Auditory and Vestibular Research

This volume describes current RNA, protein, and imaging protocols that have provided insights into genetic regulation as well as a greater understanding of the genes and pathogens involved in diseases of the ear.

Hearing is a sensory modality critical to both language and cognitive development. In its absence, and without sensory input through another modality, such as the manual/visual modality of sign language, cognitive and language development can be severely impaired in the earliest formative years of a child. In its endeavor to d- cover the mechanisms underlying audition, the ?eld of auditory science has provided rich comparative physiological studies, allowing insights into both the micromecha- cal and electrochemical world of this system. For many years, the auditory/vestibular sciences have been in?uenced by the discoveries of electrical engineers and sensory physiologists, who have provided insights into the functions of this dynamic system. The early discoveries in these ?elds, as well as advancements in microprocessing and materials technologies, provided a means whereby hearing could be regained partly through the use of a bionic device, known as a cochlear implant. Presently, this device and the auditory brainstem implant are the only ones to prosthetically replace brain function. With the advent of molecular biology tools, such as RT-PCR, the auditory and vestibular ?elds have made great strides in understanding the genetic basis for various hearing and balance disorders over the past ?fteen to twenty years. These technologies permitted the discovery of genes that control inner ear structure and function by ov- coming the hurdle of working with small amounts of tissue, as found in the inner ear.

Lists all the reagents, step-by-step techniques, and limitations for reproducible results in DNA/RNA tracing, protein identification, and microscopic imaging Clearly describes the protocols necessary for in vivo and in vitro gene delivery techniques Provides the tools to identify and isolate cochlear stem cells for the treatment of inner ear pathologies Includes lineage and fate mapping techniques for peripheral and central studies of auditory/vestibular development Describes techniques for identification and verification of protein-protein interactions Provides important tools for the basic and clinical researcher to identify genes and microbes underlying diseases of the ear including the cloning of deafness genes Includes supplementary material: sn.pub/extras

Klappentext

While early studies of the auditory/vestibular sciences provided insights into the anatomy and neurophysiology of these systems and produced a prosthetic cochlear implant, the rise of molecular biology now permits a clear examination of the genetic basis for various hearing and balance disorders. In Auditory and Vestibular Research: Methods and Protocols, specialists in the field describe current RNA, protein, and imaging protocols that have provided insights into genetic regulation as well as a greater understanding of the genes and pathogens involved in diseases of the ear. This overview utilizes both mammalian and non-mammalian animal models as well as techniques applicable to clinical studies in order to best provide an up-to-date perspective on basic research. Written in the highly successful Methods in Molecular Biology™ series format, chapters include brief introductions to their subjects, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and Notes sections, highlighting tips on troubleshooting and avoiding known pitfalls.

Comprehensive and cutting-edge, Auditory and Vestibular Research: Methods and Protocols is an ideal guide to the field and a useful tool for exploring genes and proteins in other systems as well, especially systems in which tissues are scarce and a comparative approach lends itself to discovering the underlying causes of human disorders.

Inhalt
Nucleic Acid Protocols.- RNA Isolation from Xenopus Inner Ear Sensory Endorgans for Transcriptional Profiling and Molecular Cloning.- Synthesis of Biotin-Labeled RNA for Gene Expression Measurements Using Oligonucleotide Arrays.- In situ Hybridization Approach to Study mRNA Expression and Distribution in Cochlear Frozen Sections.- Lineage Analysis of Inner Ear Cells Using Genomic Tags for Clonal Identification.- Genetic Fate-Mapping Approaches: New Means to Explore the Embryonic Origins of the Cochlear Nucleus.- The Practical Use of Cre and loxP Technologies in Mouse Auditory Research.- Helios® Gene GunMediated Transfection of the Inner Ear Sensory Epithelium.- Electroporation-Mediated Gene Transfer to the Developing Mouse Inner Ear.- Isolation of Sphere-Forming Stem Cells from the Mouse Inner Ear.- Molecular Biology of Vestibular Schwannomas.- Multilocus Sequence Typing and Pulsed Field Gel Electrophoresis of Otitis Media Causing Pathogens.- Fluorescence In Situ Hybridization for the Detection of Biofilm in the Middle Ear and Upper Respiratory Tract Mucosa.- Positional Cloning of Deafness Genes.- Amino Acid Protocols.- Twist-Off Purification of Hair Bundles.- Yeast Two-Hybrid Screening to Test for ProteinProtein Interactions in the Auditory System.- The Use of 2-D Gels to Identify Novel ProteinProtein Interactions in the Cochlea.- Identification of Functionally Important Residues/Domains in Membrane Proteins Using an Evolutionary Approach Coupled with Systematic Mutational Analysis.- In Vivo Verification of Protein Interactions in the Inner Ear by Coimmunoprecipitation.- Identification of Transcription FactorDNA Interactions Using Chromatin Immunoprecipitation Assays.- Surface Plasmon Resonance (SPR) Analysis of Binding Interactions of Proteins inInner-Ear Sensory Epithelia.- Multiplexed Isobaric Tagging Protocols for Quantitative Mass Spectrometry Approaches to Auditory Research.- Imaging Protocols.- Fluorescence Microscopy Methods in the Study of Protein Structure and Function.- Ion Imaging in the Cochlear Hair Cells.- Atomic Force Microscopy in Studies of the Cochlea.