Main description:

Rapid advances in knowledge have led to an increasing interest in neuro biology over the last several years. These advances have been made possible, at least in part, by the use of increasingly sophisticated methodology. Furthermore, research in the most rapidly advancing areas is essentially multidisciplinary and is characterized by contributions from many investi gators employing a variety of techniques. While a grasp of fundamental neurobiological concepts is an obvious prerequisite for those who wish to follow or participate in this field, critical awareness and evaluation of neurobiological research also requires an understanding of sophisticated methodologies. The objective of Methods in Neurobiology is the development of such critical abilities. The reader is exposed to the basic concepts, principles, and instrumentation of key methodologies, and the application of each meth odology is placed in the special context of neurobiological research. The reader will gain familiarity with the terminology and procedures of each method and the ability to evaluate results in light of the particular features of neurobiological preparations and applications.

Contents:

1 Behavioral Techniques in Pharmacological and Neuropharmacological Analysis.- 1. Introduction.- 2. Basic Principles.- 2.1. Routes of Administration.- 2.2. Central vs. Peripheral Drug Effects.- 2.3. Dose-Response Relationships.- 2.4. Drug Receptors.- 2.5. Drug Effects on Neurotransmitter Activity.- 2.6. Transmitter Storage: Drug-Behavior Relationships.- 2.7. Transmitter Turnover.- 2.8. Drug Interactions-Effects of Chronic Depletion.- 2.9. Specificity of Drug Effects-Neurochemical Interactions.- 2.10. Drug Synergism.- 2.11. Species and Strain Factors.- 3. Locomotor Excitation and Inhibition.- 3.1. Vertical-Horizontal Activity.- 3.2. Response Excitation and Response Disinhibition.- 3.3. Activity-Reactivity.- 3.4. Stereotypy.- 3.5. Circling-Rotational Behavior.- 3.6. Perseveration.- 4. Habituation.- 4.1. Habituation and Sensitization.- 4.2. Temporal Evaluation.- 4.3. Spontaneous Alternation.- 4.4. Carry-Over Designs.- 4.5. Exploration and Startle.- 4.6. Summary.- 5. Appetitively Motivated Operant Behaviors.- 5.1. Operant-Appetitive Paradigms.- 5.2. Schedules of Reinforcement.- 5.3. Adjunctive Behavior.- 5.4. Observing Behavior.- 5.5. Drug Effects and the Experimental Analysis of Behavioral Processes.- 5.6. Summary.- 6. Aversively Motivated Behaviors.- 6.1. Components of Avoidance Behavior.- 6.2. Avoidance Techniques: Task Manipulations and Multiple-Testing Procedures.- 6.3. Discriminated Avoidance.- 6.4. Transfer Designs.- 6.5. Summary.- 7. Overview.- References.- 2 Behavioral and Biochemical Methods to Study Brain Responses to Environment and Experience.- 1. Introduction.- 2. Environmental and Training Techniques.- 2.1. Differential Environments.- 3. Behavioral Techniques to Test Effects of Prior Experience.- 3.1. Hebb-Williams Maze.- 4. Biochemical Techniques.- 4.1. General Considerations.- 4.2. Weights of Brain Regions.- 4.3. RNA and DNA Content.- 4.4. AChE and ChE Activities.- References.- 3 Cell Fractionation.- 1. Introduction.- 1.1. Brief History of Centrifugal Fractionation.- 2. Basic Information on Centrifugation.- 2.1. Centrifugal Force.- 2.2. Centrifugal Force and Sedimentation.- 3. Centrifuges.- 3.1. Determination of Relative Centrifugal Force.- 3.2. Analytical vs. Preparative Centrifugation.- 4. Homogenization.- 4.1. Coaxial Homogenizers.- 4.2. Blenders.- 4.3. Cellular Heterogeneity and Tissue Preparation.- 4.4. Homogenization Media.- 5. Differential Centrifugation.