Neurophysiology – lecture 4

January 18, 2011

1 Review:
1. Using the Hydraulic Model it is easily seen that force applied to the pump handle for a period of time
does work on the fluid in the pump by moving the piston in the pump a distance ∆s and hence moving
all the fluid in the tubing around the closed circuit.
If the force applied to the pump handle is increased but applied for the same period of time as above
then more work is done and the piston will move farther in the pump and more fluid will be moved
around the tubing.
That is, the more work is done on the fluid the more current is produced.
2. Similarly in the Electrical Model Chemical energy stored in the battery is converted to voltage to
perform work on the charges in the electrical circuit to drive current around the circuit. The more
energy that is thus converted the more current that is produced.
3. In Figures 1D and E it is seen that a constriction in the tube requires more work to be done to move
the same volume of fluid through the tubes. If the length of the constriction is increased an even
greater amount of work is required to move the same volume of fluid through the tubes in the same
period of time.
4. The constriction seen in the Hydraulic Model is analogous to a resistor in the Electrical Model. Thus
the larger the resistor in the electrical circuit the more work is required to produce the same electrical
current passing through the resistor. This qualitative conclusion is generally quantified by plotting the
current produced on the ordinate and the work/unit charge (voltage) on the abscissa.
This relationship is linear passing through the origin and having a constant slope. Since ∆V = I × R,
∆V
the slope of this linear relationship = R = .
I
5. Using glass microelectrodes which have a 1-2 µm opening at their tip a small area of cell membrane
can be bound to the glass while stretched across the opening. If this membrane patch is freed from the
rest of the cell, it is found that this patch of membrane has an exceedingly high resistance - greater
than 100 MΩ.
6. Some membrane patches produce small and brief periods of current crossing the membrane. (Figures
2C and E.)

2 Patch clamp records

1. The properties of these brief periods of current through the membrane include:

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 (a) The amplitude of these current steps is constant.
(b) Their onset and duration is not predictable.
(c) They are brief < 100ms long.
2. Using the Hydraulic Model (Figure 2D) we can explain this current behavior by assuming there are 2
ways that current can cross the membrane:
(a) one way is by a very narrow tube that does not allow much current to pass but is always open
and
(b) is by a larger tube that will allow more current to pass but which intermittently opens and closes.
This latter feature is suggested by the sliding valve.
3. Using an Electrical Model we can explain this current behavior by again assuming there are 2 ways
that electrical current can cross the membrane:
(a) one way is through a very large resistor which is always present and
(b) is through a smaller resistor which passes more current but is sometimes present in the circuit
and sometimes removed from the circuit when a switch is opened.
4. In Figure 3D we see that the magnitude and direction of the currents recorded through the patch
clamp electrode can be altered by changing the voltage present in the electrode. With the inside of the
electrode set at +50mV the recorded current is plotted upward from the base line. While at -50mV
the recorded current is plotted downward from the baseline. This change in the direction with which
the current is plotted represents a change in the direction of the current relative to the cell membrane.
5. By convention if recorded current is coming into the cell across its membrane this current is plotted
downward both in the original recordings and in the I-V graphs summarizing the data.
Conversely, if the current is coming out of the cell across its membrane this current is plotted upward
both in the original recordings and in the I-V graphs summarizing the data.
6. The summary I-V plot at the bottom of Figure 2E shows that the recorded current varies linearly with
the voltage difference applied across the membrane patch.
Thus, ∆V is directly proportional to I. That is, the membrane patch currents are behaving like current
being passed through a resistor.
7. Therefore, these currents are called ionic currents assuming that they are produced by ions passing
through the membrane, like electrons pass through a resistor.

3 The number of ions passing through the membrane during a

current step recorded from a patch clamp electrode
1. Let us assume a patch clamp current of 3pA as is evident in Figure 2C.
2.
3 · 10−12 coulombs 1 mole 6.022 · 1023 ions
3pA = 3 · 10−12 A = · · = 1.8721 · 107 ions · sec−1
second 96500 coulombs 1 mole

3. That is almost 19 million ions/second pass through the membrane when the channel is conducting
ions!! It is hard to imagine anything occurring at that speed.

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 4. It is worth noting that biological enzymes have turnover rates that are nowhere near this fast. For
instance a fast enzyme like acetylcholine esterase (AchE) has a turnover rate of about 100,000 or only
1/180th that of ion movement in a channel.
So the current produced through an ion channel can be way too fast to be due to a biochemical pumping
process.
5. Instead this high rate of charge movement appears to be produced by diffusion.
6. However, diffusion of solutes over any kind of a distance is a very slow process, not a fast one. This is
because diffusing molecules or atoms or ions may travel along a straight path with high velocity but
they are constantly running into other molecules or atoms or ions and having their motion redirected
along a different course which markedly slows their “forward” progress.
7. But biological membranes are so wonderfully thin (5nM) that ions are not running into other ions
during their transit of the membrane and hence are much less impeded in traversing the membrane.
8. Further the path through the membrane is so short and the speed of ion movement at body tempera-
tures is so great that it take only about 10-13 seconds for an ion to traverse a membrane.

4 Properties of ion channels – what creates a fluid-filled channel

or pore?
1. The Skate is a flat fish that generates sizeable electric charges for defensive purposes and to stun
prey. It generates these charges by the use of a large set of modified muscle cells. These muscle cells
are innervated by Acetylcholine-containing axons and have large areas of muscle fiber covered with
acetylcholine receptors.
2. Figure 3A is an electron micrograph of a small part of the surface of one of these muscle fibers showing
the dense distribution of the acetylcholine receptors.
At the bottom of the picture are 10 frames showing individual receptors or a pair of receptors in each
frame. In these frames it is clear that the receptor forms a kind of ring around a dark spot in the
center of the ring. The dark spot is felt to be the fluid-filled channel at the center of the receptor.
3. The “ring” is actually composed of protein subunits. In the case of acetylcholine there appear to be 5
subunits which form the ring around the central pore or channel.
4. In the case of acetylcholine receptors and many other ion channel subunits the entire sequence of
nucleotides in the DNA has been determined for each of these subunits. This DNA can be synthesized
using PCR (polymerase chain reaction).
5. This synthesized DNA can then be injected into frog eggs (oocytes). If DNA for all the subunits of the
acetylcholine receptor are injected then after several days functional acetylcholine receptors are found
in the membrane of the oocyte as determined by recording small currents like those in Figure 2C from
the membrane.
6. These data give clear evidence that membrane channels are composed of protein subunits.
7. In Figure 3B similar analysis shows that all known ion channels are composed of subunits positioned
around a central pore, but that the number of subunits found in different channels can vary.