Voltage- and Patch-Clamp Experiments in Virtual Computer Laboratories (cLABs-Neuron).

Hans A. Braun; Horst Schneider; Bastian Wollweber; Heiko B. Braun & Karlheinz Voigt

Institute of Physiology, Deutschhausstr. 2, 35037 Marburg, Germany

• „cLABs“ is a series of multimedial programs for teaching dynamic biological and physiological mechanisms in an interactive and virtual environment which allows users to perform their own sets of experiments.

• cLABs programs are equally suitable for universities and biology classes at schools as well as for private studies.

Introduction• “cLABs-Neuron”, our latest cLABs-Software, demon-strates the interrelations between ion channel dynamics and membrane currents and voltages. It consists of the modules:

I. Membrane PropertiesII. Ion ChannelsIII. Voltage-/Current-Clamp Experiments

I. Membrane PropertiesThis module provides animations and simulations to basic functions of neuronal membrane properties which are described in terms of their electrical equivalents.

Electrical Equivalents Illustrates the functional membrane properties (= bilipid layer and ion channels) and their electrical equivalents (= membrane capacitance and resistance).

RC-Circuit Visualises current flows and potential changes across a resistor and a capacitor in a RC-parallel circuit.

RC-LabAllows experiments to examine the voltage changes that occur in a RC-parallel circuit when current pulses are applied and no active elements (e.g. ion channels) are involved.

• A variable number of current pulses of pre-selectable amplitude, duration and delay can be applied.

• Resistance and capacitance of the RC-circuit can be changed

ConductanceThis interactive section can be used to learn how the membrane potential can be changed with alterations of ionic conductances. For an intuitive understanding we use a simplified circuit which only consider the Na and K currents and even neglect the membrane capacitance (stationary condition).• You can move the sliders of the potentiometers to see how the membrane potential and currents change as a function of the ionic conductances.

• The buttons allow pre-sets of the conductances to minimal, maximal or equal values and also simulates the changes during an action potential.

Available from:Sheffield BioScience ProgramsFlat 1 Salisbury Heights31 Salisbury RoadEdinburgh EH16 5AA, UKTel: + 4 131 662 8225E-mail: d.dewhurst@ed.ac.uk Price: €490 £310 (Institutional, multi-user licence)

Closing the circuit leads to a constant current flow which divides into a current via the capacitor and the resistor (1-2). At first the major current is flowing on the capacitor. The more the capacitor is charged the more current is flowing through the resistor. When the circuit is opened (3-5), the capacitor discharges across the resistor.

(1) (2) (3) (4) (5)

(1)(1) Superposition of the potential changes during stimulation with repetitive current pulses.

(2) Effect of capacitance changes on the membrane potential after stimulation by a constant current pulse.

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supported by: BM&T Heidelberg/Marburg DAQ-Solutions, Lohra, TransMIT, Giessen

II. Ion Channels

The ion-channel module embraces the structure, gating mechanisms and pharmacological sensiti-vity of voltage dependent Na and K ion channels.

• The first part of the program introduces and describes the main characteristics of one- and two-gate ion channels with a selectivity for potassium and sodium, respectively (1-2).• A following interactive section simulates single channel Na- and K-currents. Here, the user can de- or repolarize a virtual membrane and record the resulting ion currents, as well as watch the gating mechanisms (3).

(1) (2) (3)

Single Channel Lab This virtual laboratory is the main part of the module and shows the time course of Na- and K-currents, according to the number of ion channels in a virtual cell membrane.

The user can stimulate a certain number of single Na- and K-ion channels and the record the respective currents, which are add up to a virtual whole cell current (4).

(4) (5)

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• The Nernst reversal potential (Urev) is calculated from the setting of the inner and outer Na- and K-ion concentration (5). Pharmacological effects can be tested by selection of ion channel blockers, like TEA or TTX (5).

• Various stimulus pulse parameters can be set (6), e.g. to demonstrate that the direction of the Na current is reversed above a certain command potential (Urev)(7).

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III. Voltage-/Current-clamp Experiments

The third module provides a virtual lab for voltage- and current-clamp experiments with different types of neurones and also explains background and concepts of the recording techniques.

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Recording Techniques

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This interactive module shows step-by-step how the generation of actions potentials on current injection (1) leads to the concepts of current-recordings in a voltage-clamp circuit (2). Recording and stimulating electrodes can be inserted into virtual neurons. Voltage- and current-clamp recordings during the application of hyper- or depolarising stimuli of different amplitudes illustrate the transitions from purely passive to active neuron responses.

Voltage/Current-Clamp Lab (2)(1) (3)

The Voltage/Current Clamp Lab:

a virtual computer laboratory for both current- and voltage-clamp experiments with mathe-matically simulated neurons (see recording examples).

The simulations are based on simplified Hodgkin-Huxley (HH) type algorithms.

A neuron editor (left) allows the user to change the neuron’s parameters and to develop and save his/her own favourite neurons.

Recording examples (1-3)

(1) Voltage-clamp recording which also shows individual ionic conductance's and currents.

(2) Passive response and action potentials, one of them drastically lengthened because of application of TEA.

(3) Impulse sequences recorded from a HH-type neuron which was converted into a „pacemaker“ neuron with slight modifi-cations of the activation curves.