Current Protocols in Neuroscience

Table of Contents

• Foreword
• Preface
• Chapter 1 Neuroanatomical Methods
  • Introduction
  • UNIT 1.1 Basic Neuroanatomical Methods
  • UNIT 1.2 Immunohistochemical Localization of Proteins in the Nervous System
  • UNIT 1.3 Hybridization Histochemistry of Neural Transcripts
  • UNIT 1.4 Receptor Binding Techniques
  • UNIT 1.5 Transneuronal Circuit Analysis With Pseudorabies Viruses
  • UNIT 1.6 Metabolic Mapping
  • UNIT 1.7 Targeted Toxins
  • UNIT 1.8 Cellular Compartment Analysis of Temporal Activity by Fluorescence In Situ Hybridization (catFISH)
  • UNIT 1.9 Detection of DNA Damage in Tissue Sections by In Situ Nick Translation
  • UNIT 1.10 Quantitative In Situ Hybridization for the Study of Gene Expression at the Regional and Cellular Levels
  • UNIT 1.11 Counting Cells in Sectioned Material: A Suite of Techniques, Tools, and Tips
  • UNIT 1.12 Intracellular and Juxtacellular Staining with Biocytin
  • UNIT 1.13 Quantified Assessment of Terminal Density and Innervation
  • UNIT 1.14 Anterograde Axonal Tract Tracing
  • UNIT 1.15 Histochemical Methods for the Detection of Apoptosis in the Nervous System
  • UNIT 1.16 Biochemical Fractionation of Brain Tissue for Studies of Receptor Distribution and Trafficking
  • UNIT 1.17 Retrograde Axonal Tracing with Fluorescent Markers
  • UNIT 1.18 Evolution of Neurotoxins: From Research Modalities to Clinical Realities
  • UNIT 1.19 Mapping Lineage Using BAC‐Cre Reporter Lines
• Chapter 2 Imaging
  • Introduction
  • UNIT 2.1 Fluorescence Microscopy: A Concise Guide to Current Imaging Methods
  • UNIT 2.2 Basic Confocal Microscopy
  • UNIT 2.3 Imaging Nervous System Activity
  • UNIT 2.4 Electronic Imaging in Neuroscience
  • UNIT 2.5 Loading Neurons with Dextran‐Conjugated Calcium Indicators in Intact Nervous Tissue
  • UNIT 2.6 Use of FM1‐43 and Other Derivatives to Investigate Neuronal Function
  • UNIT 2.7 Multidisciplinary Approaches for Characterizing Synaptic Vesicle Proteins
  • UNIT  2.8 3‐D Reconstruction of Neurons from Multichannel Confocal Laser Scanning Image Series
  • UNIT 2.9 Two‐Photon Imaging in Live Rodents
  • UNIT 2.10 Whole‐Mount Immunohistochemistry of the Brain
  • UNIT 2.11 Ballistic Labeling with Fluorescent Dyes and Indicators
  • UNIT 2.12 The Importance of Titrating Antibodies for Immunocytochemical Methods
• Chapter 3 Cellular and Developmental Neuroscience
  • Introduction
  • UNIT 3.1 Culture of Neuroepithelial Stem Cells
  • UNIT 3.2 Long‐Term Culture of Hippocampal Neurons
  • UNIT 3.3 Culture of Substantia Nigra Neurons
  • UNIT 3.4 Isolation and Purification of Primary Oligodendrocyte Precursors
  • UNIT 3.5 Isolation and Purification of Primary Rodent Astrocytes
  • UNIT 3.6 Differentiation of Embryonic Stem Cells
  • UNIT 3.7 Immortalizing Central Nervous System Cells
  • UNIT 3.8 Identification of Neural Programmed Cell Death Through the Detection of DNA Fragmentation In Situ and by PCR
  • UNIT 3.9 Quantitative Analysis of In Vivo Cell Proliferation
  • UNIT 3.10 Neural Stem Cell Transplantation in Mouse Brain
  • UNIT 3.11 Calcium Phosphate Transfection of DNA into Neurons in Primary Culture
  • UNIT 3.12 Purification of Oligodendrocytes and Their Progenitors Using Immunomagnetic Separation and Percoll Gradient Centrifugation
