Electrophysiology in nanometer scale

Subject description

The course is divided into several sections: Backgroung and history of bioelectrical phenomena, analog electrical circuit of cell membrane, mechanisms and demonstration of resting membrane potential, mechanisms and demonstrations of postsynaptic potentials, action potentials, single channel recording techniques, the voltage-clamp techniques, measurements of membrane capacitance, combining electrophysiological techniques with optical techniques, measurements of cytosolic calcium activity, combined use of electrophysiology and molecular biology techniques to sample cytosol and manipulate gene expression in single cells, combining quantitative imaging techniques to study subcellular structures. Students will use the learned knowledge by using these techniques in their own research. Moreover, by studying the literature they will be able to identify physiological problems that can be addressed with the learned approaches.

Dr. Jernej Jorgačevski, Dr. Mataž Stenovec and other invited lecturers will cooperate at the course.

The subject is taught in programs

Objectives and competences

Electrophysiology represents a subdivision of Physiology, which has been used in routine clinical work and clinical research (recording EEG, ECG, EMG) as well as in fundamental research. In the latter area electrophysiological techniques are used for extracellular, bipolar or unipolar signal registration, but in recent years mostly as nano and microelectrophysiological recordings for the study of single molecules or isolated membrane patches or even isolated subcellular structures. For these approaches different glass micropipettes are used. For example, for the registration of synaptic potentials with intracellular recording techniques very sharp glass microelectrodes are used. For measuring synaptic currents the voltage-clamp method is used. Properties of single ion channels is studied with fire-polished glass pipettes with relatively wide tips under the voltage-clamp conditions (The patch-clamp method). Similarly, measuremnts of exo- and endocytosis in secretory cells and the properties of the fusion pore, with diamenter at the subnanometer scale, the method of patch clamping is used (either in the whole-cell recording, cell-attached recording, inside-out and outside-out recording configurations). The aim of the course is to offer the candidates the key knowledge required to conduct the physiological research of membrane processes at the level of single cells as well as the subcellular membrane bound organelles. Thus the candidates will gain key competences and expertise to conduct their own research in this field.

Teaching and learning methods

Students will focus into the study through tutor-student consultations; work in the lab with cells and equipment, through essay work and through their own research and learning about data analysis.

Expected study results

Knowledge and understanding:

Students will gain fundamental knowledge of the: The voltage-clamp method of whole cells. The voltage-clamp method of an isolated membrane patch, i.e. the patch-clamp technique. Both are used for the measurements of currents and voltages across the membrane of whole cells and for measurements of currents through single ion channels. The latter methods are ideal also for measurements of macroscopic and microscopic changes in membrane capacitance, a parameter that is linear to the membrane area. Membrane area fluctuations reflect the processes of exo- ad endocytosis. Together with electrophysiological techniques, the candidates will learn about combining these methods with optophysiological techniques (measurements of cytosolic calcium and other second messengers) with fluorescent methods to label subcellular structures.

Basic sources and literature

Katz B: Nerve. muscle, synapse. McGraw-Hill, 1966

Aidley DJ: The physiology of excitable cells. Cambridge Univ. Press, 1991.

Hille B (1992). Ionic channels of excitable membranes. Sinauer Associates.;

Sperelakis N (2012) Cell Physiology. Academic Press.

Zorec, R (1988). Ionski kanalčki v celični membrani. Kaj lahko raziskujemo z metodo "patch clamp"?. Med. razgl., 27, št.1 (1988), str.37-53.

Sakmann B in Neher E (1995) Single Channel Recording 2nd ed. Plenum Press, New York