivo model of cervical spinal cord laceration injury in the mouse is described. This technique is based on spine stabilization by fixation of the cervical facets and laceration of the spinal cord using an oscillating blade with an accuracy of ±0.01 mm.
JoVE Neuroscience
Tissue Preparation and Immunostaining of Mouse Sensory Nerve Fibers Innervating Skin and Limb Bones
1Department of Pharmacology, The University of Iowa, 2Department of Anesthesia, Roy J. and Lucille A. Carver College of Medicine, The University of Iowa
Immunocytochemical identification of peripheral sensory nerve fiber subtypes (and detection of protein expression therein) are key to the understanding of molecular mechanisms underlying peripheral sensation. Here we describe methods for preparation of peripheral/visceral tissue samples, such as skin and limb bones, for specific immunostaining of peripheral sensory nerve fibers.
JoVE Clinical and Translational Medicine
Three-dimensional Imaging of Nociceptive Intraepidermal Nerve Fibers in Human Skin Biopsies
1Department of Neurology, University of Michigan, 2Department of Internal Medicine, University of Michigan
In order to study the changes of nociceptive intraepidermal nerve fibers (IENFs) in painful neuropathies (PN), we developed protocols that could directly examine three-dimensional morphological changes observed in nociceptive IENFs. Three-dimensional analysis of IENFs has the potential to evaluate the morphological changes of IENF in PN.
JoVE General
Homarus Americanus Stomatogastric Nervous System Dissection
Volen Center for Complex Systems, Brandeis
We describe the fine dissection of the stomatogastric nervous system from the stomach of the American lobster (Homarus americanus).
JoVE Clinical and Translational Medicine
Corneal Confocal Microscopy: A Novel Non-invasive Technique to Quantify Small Fibre Pathology in Peripheral Neuropathies
Division of Cardiovascular Medicine, University of Manchester
Corneal Confocal microscopy is a non-invasive clinical technique which may be used to quantify C fibre damage to diagnose and stratify patients with increasing neuropathic severity.
JoVE Neuroscience
Methods for Experimental Manipulations after Optic Nerve Transection in the Mammalian CNS
Department of Surgery, University of Toronto
Optic Nerve transection is a widely used model of adult CNS injury. This model is ideal for performing a number of experimental manipulations that target the retina globally or directly target the injured neuronal population of retinal ganglion cells.
JoVE General
A High-Throughput Method For Zebrafish Sperm Cryopreservation and In Vitro Fertilization
1Molecular and Cellular Biology, University of California, Davis, 2Division of Basic Science, Fred Hutchinson Cancer Research Center - FHCRC
This is a high-throughput sperm cryopreservation protocol for zebrafish. Sperm cryopreserved using this protocol has an average of 25% fertility in subsequent vitro fertilization and is stable over many years.
JoVE Clinical and Translational Medicine
Quantifying Glomerular Permeability of Fluorescent Macromolecules Using 2-Photon Microscopy in Munich Wistar Rats
Medicine/Nephrology, Indiana University School of Medicine
A technique utilizing high resolution intavital 2-photon microscopy to directly visualize and quantify gloemrular filtration in surface glomeruli. This method allows for direct determination of permeability characteristics of macromolecules in both normal and diseased states.
JoVE General
Efficient and Rapid Isolation of Early-stage Embryos from Arabidopsis thaliana Seeds
Institute of Plant Biology and Zürich-Basel Plant Science Center, University of Zürich
We report an efficient and simple method to isolate embryos at early stages of development from Arabidopsis thaliana seeds. Up to 40 embryos can be isolated in 1 hr to 4 hr, depending on the downstream application. The procedure is suitable for transcriptome, DNA methylation, reporter gene expression, immunostaining and fluorescence in situ hybridization analyses.
JoVE General
Loading Drosophila Nerve Terminals with Calcium Indicators
Department of Physiology, University of Texas Health Science Center at San Antonio (UTHSCSA)
Calcium is a ubiquitous messenger in the nervous system, essential for triggering neurotransmitter release and changes in synaptic strength. Here we demonstrate a technique for loading Ca2+-indicators into Drosophila nerve terminals. We also demonstrate fabrication of the required apparatus and emphasize points critical for the technique's success.
JoVE Neuroscience
Subcutaneous Administration of Muscarinic Antagonists and Triple-Immunostaining of the Levator Auris Longus Muscle in Mice
1Biology Department, Arcadia University, 2Shriners Hospitals Pediatric Research Center, Temple University School of Medicine, 3Shriners Hospitals Pediatric Research Center and Department of Anatomy and Cell Biology, Temple University School of Medicine
We describe procedures for repeated administration of inhibitors of muscarinic signaling to the levator auris longus (LAL) muscle of young adult mice and for subsequent immunostaining of its neuromuscular junctions (NMJs) in wholemounts. The LAL muscle has unique advantages for revealing in vivo pharmacological effects on NMJs.
JoVE General
Cancer Borealis Stomatogastric Nervous System Dissection
Volen Center for Complex Systems, Brandeis
The stomatogastric nervous system (STNS) of the Jonah crab (C. borealis) can be used for electrophysiology, immunohistochemistry, and cell culture studies. The STNS extraction is done in two parts: the gross and fine dissection.
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