  • UNIT 3.13 Isolation and Generation of Oligodendrocytes by Immunopanning
  • UNIT 3.14 Generation and Utilization of Phosphorylation State–Specific Antibodies to Investigate Signaling Pathways
  • UNIT 3.15 Neuronal Transfection Using Particle‐Mediated Gene Transfer
  • UNIT 3.16 Gene Expression in Transgenic Mice Using Neural Promoters
  • UNIT 3.17 Isolation and Purification of Primary Schwann Cells
  • UNIT 3.18 Fluorescence‐Based Sorting of Neural Stem Cells and Progenitors
  • UNIT 3.19 Rodent Sensory Neuron Culture and Analysis
  • UNIT 3.20 Dissection and Culture of Embryonic Spinal Commissural Neurons
  • UNIT 3.21 Culture of Postnatal Mesencephalic Dopamine Neurons on an Astrocyte Monolayer
• Chapter 4 Gene Cloning, Expression, and Mutagenesis
  • Introduction
  • UNIT 4.1 Homology Cloning of cDNA or Genomic DNA
  • UNIT 4.2 PCR Cloning of Neural Gene Products
  • UNIT 4.3 Expression Cloning of Neural Genes Using Xenopus laevis Oocytes
  • UNIT 4.4 Interaction Trap/Two‐Hybrid System to Identify Interacting Proteins
  • UNIT 4.5 Overview of Neural Gene Expression
  • UNIT 4.6 Selection of Transfected Mammalian Cells
  • UNIT 4.7 Transient Expression of Proteins Using COS Cells
  • UNIT 4.8 cDNA Expression Cloning in Mammalian Cells
  • UNIT 4.9 Mutagenesis Approaches for Elucidation of Protein Structure‐Function Relationships
  • UNIT 4.10 Oligonucleotide‐Directed Mutagenesis Without Phenotypic Selection
  • UNIT 4.11 Directed Mutagenesis Using the Polymerase Chain Reaction
  • UNIT 4.12 Overview of Gene Delivery into Cells Using HSV‐1‐Based Vectors
  • UNIT 4.13 Generation of High‐Titer Defective HSV‐1 Vectors
  • UNIT 4.14 Gene Delivery Using Helper Virus–Free HSV‐1 Amplicon Vectors
  • UNIT 4.15 The Substituted‐Cysteine Accessibility Method (SCAM) to Elucidate Membrane Protein Structure
  • UNIT 4.16 Protein Expression in the Drosophila Schneider 2 Cell System
  • UNIT 4.17 Production of Recombinant Adeno‐Associated Viral Vectors and Use for In Vitro and In Vivo Administration
  • UNIT 4.18 Overview of the Baculovirus Expression System
  • UNIT 4.19 Protein Expression in the Baculovirus System
  • UNIT 4.20 Calcium Flux Assay in Xenopus Oocytes
  • UNIT 4.21 Production and Titration of Lentiviral Vectors
  • UNIT 4.22 Alphaviruses: Semliki Forest Virus and Sindbis Virus Vectors for Gene Transfer into Neurons
  • UNIT 4.23 Gene Transfer into Neural Cells In Vitro Using Adenoviral Vectors
  • UNIT 4.24 Gene Transfer into Rat Brain Using Adenoviral Vectors
  • UNIT 4.25 Overview of Nucleic Acid Arrays
  • UNIT 4.26 Preparation of mRNA for Expression Monitoring
  • UNIT 4.27 cDNA Library Construction from Single Cells
  • UNIT 4.28 Gene Expression Analysis of Neural Cells and Tissues Using DNA Microarrays
  • UNIT 4.29 Overview of Gene Targeting by Homologous Recombination
  • UNIT 4.30 Production of a Heterozygous Mutant Cell Line by Homologous Recombination (Single Knockout)
  • UNIT 4.31 Conditional Gene Expression and Targeting in Neuroscience Research
  • UNIT 4.32 Transfection of Cultured Primary Neurons via Nucleofection
  • UNIT 4.33 Generation of Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) Using Directed Molecular Evolution
• Chapter 5 Molecular Neuroscience
  • Introduction
