Membrane Properties Lab

Summary:
Today you will utilize a model membrane to begin to understand the principles of
electrophysiological recording.

Background:
A fundamental knowledge of electrical circuits is a valuable tool for understanding
and conceptualizing many aspects of physiological experimentation and theory.
This exercise is intended to acquaint you with some general principles involving
voltage sources, electrical resistance, and intracellular and extracellular recording.
These introductory concepts provide the background for understanding such
phenomena as synaptic transmission and the spread of electrical signals along a
nerve fiber. Today’s lesson will address passive membrane properties.

A word of caution: In standard everyday electrical circuits composed of metal

wires and resistors, current is carried by electrons, which have a negative charge.
In biological systems however, current is carried by ions, which may have one or
more positive or negative charges. By convention, current flows from positive to
negative (although, of course, electrons actually move the other way).

Intracellular and extracellular recordings of electrical activity in the nervous

system are the basic tools of neurophysiology. From them have descended
today’s most powerful methods of studying nerve cell activity: voltage clamping
and patch clamping. Passive electrical properties of excitable tissue are critical
factors that determine electrical conduction in nerve cells (Siegelbaum and
Koester, 2013). For example, space and time constants determine the conduction
velocity of action potentials and the temporal/spatial integration of synaptic
potentials in the brain (Byrne and Shepherd, 2009; Koester and Siegelbaum,
2013).

General Principles
The basic properties of electrical circuits (using direct current) are
described by Ohm’s Law V = IR where V = voltage in volts (V), I = current in amps
(A), and R = resistance in ohms (Ω). This law governs the potentials that are
observed in nerve cells with electrodes.
Exercise 1
Use Ohm’s Law to calculate the current flowing through the following
circuit:

9V 10 K Ω

Current (Network 1) in A: ___________

Convert this current to mA: ___________

This circuit can be useful in modeling electrical properties in cells, but we are
missing one component: capacitance. This refers to the time that it takes to
“charge up” the cell membrane. Lipid bilayers store charge in a circuit and when
the voltage changes, there is a short delay as this capacitance changes. This can
be seen in the curved response of a cell in response to a square voltage change:
(from Cellular Neurophysiology, Stony Brook University, Creative Commons)

Exercise 2
Set up a simple circuit on the breadboard

We will be using a Breadboard to model neuronal membranes. When using a

breadboard there are two rows on top and bottom that are electrically coupled.
These are often used for the input and output leads but we will not be using these
rows. In the middle, between the two sets of rows, are multiple columns of five
cells, each of which is independent of the others. All of the cells in a column of
five are connected to one another, but isolated from the other rows. Finally, the
breadboard is divided into two halves by a midline that is not electrically coupled
to either side. Refer to Figure 1.
 Figure 1. Breadboard Circuit Layout

Find a 10 KΩ resistor and gently insert one end into row D and one end into row I
so that it spans the middle groove of the breadboard. Place the black wire from a
9 V battery into row A, in the same column as you placed the resistor and the red
wire from the batter into row J, again, in the same column (Fig. 2). Set up your
voltmeter: Plug the black (negative) lead into the jack marked “COM” and the red
(positive) lead into the jack marked “VΩmA”. Turn the dial to “DCV 20”. You can
now measure the voltage in the circuit by gently placing the probes from the
voltmeter on the legs on either side of the resistor. Note that you are measuring
in parallel with the resistor. Confirm that the voltage reads approximately 9 V.

Fig. 2

Now check the current calculation you made in Exercise 1. To measure current,
you will have to change the circuit slightly so that the voltmeter is in series with
the resistor. Leave the black input in place but move the red input over a couple
of rows. Think about what this has done to current flowing in the circuit – you
have created a gap over which the current can’t flow because the rows are not
connected to each other. Switch the knob on the voltmeter over to DCA 20m.
This will allow you to measure the current. Now gently touch the leg of the first
resistor with the black multimeter probe on the read side of the resistor and the
red battery wire with the red probe.

Measured current: _______________________________ A

We will now use a battery and a model membrane to simulate passive electrical
activity of a neuron. The model membrane is a ladder-like chain of resistors that
approximates the steady-state electrical characteristics of a real cell membrane.

Figure 3

Figure 3 represents the model membrane schematically. In this model:

 Rm (membrane resistance) represents the resistance of all the ion channels
in the cell membrane
 Ri (internal resistance) represents the resistance of the cytoplasm
 Ro (outside resistance) represents the resistance of the extracellular fluid.

A battery will simulate an electrical potential traveling through the membrane.

Instead of the electrodes, amplifiers, and oscilloscope that are required for real
neural recordings, you will use a voltmeter to examine the response of the model
membrane. Figure 4 shows how the recording techniques you will use with the
model membrane correspond to those used in real cells.
Figure 4. Real and Model Membranes (ion channels are omitted for simplicity). A. In intracellular
recording, an electrode is inserted through the membrane and a voltmeter or oscilloscope records the
difference in potential between the inside of the cell (+ electrode) and the outside of the cell (-
electrode). B. In extracellular recording, both electrodes are outside the cell, so the voltmeter or
oscilloscope records the difference in potential between two points ( + and – electrodes) near the
external surface of the membrane.

