The McGill Physiology Virtual Lab

Compound Action Potential

Background > Recording technique

In this laboratory, you will be recording an extracellular, biphasic, compound action potential. Clarifying  those three terms in the context of the experiment will help you to understand and interpret your results.


Intracellular versus Extracellular recording



One can measure a  single trans-membrane potential  by inserting a glass pipette into one cell and recording the potential changes with respect to an extracellular reference electrode.  This intracellular technique is used, for example, to record the resting membrane potential of a muscle in the Resting Membrane Potential Lab.

The intracellular recording technique does allow for very accurate assessment of the electrical activity of a single cell, but it is very difficult to do in vertebrate nerve fibres and can involve considerable damage to the membrane around the electrode tip.
A far less demanding technique, extracellular recording, involves placing one electrode in close proximity to the excitable cell and the reference electrode at some location in the extracellular fluid.  This arrangement records potential changes at the membrane surface rather than across the membrane.

Extracellular positioning of the recording electrode

Action potentials recorded extracellularly differ from those recorded intracellularly in several important respects.  The size of any one action potential will be obviously reduced.  The shape of the waveform for any one action potential will depend on the exact geometry of its contact with the electrode. Extracellular techniques are therefore better suited where one only wants to know that an action potential has occurred or to record the activity of an entire population of cells.


Biphasic recording

In this lab you will study the response of the sciatic nerve of the frog to electrical stimulation using two pairs of stainless steel wire electrodes that are both in contact with the nerve, therefore recording the potential difference between two points on the nerve.  There are 12 such recording electrodes in the nerve bath, but only two can be connected to the differential amplifier at any one time. 

Modified from ADinstruments. All Rights Reserved.

Delivering a sufficiently large stimulus to the nerve will result in an action potential that is quite a bit larger than a single intracellular action potential but looks remarkably similar.
This compound action potential (CAP) is the algebraic summation of all the action potentials produced by all the fibres that were fired by that stimulus.  The nerve is made of thousands of axons whose size, myelination and position with respect to the stimulating and recording electrodes all affect the size of their contribution to the compound action potential. 
Both the classic intracellular action potential and the compound action potential are biphasic.  In other words, they have both positive and negative deflections, but for different reasons.  The negative phase of the intracellular action potential is attributed to the mechanism of after-hyperpolarization.  The negative phase of the CAP is due to the manner in which it is recorded, which will be explained below.
There are two wire recording electrodes (R1 and R2) touching the nerve, each connected to one input of the differential amplifier.  The animation below illustrates how the shape of the CAP depends on the position of the two electrodes with respect to the travelling CAP.  For an in-depth explanation, please read on below.

Before the stimulus is delivered, both wires should be measuring basically the same voltage.  There will be no deflection recorded because the amplifier takes the difference of the two inputs before passing the signal on to the A/D converter.

The situation changes as the CAP travels along the nerve.  The shape of the CAP will depend on the relationship between the inter-electrode distance, the length of the axon segments depolarized by the action potentials, and the conduction velocities of the axons.  When the CAP has reached the first recording electrode (R1, proximal), the proximal electrode becomes transiently negative to the distal electrode; the potential difference between the two is detected and the trace is displayed as an upward deflection on the screen. 
As the CAP progresses between both recording electrodes, the recorded potential returns to the base line (no voltage difference between the two recording electrodes).

As the CAP passes the second electrode (R2), a deflection of the same size but opposite sign will be recorded.  The sign is negative because of the way the amplifier compares the two inputs.

Theoretically, if the electrodes are sufficiently far apart, a short segment of 0 deflection will be recorded before the CAP reaches the second electrode. It may not or may not be difficult to observe this very short segment: the recording electrodes have to be sufficiently far apart. If in a frog nerve at room temperature, the duration of the action potential is about 1.5 ms, and the conduction velocity is about 20 m/s, what would be the distance occupied by the active region of the action potential? How far apart should the recording electrodes be in order to see a short segment of 0 deflection?            
If the electrodes are not separated by such a large distance, the two phases will not be of equal amplitude.  The CAP will not have completely passed the first electrode before reaching the second.  Adding the two opposite signed deflections will reduce the amplitude of the negative phase and decrease the apparent width of both.
If one were to crush the nerve at the second electrode -effectively permanently depolarizing the membrane at this location -  a monophasic CAP would result.


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