  • UNIT 5.1 RNA Analysis by Nuclease Protection
  • UNIT 5.2 Differential Display of mRNA by PCR
  • UNIT 5.3 Analysis of mRNA Populations from Single Live and Fixed Cells of the Central Nervous System
  • UNIT 5.4 Reducing Gene Expression in the Brain via Antisense Methods
  • UNIT 5.5 Production of Polyclonal Antisera
  • UNIT 5.6 Production of Antipeptide Antisera
  • UNIT 5.7 Production of Antisera Using Fusion Proteins
  • UNIT 5.8 Epitope Tagging of Recombinant Proteins
  • UNIT 5.9 Overview of Membrane Protein Solubilization
  • UNIT 5.10 Metal‐Chelate Affinity Chromatography
  • UNIT 5.11 Immunoaffinity Chromatography
  • UNIT 5.12 The Use of Phage Display in Neurobiology
  • UNIT 5.13 Fluorescent In Situ Transcription in Cells and Tissues
  • UNIT 5.14 Detection of Protein Phosphorylation in Tissues and Cells
  • UNIT 5.15 Use of the A. Victoria Green Fluorescent Protein to Study Protein Dynamics In Vivo
  • UNIT 5.16 Green Fluorescent Protein in the Study of Neuronal Signaling Pathways
  • UNIT 5.17 Analysis of RNA by Northern and Slot‐Blot Hybridization
  • UNIT 5.18 Using Phage Display in Neurobiology
  • UNIT 5.19 Immunoblotting and Immunodetection
  • UNIT 5.20 An Overview on the Generation of BAC Transgenic Mice for Neuroscience Research
  • UNIT 5.21 Modification of Bacterial Artificial Chromosomes (BACs) and Preparation of Intact BAC DNA for Generation of Transgenic Mice
  • UNIT 5.22 Imaging Protein‐Protein Interactions by Fluorescence Resonance Energy Transfer (FRET) Microscopy
  • UNIT 5.23 Monitoring Protein‐Protein Interactions in Living Cells by Bioluminescence Resonance Energy Transfer (BRET)
  • UNIT 5.24 Immunoprecipitation
  • UNIT 5.25 Detection of Protein‐Protein Interactions by Coprecipitation
  • UNIT 5.26 Lentiviral Strategies for RNAi Knockdown of Neuronal Genes
  • UNIT 5.27 Split‐Ubiquitin System for Identifying Protein‐Protein Interactions in Membrane and Full‐Length Proteins
  • UNIT 5.28 Identifying Novel Protein‐Protein Interactions Using Co‐Immunoprecipitation and Mass Spectroscopy
• Chapter 6 Neurophysiology
  • Introduction
  • UNIT 6.1 The Electrophysiology Setup
  • UNIT 6.2 Principles of Signal Conditioning
  • UNIT 6.3 Fabrication of Patch Pipets
  • UNIT 6.4 Preparation of Hippocampal Brain Slices
  • UNIT 6.5 Acute Isolation of Neurons from the Mature Mammalian Central Nervous System
  • UNIT 6.6 Whole‐Cell Voltage Clamp Recording
  • UNIT 6.7 Patch‐Pipet Recording in Brain Slices
  • UNIT 6.8 Single‐Channel Recording
  • UNIT 6.9 Rapid Solution Application Methods
  • UNIT 6.10 Recording and Analyzing Synaptic Currents and Synaptic Potentials
  • UNIT 6.11 Preparation and Maintenance of Organotypic Slice Cultures of CNS Tissue
  • UNIT 6.12 Voltage Clamp Recordings from Xenopus Oocytes
  • UNIT 6.13 Synaptic Plasticity in the Hippocampal Slice Preparation
  • UNIT 6.14 Amperometry and Cyclic Voltammetry with Carbon Fiber Microelectrodes at Single Cells
  • UNIT 6.15 Preparation and Maintenance of Organotypic Cultures for Multi‐Electrode Array Recordings
  • UNIT 6.16 Chronic Recording of Extracellular Neuronal Activity in Behaving Animals
  • UNIT 6.17 Imaging Nervous System Activity with Voltage‐Sensitive Dyes
  • UNIT 6.18 Recording in the Cerebellar Slice
  • UNIT 6.19 Patch‐Clamp Recording from Neuronal Dendrites
