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35 articles published in JoVE
A Minimally Invasive Lesion Technique for Muscles Intrinsic to the Odontophore of Aplysia californica Catherine Kehl1, Hillel J. Chiel1,2,3,4 1Department of Biology, Case Western Reserve University, 2Department of Biology, Case Western Reserve University, 3Department of Neurosciences, Case Western Reserve University, 4Department of Biomedical Engineering, Case Western Reserve University Here we present a protocol for minimally invasive surgical lesioning of muscles intrinsic to the feeding apparatus of the marine mollusk Aplysia californica to understand the roles of these muscles during feeding behavior.
A Novel Single Animal Motor Function Tracking System Using Simple, Readily Available Software Keith R. Dona1,2, Monika Goss-Varley1,2, Andrew J. Shoffstall1,2, Jeffrey R. Capadona1,2 1Advanced Platform Technology Center, Rehabilitation Research and Development, Louis Stokes Cleveland Department of Veterans Affairs Medical Center, 2Department of Biomedical Engineering, Case Western Reserve University The current study aimed to automate the quantification of motor deficits in rats. The initial evaluation model assesses motor loss resulting from an intracortical microelectrode implantation in the motor cortex. We report on the development and use of a tracking algorithm using easily adaptable, simple, and readily available coding software.
Quantitative Analysis of Alternative Pre-mRNA Splicing in Mouse Brain Sections Using RNA In Situ Hybridization Assay Xuan Guo1,2, Yiqing Zhao1,3, Hieu Nguyen1, Tonghua Liu2, Zhenghe Wang1,3, Hua Lou1,3,4 1Department of Genetics and Genome Sciences, Case Western Reserve University, 2Department of Endocrinology, Dongfang Hospital of Beijing University of Chinese Medicine, 3Case Comprehensive Cancer Center, Case Western Reserve University, 4Center for RNA Science and Therapeutics, Case Western Reserve University An in situ hybridization (ISH) protocol that uses short antisense oligonucleotides to detect alternative pre-mRNA splicing patterns in mouse brain sections is described.
Rodent Behavioral Testing to Assess Functional Deficits Caused by Microelectrode Implantation in the Rat Motor Cortex Monika Goss-Varley1,2, Andrew J. Shoffstall1,2, Keith R. Dona1,2, Justin A. McMahon1,2, Sydney C. Lindner1,2, Evon S. Ereifej1,2, Jeffrey R. Capadona1,2 1Advanced Platform Technology Center, Rehabilitation Research and Development, Louis Stokes Cleveland Department of Veterans Affairs Medical Center, 2Department of Biomedical Engineering, Case Western Reserve University We have shown that a microelectrode implantation in the motor cortex of rats causes immediate and lasting motor deficits. The methods proposed herein outline a microelectrode implantation surgery and three rodent behavioral tasks to elucidate potential changes in the fine or gross motor function due to implantation-caused damage to the motor cortex.
Conducting Elevated Temperature Normal and Combined Pressure-Shear Plate Impact Experiments Via a Breech-end Sabot Heater System Bryan Zuanetti1, Tiaxue Wang1, Vikas Prakash1 1Department of Mechanical and Aerospace Engineering, Case Western Reserve University Here, we present a detailed protocol of a new approach for conducting elevated temperature reverse normal plate impact, and combined pressure-and-shear plate impact. The approach involves the use of a breech-end resistive coil heater to heat a sample held at the front-end of a heat-resistant sabot to the desired temperature.
Development and Functional Characterization of Murine Tolerogenic Dendritic Cells Hsi-Ju Wei1, John J. Letterio2,3, Tej K. Pareek2,3 1Department of Biochemistry, School of Medicine, Case Western Reserve University, 2Department of Pediatrics, Division of Pediatric Hematology/Oncology, Case Western Reserve University, 3Angie Fowler Cancer Institute, Rainbow Babies & Children's Hospital, University Hospitals, Cleveland Here, we present a protocol to develop and characterize tolerogenic dendritic cells (TolDCs) and evaluate their immunotherapeutic utility.