Protocol:

To begin, construct your model membrane by following the circuit diagram in

Figure 5.

Figure 5. Model Membrane Circuit Diagram

Your model membrane should run the length of the breadboard.
 The line of Ro resistors should be on one side of the center channel (running
the length of a lettered row B)
 The Ri resistors should be on the opposite side of the channel (running the
length of lettered row I)
 Note that these two lines of resistors should be spaced close enough to the

center channel that the Rm resistors can span the distance between the R o

and Ri lines
 Also note that the numbered columns (consisting of 5 holes each) are
already electrically connected, which will allow you to place the R m resistors
in a separate, but still electrically linked, hole from the R i or Ro lines
 The Rm resistors should be inserted into rows C and H of the same
numbered column.

Set up your voltmeter: Plug the black (negative) lead into the jack marked “COM”
and the red (positive)lead into the jack marked “VΩmA”. Turn the dial to “DCV
20”.
Intracellular Recording
Figure 6 shows the setup for intracellular recording.

1. Connect the voltmeter at the middle of the model membrane (touching

either side of the 7 , or most medial, R resistor works well).
th
m

2. Be sure that the positive (red) electrode is placed on the designated inside
of the resistor ladder and that the negative (black) electrode is directly
across from it on the outside.
3. Start with the battery at the left end of the model with the positive (red)
terminal connected to the inside and the negative (black) terminal
connected to the outside (for your battery connections, use Rows A and J of
whatever column contains your leftmost R resistor).
m

4. Double check that your battery wires are plugged into the correct,
electrically-connected column.
5. Record the voltage at the voltmeter.
6. Now move the battery to the right, moving both the positive and negative
terminals over to the next junction (the column with the next R resistor)
m
 and record the voltage for that battery position. Continue this procedure
until the battery has reached the right end of the model membrane. At
each battery position, record the voltage level and the position of the
battery (node 1, 2, 3, etc.). You should end up with 13 voltage readings.

Figure 6. Intracellular Recording. The voltmeter electrodes span the membrane with the positive
electrode inside the cell. The battery, representing changes in membrane potential, moves along the
model membrane.

Table 1. Intracellular Data

Position / Node Voltage (mV)

9
 10

11

12

13

Extracellular Recording
Figure 7 shows the setup for extracellular recording.

1. This time, place both terminals of the voltmeter on the outside edge of R m

resistors 6 (negative/black) and 8 (positive/red). This should leave two R o

resistors between the terminals, as shown.

2. As before, move the battery from one end of the model membrane to the
other, recording the position and voltage at each node.

Figure 7. Extracellular Recording. The voltmeter electrodes are now both outside the cell. The battery,
representing a change in membrane potential, moves along the model membrane.

Table 2. Extracellular Data

Position / Node Voltage (mV)

2
 3

9.0

10

11

12

13

Lab Questions
Typed answers to the following thought questions are due Jan 24. Please upload
your assignment to the appropriate Assignment folder on WorldClass. You are
welcome (and encouraged) to consult with your classmates, but make sure your
responses are written individually. Worth 80 points.
1. Think about Ohms law. For a given voltage input, how does current flow
change as resistance increases (i.e. does it get bigger or smaller)?
2. For a given current input, how does voltage change as resistance increases?
3. If a capacitor is introduced into a circuit, how would this change the shape of
the cell response to a square voltage input?
4. Make a graph plotting voltage (y-axis) vs. position (x-axis) for your
intracellular recording, using Table 1. Use Excel or Google Sheets.
5. Make a graph plotting voltage (y-axis) vs. position (x-axis) for your
extracellular recording.
 For help answering questions 3-5, refer to the videos posted on the course
website and note that the voltmeter measures the difference in potential
between the positive and negative electrodes.
6. Is the maximum voltage the same for both types of recording? Why or why
not?
7. The extracellular recording changed polarity when the battery passed the
electrodes. Why? Why didn’t this happen with the intracellular recording?
8. What happened to the extracellular recording when the battery was
between the voltmeter electrodes? Why?

Note that the closer the battery inputs are to the recording electrodes in both
cases, the larger the measured voltage drop. This is due to a property called the
length constant, or lambda (λ). It specifies the distance over which a potential
difference (V) across a membrane declines to 37% of its original value.

9. Calculate the length constant for our membrane circuit using the following

equation where R is 10 kΩ, R is 1kΩ, and R is 100Ω in the

m i o

model membrane.

10.What would happen to λ if the membrane resistance decreased? How would

this affect recording of electrical signals?

11. What would happened to λ if the membrane resistance increased? Can you
think of a biologically relevant way that membrane resistance might be
increased?