Introduction to Electrophysiological
Methods and Instrumentation
Franklin Bretschneider Department of Biology, Utrecht University, Utrecht, The Netherlands
Jan R. de Weille Institut des Neurosciences de Montpellier, Hôpital Saint Eloi,
Montpellier, France
All living cells use electricity for some function or another. Even bacteria communicate with each other through electricity and have been shown to elicit electrical spikes of millisecond duration, uncannily akin spiking by our own neurones. Mitochondria, which are thought to be of bacterial origin, use electricity to fuel the phosphorylation of ADP into the energy carrier ATP. Perhaps viruses are the sole creatures that are electrically inert, even if some viruses contain DNA coding for ion channels. The sensitive plant or touch-me-not (Mimosa pudica) closes its leaflets upon mechanical stimulation. An action potential carried by potassium ions propagates slowly away from the site of stimulation causing other leaflets to close as well. The Venus flytrap, Dionaea muscipula, fires a couple of action potentials before closing the trap over an insect within a fraction of a second. Plants do not merely use electricity for exotic behaviour, but more generally rely on ion channels for volume regulation, most notably that of guard cells, as well as signalling involving photosynthesis and chloroplast movement. Electrophysiological signals are the fastest in living nature: The brain resolves time differences of less than 20 microsecond between the arrival of sound in the left and right ears in real time to determine of the direction the sound is coming from. In addition to fast signalling, electrical processes are implied in the sensitive detection of weak signals from the environment. Certain fish bestowed with electroreceptors, sense organs for electricity, react to voltages as small as 1 µV across their body wall.
Ever since the archetypical protozoan ion channel came into being, evolution has engendered a wealth of ion channels with diverse functions, modes of operation and selectivities. The human genome harbours hundreds of genes coding for (units of) ion channels of which over 200 are destined for the plasma membrane alone. An equally impressive number of ion transporters adds to the diversity of electrical signals in living cells. The study of all those channels and transporters in isolation or in their physiological context is a valid goal by itself and is much the realm of electrophysiology.
Electrical signals can equally be used as tokens of health and disease, for diagnostic purposes as in clinical medicine. The electrocardiogram (ECG) and the electroencephalogram (EEG) are standard techniques used in that sense. Even if it might be conceivable to routinely operate equipment for diagnostic purposes without the slightest inkling of its inner workings as if it were a kitchen appliance, such lack of comprehension is certainly not to be recommended in the context of research. Whenever we try to measure something, we cannot prevent perturbing the object of observation to some extent. This is of course a general problem in science, but it appears most acute in the practice of electrophysiology, where it seems to be far much easier to record artefacts than the real events. The best way to dominate these problems is knowledge of the potential interactions between equipment and object under study. This is to justify "Instrumentation" in the title of the book.
The "Methods" aspect of the book partly encompasses instrumentation, since selecting the right equipment for the job is part of a method to record particular events. Other methods apply to the subsequent analysis and presentation of the recorded data.
"Introduction" alludes to our target group, which are graduate students who wish for a solid and up to date basis in electrophysiology. It is specifically destined for readers without a formal training in electronics, signal analysis or electrochemistry and serves as a thorough, yet easy to digest introduction that should lead all the way up from a first recognition of principles to the understanding and the routine application of the various methods. In the early days of electrophysiological recording, amplifiers and other tools were often built by the physiologists themselves. Nowadays, many types of instruments for recording, processing and stimulation, versatile and almost perfect, can be delivered off the shelf. Despite the streamlined technology and the many computer algorithms available for filtering or post-processing of the signals, all students of electrophysiology should gain proper insight in the working principles of their principal tools, specifically the vital stages like preamplifiers and electrodes that are connected to the living preparation under study. In planning experiments with the concomitant purchase of instruments, one has to know the possibilities to choose from and their consequences for the validity of the measurements. Since most of these instruments depend heavily on electronic circuitry, introductory electronics takes a significant part in this book. The chapters concerning it are preceded by a summary of the basics of electricity theory.
An important source of artefacts in electrophysiological records concerns the chemistry of electrodes. Electrodes aren't inert objects but exchange ions with the medium possibly creating an oxidative, reductive or hyperosmotic environment. They may induce the formation of capacitive double layers that are a blessing in some situations and a nuisance in others. The description of electrode properties could be the subject of an entire volume of which our book resumes the principal aspects.
The second edition contains several updates that we will not enumerate at length here. We just mention a few. Notably, the chapter that overviews some of the classical techniques of recording electrophysiological data includes several updates since the first edition. Although electrophysiology is still the field of predilection of skilled personnel, several companies make efforts to shift it to a push-the-button exercise. Their efforts are possibly sparked by the huge interest that the pharmaceutical industry has in the development of automated electrophysiology due to the compulsory electrophysiology tests imposed by the FDA and the EMA. In the wake of these developments several unrelated "on-a-chip" applications have appeared that we now discuss as well in the 2nd edition. The chapter about optical methods in electrophysiology is new to the second edition too. Of course, every electrophysiologist has to know how to use a microscope. In addition, several fluorescent dyes and engineered proteins have become available with which cells can be manipulated and examined optically.
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