- 5.1. Normal-Rate Separation.- 5.2. Factors That Influence Sedimentation.- 5.3. Density Gradients.- 6. Analytical vs. Preparative Strategies.- 6.1. Analytical Approach.- 6.2. Validation of Fractionation Schemes.- 6.3. Balance Sheets.- References.- Selected Bibliography of Fractionation Literature.- Whole Cells: Neurons and Glia.- Nuclei.- Mitochondria.- Lysosomes.- Synaptosomes.- Plasma Membranes: Synaptic and Glial Membranes.- Synaptic Vesicles.- Endoplasmic Reticulum; Ribosomes, Polysomes.- Filaments, Tubules, and Other Neuronal Inclusions.- Myelin.- Soluble Phase.- 4 Polyacrylamide Gel Electrophoresis: Principles, Techniques, and Micromethods.- 1. Introduction.- 2. Basic Concepts.- 3. Outline of Electrophoretic Techniques.- 3.1. General Terminology.- 3.2. Moving-Boundary Technique.- 3.3. Zone Electrophoresis.- 3.4. Disk Electrophoresis.- 3.5. Isotachophoresis.- 3.6. Isoelectric Focusing.- 3.7. Two-Dimensional Techniques.- 3.8. Buffer Systems.- 3.9. Electrophoresis in Sodium Dodecyl Sulfate.- 4. Practical Aspects of Electrophoresis and Micromethods.- 4.1. The Case for Miniaturization.- 4.2. Zone Microelectrophoresis.- 4.3. Disk Microelectrophoresis.- 4.4. Gradient Microelectrophoresis.- 4.5. Microisoelectric Focusing.- 4.6. Two-Dimensional Microelectrophoresis.- 4.7. Microelectrophoresis in Sodium Dodecyl Sulfate.- 4.8. Microelectrophoresis of Nucleic Acids.- 4.9. Equipment for Microelectrophoresis.- 4.10. Recent Advances.- 5. Related Techniques and Analysis of Gel Patterns.- 5.1. Qualitative and Quantitative Analysis of Gel Patterns.- 5.2. Determination of Molecular Weight.- 5.3. Sources of Artifacts.- 6. Interfacing Electrophoresis with Other Techniques.- 6.1. Elution and in Situ Histochemistry.- 6.2. Immunological and Autoradiographic Techniques.- 6.3. Peptide Mapping and Amino Acid Analysis of Proteins Following Electrophoresis.- 7. Concluding Remarks.- References.- 5 Classic Methods in Neuroanatomy.- 1. Introduction.- 2. Dissection and Gross Anatomy of the Brain.- 3. Reconstruction Methods.- 4. Fixation and General Histological Procedure.- 4.1. Fixation.- 4.2. Frozen Sections.- 4.3. Paraffin Embedding.- 4.4. Celloidin and Plastic Embedding.- 5. Nissi Stains.- 5.1. Nissl's Original Method and Its Modifications.- 5.2. Cytoarchitectonics.- 5.3. Axonal Reaction of the Perikaryon.- 6. Myelin Stains.- 6.1. Staining of the Lipids in the Myelin Sheath.- 6.2. Mordanting of Myelin: Weigert and Haggqvist Methods.- 6.3. Kluver-Barrera Method.- 6.4. Myeloarchitecture: Counting and Measuring of Nerve Fibers.- 6.5. Myelogenesis, Myelin Degeneration, and Demyelinization.- 7. Reduced-Silver Methods.- 7.1. Neurofibrillar Stains.- 7.2. Silver Impregnation of Degenerating Axons and Axon Terminals.- 8. Golgi Method.- 9. Histological Methods.- 9.1. Gelatin Embedding.- 9.2. Paraffin Embedding.- 9.3. Celloidin Embedding.- 9.4. Nissl Stains.- 9.5. Myelin Stains.- 9.6. Reduced-Silver Methods.- 9.7. Golgi Methods.- References.- 6 Fluorescence Microscopy of Biogenic Monoamines.- 1. Introduction.- 2. Fluorescence Microscopy and Microspectrofluorometry.- 2.1. The Fluorescence Microscope.- 2.2. The Microspectrofluorometer.- 3. Visualization of Biogenic Monoamines.- 3.1. Chemical Background.- 3.2. Practical Performance of the Formaldehyde and Glyoxylic Acid Methods.- 4. Fluorescence-Histochemical Techniques for Identification and Differentiation of Biogenic Monoamines.- 4.1. Tests for Specificity.- 4.2. Identification and Differentiation of Biogenic Amines and Related Compounds after Formaldehyde Treatment.- 4.3. Identification and Differentiation of Biogenic Amines and Related Compounds after Glyoxylic Acid Treatment.