Ultrasound-guided Intracardiac Injection of Human Mesenchymal Stem Cells to Increase Homing to the Intestine for Use in Murine Models of Experimental Inflammatory Bowel Diseases Maneesh Dave1, Paola Menghini1, Keiki Sugi2,3, Rodrigo A. Somoza5, Zhenghong Lee4, Mukesh Jain2,3, Arnold Caplan5, Fabio Cominelli1 1Division of Gastroenterology and Liver Disease, University Hospitals, Digestive Health Research Institute, Case Western Reserve University, 2Case Cardiovascular Research Institute, School of Medicine, Case Western Reserve University, 3Department of Medicine, Harrington Discovery Institute, Harrington Heart and Vascular Institute, University Hospitals Case Medical Center, 4Department of Radiology, University Hospitals Case Medical Center, 5Department of Biology - Skeletal Research Center, Case Western Reserve University Murine studies in models of colonic inflammation have demonstrated that a small percentage (1 - 5%) of mesenchymal stem cells (MSC) injected intravenously or intraperitoneally home to the inflamed colon1,2. This study shows that ultrasound-guided intracardiac injections of MSCs result in increased localization to the intestine.
Intrarenal Injection of Escherichia coli in a Rat Model of Pyelonephritis Karishma Gupta1, Shannon B Donnola1, Zhina Sadeghi3, Lan Lu1,3, Bernadette O. Erokwu1, Michael Kavran1, Adonis Hijaz3, Chris A. Flask1,2,4 1Department of Radiology, Case Western Reserve University, 2Department of Pediatrics, Case Western Reserve University, 3Department of Urology, Case Western Reserve University, 4Department of Biomedical Engineering, Case Western Reserve University This manuscript describes a rat surgical model of pyelonephritis using direct intra-renal infection by Escherichia coli into the renal pelvis. The experimental procedure can be utilized to study the pathogenesis of pyelonephritis as well as the associated inflammation and fibrosis.
Uncontrolled Hemorrhagic Shock Modeled via Liver Laceration in Mice with Real Time Hemodynamic Monitoring Mitchell Dyer1, Shannon Haldeman1, Andres Gutierrez1, Lauryn Kohut1, Anirban Sen Gupta2, Matthew D. Neal1 1Department of Surgery, University of Pittsburgh, 2Department of Biomedical Engineering, Case Western Reserve University Uncontrolled hemorrhage, an important cause of mortality among trauma patients, can be modeled using a standard liver laceration in a murine model. This model results in consistent blood loss, survival, and allows for testing hemostatic agents. This article provides the step-by-step process to perform this valuable model.
Fabrication of High Contact-Density, Flat-Interface Nerve Electrodes for Recording and Stimulation Applications Yazan M. Dweiri1,3, Matthew A. Stone2, Dustin J. Tyler1, Grant A. McCallum1, Dominique M. Durand1 1Neural Engineering Center, Case Western Reserve University, 2Advanced Platform Technology Center, U.S. Department of Veterans Affairs, 3Jordan University of Science and Technology This article provides a detailed description on the fabrication process of a high contact-density flat interface nerve electrode (FINE). This electrode is optimized for recording and stimulating neural activity selectively within peripheral nerves.
Ferric Chloride-induced Murine Thrombosis Models Wei Li1,2, Marvin Nieman3, Anirban Sen Gupta4 1Department of Cellular and Molecular Medicine, Lerner Research Institute, Cleveland Clinic, 2Department of Molecular Medicine, Cleveland Clinic Lerner College of Medicine of Case Western Reserve University, 3Department of Pharmacology, Case Western Reserve University, 4Department of Biomedical Engineering, Case Western Reserve University We report a refined procedure of the ferric chloride (FeCl3)-induced thrombosis models on carotid and mesenteric artery as well as vein, characterized efficiently using intravital microscopy to monitor time to occlusive thrombi formation.