  • UNIT 6.20 Caged Molecules: Principles and Practical Considerations
  • UNIT 6.21 Photolysis of Caged Neurotransmitters: Theory and Procedures for Light Delivery
  • UNIT 6.22 Whole‐Cell Recording In Vivo
  • UNIT 6.23 Electrophysiological Recordings from Neonatal Neocortical Brain Slices
  • UNIT 6.24 EEG Recording in Rodents, with a Focus on Epilepsy
• Chapter 7 Neurochemistry/Neuropharmacology
  • Introduction
  • UNIT 7.1 Overview of Brain Microdialysis
  • UNIT 7.2 Microdialysis in Rodents
  • UNIT 7.3 Microdialysis in Nonhuman Primates
  • UNIT 7.4 Detection and Quantification of Neurotransmitters in Dialysates
  • UNIT 7.5 Analyzing Radioligand Binding Data
  • UNIT 7.6 Saturation Assays of Radioligand Binding to Receptors and Their Allosteric Modulatory Sites
  • UNIT 7.7 Saturation Analysis of Ligand Binding Using a Centrifugation Procedure
  • UNIT 7.8 Ligand Characterization Using Microphysiometry
  • UNIT 7.9 Uptake and Release of Neurotransmitters
  • UNIT 7.10 Measurement of Chloride Movement in Neuronal Preparations
  • UNIT 7.11 Measurement of Cation Movement in Primary Cultures Using Fluorescent Dyes
  • UNIT 7.12 Measurement of Second Messengers in Signal Transduction: cAMP and Inositol Phosphates
  • UNIT 7.13 Measurement of NO and NO Synthase
  • UNIT 7.14 Measurement of Nitric Oxide in Single Cells and Tissue Using a Porphyrinic Microsensor
  • UNIT 7.15 Scintillation Proximity Assay
  • UNIT 7.16 Peripheral Type Benzodiazepine Binding Sites as a Tool for the Detection and Quantification of CNS Injury
  • UNIT 7.17 Measurement of Oxygen Radicals and Lipid Peroxidation in Neural Tissues
  • UNIT 7.18 Assessment of Cell Viability in Primary Neuronal Cultures
  • UNIT 7.19 In Vivo Measurement of Blood‐Brain Barrier Permeability
  • UNIT 7.20 Methods for Sample Preparation for Direct Immunoassay Measurement of Analytes in Tissue Homogenates: ELISA Assay of Amyloid β‐Peptides
  • UNIT 7.21 Assays for Cyclic Nucleotide‐Specific Phosphodiesterases (PDEs) in the Central Nervous System (PDE1, PDE2, PDE4, and PDE10)
• Chapter 8 Behavioral Neuroscience
  • Introduction
  • UNIT 8.1 Locomotor Behavior
  • UNIT 8.2 Sexual and Reproductive Behaviors
  • UNIT 8.3 Animal Tests of Anxiety
  • UNIT 8.4 Application of Experimental Stressors in Laboratory Rodents
  • UNIT 8.5A Assessment of Spatial Memory Using the Radial Arm Maze and Morris Water Maze
  • UNIT 8.5B Assessment of Spatial Memory Using the T Maze
  • UNIT 8.5C Cued and Contextual Fear Conditioning in Mice
  • UNIT 8.5D Social Learning of Food Preferences in Rodents: Rapid Appetitive Learning
  • UNIT 8.5E Assessment of Sustained and Divided Attention in Rats
  • UNIT 8.5F Assessment of Learning and Memory Using the Autoshaping of Operant Responding in Mice
  • UNIT 8.5G Social Transmission of Food Preference in Mice
  • UNIT 8.5H Assaying Aspects of Attention and Impulse Control in Mice Using the 5‐Choice Serial Reaction Time Task
  • UNIT 8.6A Overview of Methodological Approaches to the Study of Ingestive Behavior
  • UNIT 8.6B Basic Measures of Food Intake
  • UNIT 8.6C Direct, Continuous Behavioral Analysis of Drug Action on Feeding
  • UNIT 8.6D Sham Feeding in Rats with Chronic, Reversible Gastric Fistulas
