Neuroscience Methods-A Guide for Advanced Students

273 Pages·1998·16.457 MB·\273

by  Rosemary Martin (Author)

NEUROSCIENCE METHODS

NEUROSCIENCE METHODS
A Guide for Advanced Students
Edited by
Rosemary Martin
Australian National University
Canberra, Australia
CRC Press
Taylor & Francis Group
Boca Raton London New York
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CONTENTS
Preface ix
Contributors xi
SECTION 1 IN VITRO PREPARATIONS
Chapter 1 Slices of brain tissue 3
C. Strieker
Chapter 2 The brain in a dish: How to study acutely
dissociated and cultured neurons 11
/. M. Bekkers
Chapter 3 Painting optimal bilayers for recording ion
channels 17
J. I. Kourie
SECTION 2 ELECTROPHYSIOLOGICAL TECHNIQUES
Chapter 4 Extracellular single neuronal recordings in the
whole animal 25
T. R. Vidyasagar
Chapter 5 Equipment for measurement of membrane
potentials and currents, with special reference to
sharp microelectrodes 33
R. L. Martin
Chapter 6 Sucking up to cells: The patch-clamp technique in
neuroscience 40
J. M. Bekkers
Chapter 7 Analysis of single channel recordings 46
D. Lover
V
VI CONTENTS
SECTION 3 HUMAN ELECTROPHYSIOLOGY
Chapter 8 Recording electromyographic activity (emg)
from single motor units in human subjects 55
V. G. Macefield
Chapter 9 Intraneural recording from single nerve fibers in
awake human subjects 60
V. G. Macefield
Chapter 10 Somatosensory cortical evoked potentials recorded
from human subjects 65
V. G. Macefield
SECTION 4 APPLICATION OF DRUGS
Chapter 11 Very fast drug application to dissociated cells and
membrane patches 73
J. Clements
Chapter 12 Microelectrophoresis and pressure ejection methods 80
G. Lacey
SECTION 5 MEASUREMENT OF ION CONCENTRATIONS
Chapter 13 Ion-selective microelectrodes 87
A. I. Cowan
Chapter 14 Fluorescent dyes for measurement of intracellular
ion concentrations 94
N. Mahanty
SECTION 6 IN VIVO TECHNIQUES AND PREPARATIONS
Chapter 15 Stereotaxic placement of probes in neurobiology 107
S. P. Perrett
Chapter 16 In vivo microdialysis 113
A. J. Lawrence
Chapter 17 Electroretinography 119
A. Leon
CONTENTS VII
SECTION 7 HISTOCHEMICAL TECHNIQUES
Chapter 18 Anatomical tracing of neuronal connections 127
L. R. Marotte
Chapter 19 Immunohistochemistry 131
P. Cooper
Chapter 20 High resolution autoradiography 137
S. Oleskevich
SECTION 8 BIOCHEMICAL TECHNIQUES
Chapter 21 Membrane fractionation 143
P. R. Junankar
Chapter 22 Chromatographic techniques for protein
separation and purification 149
G. M. de Plater
SECTION 9 PRODUCTION OF ANTIBODIES
Chapter 23 Production and uses of polyclonal antibodies 157
M. F. Crouch
Chapter 24 Production and uses of monoclonal antibodies 160
W. D. Phillips
SECTION 10 BLOTTING AND HYBRIDIZATION TECHNIQUES
Chapter 25 Immunoblotting (Western blotting) 169
5. Johanson
Chapter 26 Southern and Northern blotting 174
S. Howitt
Chapter 27 Membrane and in situ hybridization:
Principles and pitfalls 179
B. Key
VIII CONTENTS
SECTION 11 EXPRESSION SYSTEMS
Chapter 28 Cellular expression of cloned and mutated
ion channels 187
P. R. Schofield
Chapter 29 Transfection in cell cultures 192
W. D. Phillips
SECTION 12 GENETIC TECHNIQUES
Chapter 30 Reverse transcription polymerase chain reaction 203
M. Vidovic
Chapter 31 Differential display of gene expression 209
F. M. Freeman
Chapter 32 Inhibition of gene expression with antisense
oligonucleotides 215
P. M. Pilowsky
Chapter 33 Creation of transgenic mice 220
P. G. Noakes
SECTION 13 NEURAL GRAFTING
Chapter 34 Neural grafting 233
L. J. Reece
Glossary 239
Index 253
PREFACE
Scientists choose to study the brain in many different ways. Some prefer to work at the
molecular level, unravelling the contribution of variations in gene expression or intra-
cellular proteins to normal function or neuropathologies. Others are engaged in
understanding the electrical properties of neurons or the chemistry of the brain and still
others prefer to study the overt behavior of an organism and surmise about the design
of the 'black box' which produces such behavior.
Not surprisingly, a large number of very specialized techniques have been developed
to allow these avenues of study. Most undergraduate, honours and new PhD students find
the plethora of techniques bewildering. As a consequence they have trouble understand-
ing original papers and find it difficult to critically appraise them.
To help solve this problem, I asked a number of colleagues to each write a few pages
about a method they used to study the nervous system. I particularly requested that they
directed their writing towards an outline of the methodological principles and potential
pitfalls of the technique they chose to describe.
It is always difficult to know where to stop with a book such as this. In the main,
the techniques described are those used in animal experiments but I have chosen to
include a few chapters on electrophysiological measurements in humans. Descriptions
of some modern imaging techniques such as NMR, PET and SPECT and behavioral
methods will have to wait, perhaps for another edition.
This book is the outcome of the labors of my many colleagues and I thank them all
for so generously giving their time. I also wish to thank Garry Rhoda for his excellent
editing of diagrams and Audra Johnstone for preparing the manuscript in its final form.
IX