Site Directed Spin Labeling and EPR Spectroscopic Studies of Pentameric Ligand-Gated Ion Channels Sandip Basak1, Soumili Chatterjee1, Sudha Chakrapani1,2 1Department of Physiology and Biophysics, Case Western Reserve University, 2School of Medicine, Case Western Reserve University This article describes methods for site-directed spin labeling and reconstitution of pentameric ligand-gated channels for Electron Paramagnetic Resonance studies. This protocol can be adapted for any membrane protein. The reconstitution method described here can also be used for patch-clamp measurements of macroscopic and single-channel currents in a defined lipid system.
Neural Activity Propagation in an Unfolded Hippocampal Preparation with a Penetrating Micro-electrode Array Mingming Zhang*1, Andrew B. Kibler*1, Luis E. Gonzales-Reyes1, Dominique M. Durand1 1Neural Engineering Center, Department of Biomedical Engineering, Case Western Reserve University We have developed an in vitro unfolded hippocampus which preserves CA1-CA3 array of neurons. Combined with the penetrating micro-electrode array, neural activity can be monitored in both the longitudinal and transverse orientations. This method provides advantages over hippocampal slice preparations as the propagation in the entire hippocampus can be recorded simultaneously.
Th17 Inflammation Model of Oropharyngeal Candidiasis in Immunodeficient Mice Natarajan Bhaskaran1, Aaron Weinberg1, Pushpa Pandiyan1 1Department of Biological Sciences, School of Dental Medicine, Case Western Reserve University Although Candida infection models are available to study host immune resistance, a model to study T cell mediated immunopathology in the context of Candida infection is absent. Here we describe a method to establish Th17 immunopathology associated with oral Candida infection in immunodeficient mice.
Modifying the Bank Erosion Hazard Index (BEHI) Protocol for Rapid Assessment of Streambank Erosion in Northeastern Ohio Sara E. Newton1, Deanna M. Drenten2 1Natural Resources, Cleveland Metroparks, 2Department of Biology, Case Western Reserve University Streambank erosion potential can be evaluated and ranked using David Rosgen's Bank Erosion Hazard Index (BEHI), however this protocol has significant limitations. Here we present protocol modifications to address time constraints, allow nonprofessionals to complete accurate assessments, and account for non-alluvial stream conditions in Northeast Ohio.
Floral-Dip Transformation of Flax (Linum usitatissimum) to Generate Transgenic Progenies with a High Transformation Rate Nasmah K. Bastaki1, Christopher A. Cullis1 1Department of Biology, Case Western Reserve University Here, we present a protocol to transform flax using Agrobacterium-mediated plant transformation via floral-dip. This protocol is simple to perform and inexpensive, yet yields a higher transformation rate than the current available methods for flax transformation.
Intravital Imaging of Axonal Interactions with Microglia and Macrophages in a Mouse Dorsal Column Crush Injury Teresa A. Evans1, Deborah S. Barkauskas2, Jay T. Myers3, Alex Y. Huang3 1Department of Neurosciences, Case Western Reserve University, 2Department of Biomedical Engineering, Case Western Reserve University, 3Department of Pediatrics, Case Western Reserve University Two-photon intravital imaging can be used to investigate interactions among different cell types in the spinal cord in their native tissue environment in a bone marrow chimeric animal with a dorsal column traumatic spinal cord crush injury.
Extracellular Wire Tetrode Recording in Brain of Freely Walking Insects Peiyuan Guo1, Alan J. Pollack1, Adrienn G. Varga1, Joshua P. Martin1, Roy E. Ritzmann1 1Department of Biology, Case Western Reserve University We previously developed a technique for implanting tetrode wires into the central complex of cockroach brains that allows us to monitor activity in individual units of tethered cockroaches. Here we present a modified version of that technique that allows us to also record brain activity in freely moving insects.
Experimental Protocol for Manipulating Plant-induced Soil Heterogeneity Angela J. Brandt1, Gaston A. del Pino1, Jean H. Burns1 1Department of Biology, Case Western Reserve University Understanding the role of environmental heterogeneity in species coexistence has typically focused on types of heterogeneity that are extrinsic to the community's species composition. We provide novel detailed methods for creating soil heterogeneity treatments using soils subject to plant-soil feedback conditioning, or heterogeneity intrinsic to the community composition.