  • UNIT 8.6E Conditioned Flavor Aversions: Assessment of Drug‐Induced Suppression of Food Intake
  • UNIT 8.6F Conditioned Flavor Preferences: Evaluating Postingestive Reinforcement by Nutrients
  • UNIT 8.6G Macronutrient Selection in Experimental Animals
  • UNIT 8.7 Measurement of Startle Response, Prepulse Inhibition, and Habituation
  • UNIT 8.8 Measurement of Rodent Stereotyped Behavior
  • UNIT 8.9 Models of Nociception: Hot‐Plate, Tail‐Flick, and Formalin Tests in Rodents
  • UNIT 8.10A Rodent Models of Depression: Forced Swimming and Tail Suspension Behavioral Despair Tests in Rats and Mice
  • UNIT 8.10B Rodent Models of Depression: Learned Helplessness Using a Triadic Design in Rats
  • UNIT 8.10C Rodent Models of Depression: Learned Helplessness Induced in Mice
  • UNIT 8.11A Fear‐Potentiated Startle in Rats
  • UNIT 8.11B Fear‐Potentiated Startle in Mice
  • UNIT 8.12 Motor Coordination and Balance in Rodents
  • UNIT 8.13 Latent Inhibition
  • UNIT 8.14 Ultrasonic Vocalizations in Rat and Mouse Pups
  • UNIT 8.15 Parental Behaviors in Rats and Mice
  • UNIT 8.16 Mouse Social Recognition and Preference
  • UNIT 8.17 Assessment of Murine Startle Reactivity, Prepulse Inhibition, and Habituation
  • UNIT 8.18 Assessment of Developmental Milestones in Rodents
  • UNIT 8.19 Defensive Responses to Predator Threat in the Rat and Mouse
  • UNIT 8.20 Olfactometry with Mice
  • UNIT 8.21A Overview of Methods for Assessing the Mouse Auditory System
  • UNIT 8.21B Measurement of the Auditory Brainstem Response (ABR) to Study Auditory Sensitivity in Mice
  • UNIT 8.21C Assessment of Cochlear Function in Mice: Distortion‐Product Otoacoustic Emissions
  • UNIT 8.21D Behavioral Assessment of Hearing in Mice—Conditioned Suppression
  • UNIT 8.22 Delay Discounting and Impulsive Choice in the Rat
  • UNIT 8.23 Fear Extinction in Rodents
  • UNIT 8.24 Simple Behavioral Assessment of Mouse Olfaction
• Chapter 9 Preclinical Models of Neurologic and Psychiatric Disorders
  • Introduction
  • UNIT 9.1 Dural Inflammation Model of Migraine Pain
  • UNIT 9.2 Traumatic Brain Injury in the Rat Using the Fluid‐Percussion Model
  • UNIT 9.3 Preclinical Models of Hepatic Encephalopathy
  • UNIT 9.4 Preclinical Models of Parkinson's Disease
  • UNIT 9.5 Rodent Models of Global Cerebral Ischemia
  • UNIT 9.6 Rodent Models of Focal Cerebral Ischemia
  • UNIT 9.7 Experimental Autoimmune Encephalomyelitis (EAE)
  • UNIT 9.8 A Murine Model of Retroviral Encephalopathy
  • UNIT 9.9 Animal Models of Retroviral Encephalopathies: Feline Model
  • UNIT 9.10 Overview of Rodent Models for Obesity Research
  • UNIT 9.11 Intravenous Self‐Administration of Ethanol in Mice
  • UNIT 9.12 Preclinical Models to Evaluate Potential Pharmacotherapeutic Agents in Treating Alcoholism and Studying the Neuropharmacological Bases of Ethanol‐Seeking Behaviors in Rats
  • UNIT 9.13 Models of Amyotrophic Lateral Sclerosis
  • UNIT 9.14 Models of Neuropathic Pain in the Rat
  • UNIT 9.15 Place Preference Test in Rodents
  • UNIT 9.16 Inducing Photochemical Cortical Lesions in Rat Brain
  • UNIT 9.17 Measurement of Panic‐Like Responses Following Intravenous Infusion of Sodium Lactate in Panic‐Prone Rats
  • UNIT 9.18 Animal Models of Painful Diabetic Neuropathy: The STZ Rat Model
  • UNIT 9.19 Chemoconvulsant Model of Chronic Spontaneous Seizures