- 5. General Conclusion.- References.- 7 Electron Microscopy in Neurobiology.- 1. Introduction.- 2. The Transmission Electron Microscope: Basic Principles and Design.- 2.1. Introduction.- 2.2. Resolving Power.- 2.3. Image Formation and Contrast.- 2.4. Depth of Field and Depth of Focus.- 2.5. Magnification.- 2.6. Design of the Transmission Electron Microscope.- 2.7. Special Applications.- 3. Electron-Microscopic Histology.- 3.1. Introduction.- 3.2. Fixation.- 3.3. Dehydration.- 3.4. Embedding.- 3.5. Ultramicrotomy.- 3.6. Staining Procedures.- 3.7. Other Techniques.- 3.8. Some Critical Remarks on Electron-Microscopic Histology.- 4. Applications of Electron Microscopy in Neurobiology.- 4.1. Ultrastructural Studies in Neuroanatomy and Neurocytology.- 4.2. Electron Microscopy and Synaptic Transmission.- 4.3. Electron Microscopy in Cell and Tissue Culture.- 4.4. Electron Microscopy and Cell Fractionation.- 4.5. Electron Microscopy: Relevance and Limitations in Neurobiological Research.- 5. Quantitative Stereology in Electron Microscopy.- 5.1. Introduction.- 5.2. Basic Principles and Terminology.- 5.3. Some Practical Aspects of Quantitative Stereology.- References.- 8 Scanning Electron Microscopy: Applications to Neurobiology.- 1. Fundamentals of Scanning Electron Microscopy.- 1.1. Introduction.- 1.2. History.- 1.3. The Instrument.- 1.4. The Sample.- 1.5. Advances in Techniques of Specimen Examination and Characterization.- 2. Specific Applications of Scanning Electron Microscopy to Neurobiology.- 2.1. Introduction.- 2.2. Central Nervous System.- 2.3. Peripheral Nervous System and Individual Neurons.- 2.4. Receptors.- 2.5. Conclusion.- References.- 9 Autoradiography in the Nervous System.- 1. Introduction.- 2. Application of Autoradiography in Different Fields of Neurobiological Research.- 2.1. Introduction.- 2.2. Cell Biology.- 2.3. Embryology.- 2.4. Neuronal Connectivity.- 3. Basic Principles of Autoradiography.- 3.1. Introduction.- 3.2. Radioisotopes.- 3.3. Photographic Emulsions.- 3.4. Resolution of Autoradiographs.- 3.5. Efficiency of Autoradiographs.- 4. Methods.- 4.1. Administration of Labeled Compounds.- 4.2. Preparation of Autoradiographs.- 4.3. Analysis of Autoradiographs.- 5. Critical Evaluation of Anterograde Tracing Techniques.- 5.1. Introduction.- 5.2. The Problem of Passing Fibers.- 5.3. Injection Site.- 5.4. Termination Area.- 5.5. Differential Uptake of Precursors.- 5.6. Sensitivity of the Autoradiographic Tracing Method.- 5.7. Transneuronal Transport.- 5.8. Some Technical Advantages and Limitations of the Autoradiographic Tracing Method.- 5.9. Combination of the Autoradiographic Tracing Method with Other Techniques.- References.- 10 Isotope Methods.- 1. Introduction.- 2. Physical Background.- 2.1. Properties of the Nucleus.- 2.2. Types of Radioactive Decay.- 2.3. Rate of Radioactive Decay.- 2.4. Units and Definitions.- 3. Isotopes of Neurobiological Interest.- 4. Labeled Compounds.- 4.1. Availability.- 4.2. Position of Label.- 4.3. Manufacturing of Labeled Compounds.- 4.4. Purity of Labeled Compounds.- 4.5. Stability and Storage of Labeled Compounds.- 5. Measurement of Isotopes.- 5.1. Stable Isotopes.- 5.2. Radioactive Isotopes.- 6. Statistics of Radioactivity Detection.- 7. Data Analysis in Neurobiological Applications.- 7.1. Assay of Brain Free Amino Acids with Labeled Dansyl Chloride.- 7.2. Radioimmunoassay for Myelin Basic Protein.- 7.3. Compartmental Analysis of Tracer Kinetics.- 7.4. Determination of the Blood-Brain Exchange of Taurine.- 7.5. Determination of the Rate Constants for Amino Acid Efflux from Brain Slices.- References.