MR Molecular Imaging of Prostate Cancer with a Small Molecular CLT1 Peptide Targeted Contrast Agent Xueming Wu1, Daniel Lindner2, Guan-Ping Yu1, Susann Brady-Kalnay3, Zheng-Rong Lu1 1Department of Biomedical Engineering, Case Western Reserve University, 2Department of Translational Hematology & Oncology Research, Cleveland Clinic, Case Western Reserve University, 3Department of Molecular Biology & Microbiology, Case Western Reserve University To demonstrate MR cancer molecular imaging with a small peptide targeted MRI contrast agent specific to clotted plasma proteins in tumor stroma in a mouse prostate cancer model.
Environmentally-controlled Microtensile Testing of Mechanically-adaptive Polymer Nanocomposites for ex vivo Characterization Allison E. Hess1, Kelsey A. Potter1,2, Dustin J. Tyler1,2, Christian A. Zorman1,3, Jeffrey R. Capadona1,2 1Advanced Platform Technology Center, Rehabilitation Research and Development, Louis Stokes Cleveland Department of Veterans Affairs Medical Center, 2Department of Biomedical Engineering, Case Western Reserve University, 3Department of Electrical Engineering and Computer Science, Case Western Reserve University A method is discussed by which the in vivo mechanical behavior of stimuli-responsive materials is monitored as a function of time. Samples are tested ex vivo using a microtensile tester with environmental controls to simulate the physiological environment. This work further promotes understanding the in vivo behavior of our material.
Extracellularly Identifying Motor Neurons for a Muscle Motor Pool in Aplysia californica Hui Lu1, Jeffrey M. McManus1, Hillel J. Chiel1,2,3 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.
An In Vitro Preparation for Eliciting and Recording Feeding Motor Programs with Physiological Movements in Aplysia californica Jeffrey M. McManus1, Hui Lu1, Hillel J. Chiel1,2,3 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.
Viral Nanoparticles for In vivo Tumor Imaging Amy M. Wen1, Karin L. Lee1, Ibrahim Yildiz1, Michael A. Bruckman1, Sourabh Shukla1, Nicole F. Steinmetz2 1Department of Biomedical Engineering, Case Western Reserve University, 2Department of Biomedical Engineering, Radiology, and Materials Science and Engineering, Case Western Reserve University Plant viral nanoparticles (VNPs) are promising platforms for applications in biomedicine. Here, we describe the procedures for plant VNP propagation, purification, characterization, and bioconjugation. Finally, we show the application of VNPs for tumor homing and imaging using a mouse xenograft model and fluorescence imaging.
Isolation of Soluble and Insoluble PrP Oligomers in the Normal Human Brain Xiangzhu Xiao1, Jue Yuan1, Wen-Quan Zou1,2 1Department of Pathology, National Prion Disease Pathology Surveillance Center, Case Western Reserve University School of Medicine, 2Department of Neurology, National Prion Disease Pathology Surveillance Center, Case Western Reserve University School of Medicine A new species of cellular prion protein (PrPC) has recently been identified in uninfected human brains using the methods described here. These methods can be used to isolate various PrP species, while some of them are also useful in isolating other misfolded protein aggregates from human brains.
Flow Cytometry-based Purification of S. cerevisiae Zygotes Serendipity Zapanta Rinonos1,2, Jeremy Saks1, Jonida Toska3, Chun-Lun Ni1,2, Alan M. Tartakoff1,2 1Department of Pathology, Case Western Reserve University School of Medicine, 2Cell Biology Program, Case Western Reserve University School of Medicine, 3Case Comprehensive Cancer Center, Case Western Reserve University School of Medicine To purify zygotes of S. cerevisiae, haploid cells of opposite mating type were engineered to express red or green fluorescent proteins, co-incubated to allow zygote formation, and fractionated using a flow cytometry-based protocol. The highly-enriched fraction enables subsequent "-omic" studies, recovery of initial progeny, and systematic investigation of zygote morphogenesis.
