2011.11.25.. TÁMOP – 4.1.2-08/2/A/KMR-2009-0006 1 Development of Complex Curricula for Molecular Bionics and Infobionics Programs within a consortial* framework**
Consortium leader
PETER PAZMANY CATHOLIC UNIVERSITY
Consortium members
SEMMELWEIS UNIVERSITY, DIALOG CAMPUS PUBLISHER
The Project has been realised with the support of the European Union and has been co-financed by the European Social Fund ***
**Molekuláris bionika és Infobionika Szakok tananyagának komplex fejlesztése konzorciumi keretben
***A projekt az Európai Unió támogatásával, az Európai Szociális Alap társfinanszírozásával valósul meg.
Neurobiológia alapjai - Módszerek
BASICS OF NEUROBIOLOGY - Methods
By Imre Kalló
2011.11.25. TÁMOP – 4.1.2-08/2/A/KMR-2009-0006 3
METHODS IN NEUROBIOLOGY VII.
Electrophysiological approaches
Imre Kalló
Pázmány Péter Catholic University, Faculty of Information Technology
I. Histology techniques: light microscopic studies II. Applications using fluorescent dyes
III. Histology techniques: electron microscopic studies IV. Techniques to map neuronal connections
V. Molecular biological techniques VI. Living experimental models
VII. Electrophysiological approaches VIII. Behavioral studies
IX. Dissection, virtual dissection, imaging techniques
ELECTROPHYSIOLOGICAL APPROACHES
In vitro electrophysiology
Patch-clamp recording
Voltage clamp, Current clamp modes
Loose patch, Cell-attached, Whole cell, Perforated patch modes
In vivo electrophysiology
Anesthetised and moving animals Measuring field potentials
Multiunit recordings Single cell recordings
Juxtacellular recording Intracellular recording
TÁMOP – 4.1.2-08/2/A/KMR-2009-0006 5 2011.11.25.
IN VITRO ELECTROPHYSIOLOGY: PATCH CLAMP TECHNIQUE
Cell-attached mode:
Transfer is only through ion channels entrapped
Whole cell mode:
Content of the pipette redistribute with that of the cytoplasm
Perforated patch mode:
Ion transfer through the
perforations, but large molecules stay within the cytoplasm
IN VITRO ELECTROPHYSIOLOGY VOLTAGE CLAMP
Current meter
Membrane potential amplifier
Signal generator
Feedback amplifier Intracellular
electrode
Extracellular electrode AXON
The cell’s potential is clamped at a chosen value. The size of ionic current crossing the cell’s membrane at the chosen value is measured.
Circuit diagram depicting the feed-back amplifier charging the cell membrane of the recorded cell with the digitally subtracted transient capacitive currents.
Xu C et al Histamine innervation and activation of septohippocampal GABAergic neurones: involvement of local ACh release. J Physiol.
2004 Dec 15;561(Pt 3):657-70.
TÁMOP – 4.1.2-08/2/A/KMR-2009-0006 7 2011.11.25.
IN VITRO ELECTROPHYSIOLOGY CURRENT CLAMP MODE
The current clamp technique records the membrane potential by injecting current into a cell through the recording electrode. The membrane potential generated by the cell „spontaneously” or in response to external stimulus is measured.
Xu C et al Histamine innervation and activation of septohippocampal GABAergic neurones: involvement of local ACh release. J Physiol.
2004 Dec 15;561(Pt 3):657-70.
IN VITRO RECORDINGS – PAIRED RECORDINGS
TÁMOP – 4.1.2-08/2/A/KMR-2009-0006 9 2011.11.25.
IN VIVO EXTRACELLULAR RECORDING
IN VIVO EXTRACELLULAR RECORDING
MULTIELECTRODES
TÁMOP – 4.1.2-08/2/A/KMR-2009-0006 11 2011.11.25.
IN VIVO JUXTACELLULAR RECORDING
An extracellular recording technique
It makes a correlated electrophysiological and morphological examination possible Electrode and marker of cell filling (neurobiotin)
Fine positioning of the electrode – changes of the amplitude of action potentials Current impulse (1-10 nA, anode impulse) – modulation of neuronal activitity Duration of filling: 15-20 min
MEASURING FIELD POTENTIALS
V From the surface of the skull (EEG)
From the surface of the brain
From the brain (a certain brain region)
The field potential is the summation of spatial and temporal alterations of
synaptic and voltage-dependent currents in a defined region of the brain.
Consequently, it refers to and
characterizes the activity of a certain cell or afferent population.