  • UNIT 9.20 Chronic Intravenous Drug Self‐Administration in Rats and Mice
  • UNIT 9.21 Intravenous Self‐Administration Techniques in Monkeys
  • UNIT 9.22 Overview on Rodent Models of Alzheimer's Disease
  • UNIT 9.23A Binge Eating in Rats Produced by Combining Dieting with Stress
  • UNIT 9.23B Binge Eating in Rats with Limited Access to Vegetable Shortening
  • UNIT 9.23C Sugar Bingeing in Rats
  • UNIT 9.23D Characterization of Energy Expenditure in Rodents by Indirect Calorimetry
  • UNIT 9.24 Overview of Animal Models of Schizophrenia
  • UNIT 9.25 Ratings of L‐DOPA‐Induced Dyskinesia in the Unilateral 6‐OHDA Lesion Model of Parkinson's Disease in Rats and Mice
  • UNIT 9.26 Overview of Mouse Assays of Ethanol Intoxication
  • UNIT 9.27 Schedule‐Induced Polydipsia: A Rat Model of Obsessive‐Compulsive Disorder
  • UNIT 9.28 The Majchrowicz Binge Alcohol Protocol: An Intubation Technique to Study Alcohol Dependence in Rats
  • UNIT 9.29 Vapor Inhalation of Alcohol in Rats
  • UNIT 9.30 Animal Models of Obsessive‐Compulsive Disorder
  • UNIT 9.31 An Animal Model of Premenstrual Dysphoric Disorder Sensitive to Antidepressants
  • UNIT 9.32 Recapitulation and Reversal of a Persistent Depression‐like Syndrome in Rodents
• Chapter 10 Human Neuroscience
  • Introduction
  • UNIT 10.1 Assessment of Parkinson Disease Manifestations
  • UNIT 10.2 EEG Recording and Analysis for Sleep Research
  • UNIT 10.3 Neuropsychological Assessment in the Diagnosis of Alzheimer's Disease
• Appendix 1 Molecular Biology Techniques
  • APPENDIX 1B Optimization of Transfection
  • APPENDIX 1C Calcium Phosphate Transfection
  • APPENDIX 1D Transfection Using DEAE‐Dextran
  • APPENDIX 1E Transfection by Electroporation
  • APPENDIX 1F Transfection of Cultured Eukaryotic Cells Using Cationic Lipid Reagents
  • APPENDIX 1G Purification and Concentration of DNA from Aqueous Solutions
  • APPENDIX 1H Preparation of Genomic DNA from Mammalian Tissue
  • APPENDIX 1I Preparation of RNA from Tissues and Cells
  • APPENDIX 1J Preparation of Bacterial Plasmid DNA
  • APPENDIX 1K Quantitation of DNA and RNA with Absorption and Fluorescence Spectroscopy
  • APPENDIX 1L Introduction of Plasmid DNA into Cells
  • APPENDIX 1M Digestion of DNA with Restriction Endonucleases
  • APPENDIX 1N Agarose Gel Electrophoresis
• Appendix 2 Stock Solutions, Equipment, and Laboratory Guidelines
  • APPENDIX 2A Common Stock Solutions, Buffers, and Media
  • APPENDIX 2B Standard Laboratory Equipment
• Appendix 3 Commonly Used Techniques
  • APPENDIX 3A Preparation of an Iodinated Radioligand
  • APPENDIX 3B Techniques for Mammalian Cell Tissue Culture
• Appendix 4 Animal Techniques
  • APPENDIX 4A High Precision Stereotaxic Surgery in Mice
  • APPENDIX 4B Mouse and Rat Anesthesia and Analgesia
  • APPENDIX 4C Animal Health Assurance
  • APPENDIX 4D Handling and Restraint
  • APPENDIX 4E Animal Identification
  • APPENDIX 4F Parenteral Injections
  • APPENDIX 4G Blood Collection
  • APPENDIX 4H Euthanasia
  • APPENDIX 4I Assessing Reproductive Status/Stages in Mice
• Appendix 5 Databases
• Appendix Suppliers
  • Selected Suppliers of Reagents and Equipment

* Looking for a protocol from this title that does not appear here? You can find it on interscience.wiley.com