Stereotactic Radiosurgery for Gynecologic Cancer Charles Kunos1, James M. Brindle1, Robert Debernardo2 1Department of Radiation Oncology, University Hospitals Case Medical Center and Case Western Reserve University School of Medicine, 2Department of Obstetrics and Gynecology, Division of Gynecologic Oncology, University Hospitals Case Medical Center and Case Western Reserve University School of Medicine Stereotactic body radiotherapy (SBRT) involves image-guided, ablative radiation delivered to cancer targets refractory to chemotherapy or to conventional radiation treatment. The robotic-armed Cyberknife SBRT system, using sophisticated target localization, delivers hypofractionated radiation doses capable of sterilizing cancer targets. This article will consider new therapeutic roles of SBRT for gynecological cancers.
Intravital Imaging of the Mouse Popliteal Lymph Node H. L. Rachel Liou1, Jay T. Myers1, Deborah S. Barkauskas1, Alex Y. Huang2 1Department of Pediatrics, Case Western Reserve University, 2Department of Pediatrics, Pathology and Biomedical Engineering, Case Western Reserve University Recent advances in 2-photon microscopy have enabled real-time in situ imaging of live tissues in animal models, thereby enhancing our ability to investigate cellular behavior in both physiologic and pathologic conditions. Here, we outline the preparations required to perform intravital imaging of the mouse popliteal lymph node.
In vivo Dual Substrate Bioluminescent Imaging Michael K. Wendt1, Joseph Molter1, Christopher A. Flask1, William P. Schiemann1 1Case Comprehensive Cancer Center, Case Western Reserve University Herein we describe the methods to construct, visualize, and quantify the bioluminescent reactions of both firefly and renilla luciferase enzymes expressed in metastatic breast cancer cells during their growth and metastasis in vivo.
Cercarial Transformation and in vitro Cultivation of Schistosoma mansoni Schistosomules John N. Milligan1, Emmitt R. Jolly1 1Department of Biology, Case Western Reserve University We describe an in vitro method for culturing schistosomula of the flatworm parasite Schistosoma mansoni, via the harvesting and transformation of infective cercariae from the fresh water snail intermediate host Biomphalaria glabrata.
Reverse Total Shoulder Arthroplasty Christopher J. Lenarz1, Reuben Gobezie1 1Department of Orthopaedic Surgery – Division of Shoulder and Elbow Surgery, Case Western Reserve University Reverse total shoulder arthroplasty is indicated for the treatment of conditions that cannot be treated with conventional arthroplasty or other procedures. These primarily include degenerative and incapacitating conditions with irreparable rotator cuff and loss of the normal biomechanical coupling of the shoulder.
Environmentally Induced Heritable Changes in Flax Cory Johnson1, Tiffanie Moss1, Christopher Cullis1 1Department of Biology, Case Western Reserve University Growing some flax varieties under nutrient stress results in genomic variation within a subset of the genome and phenotypic variation. A complex insertion at a specific site is associated with growth under various nutrient regimes and with changes in gene expression around this site.
Upright Imaging of Drosophila Embryos Mirela Belu1, Michelle Javier1, Katayoun Ayasoufi1, Sarah Frischmann1, Catherine Jin1, Kuo-Chen Wang1, Rui Sousa-Neves1, Claudia Mieko Mizutani1,2 1Department of Biology, Case Western Reserve University, 2Department of Genetics, Case Western Reserve University Here we present a mounting protocol for stained Drosophila embryos in an upright position that allows imaging of cross-sections using Confocal microscopy.
Electrode Fabrication and Implantation in Aplysia californica for Multi-channel Neural and Muscular Recordings in Intact, Freely Behaving Animals Miranda J. Cullins1, Hillel J. Chiel1,2,3 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.
Surgical Induction of Endolymphatic Hydrops by Obliteration of the Endolymphatic Duct Cliff A. Megerian1, Chris Heddon1, Sami Melki1, Suhael Momin1, Janis Paulsey1, Joy Obokhare1, Kumar Alagramam1 1Otolaryngology - Head and Neck Surgery, Case Western Reserve University This video shows how to surgically obstruct the guinea pig's endolymphatic duct to produce endolymphatic hydrops.