Chapter 2: Ionic Mechanisms of Action Potentials
Medical Neuroscience Course Lecture John H. Byrne, Ph.D.
Professor, Dept of Neurobiology and Anatomy McGovern Medical School
The University of Texas Health Science Center at Houston (UTHealth) 00:03 Okay, good morning everyone. So the
00:09 question for today, the question for 00:13 today is, “Is it possible to describe
the 00:16 ionic mechanisms of the action potential
00:18 just as we described the ionic 00:21 mechanisms of the resting potential?” 00:23 Well
00:24 interestingly it turns out that at the 00:26 same time that Julius Bernstein back
in 00:29 1902 was proposing his hypothesis for
00:31 the generation the resting potential, 00:33 another early physiologist /
00:36 neuroscientists made a proposal for the
00:40 ionic mechanisms underlying the action 00:42 potential. Now Overton, knew from
00:44 observations that sodium ions in the 00:47 extracellular medium were absolutely
00:51 essential for so-called animal 00:53 electricity and nerve and muscle cell
00:55 excitability. But it wasn't until the
01:00 1940s, when three giants in the field of
01:06 physiology, Hodgkin, Huxley and Katz, 01:08 proposed a satisfactory hypothesis for
01:13 the action potential. And what they did
01:16 was to suggest that whereas the resting 01:20 potential is due to a large extent to
01:22 the fact that the membrane is highly 01:23 permeable for potassium and relatively
01:27 little permeability to other ions, and 01:30 therefore the resting potential could
be 01:32 described as a first approximation by
01:35 the Nernst equilibrium potential for 01:36 potassium. 01:37 What Hodgkin, Huxley and Katz suggested
01:39 was, during the action potential the 01:42 membrane became highly, if not
01:44 exclusively, permeable to sodium, 01:47 therefore the peak of the action
01:49 potential might be described by the 01:51 equilibrium potential for sodium ions. 01:54 And so the hypothesis was as follows:
01:57 that the membrane potential, that's that
01:59 VM, at the peak of the action potential 02:02 is equal to, and I'll put a question
mark 02:06 here, the
02:11 equilibrium potential for sodium (which 02:15 you know). We don't have to put a question
02:17 mark here; we know what it is, because 02:19 according to Nernst the equilibrium
02:21 potential for sodium is 60 times the log
02:24 of the outside sodium concentration 02:29 divided by the inside sodium
02:32 concentration. Now sometimes you see a Z
02:35 down here but Z is the valience, valence 02:38 so that's equal to one. So I'm not
02:40 putting the Z and operations that I use. 02:42 So this was the hypothesis: during the
02:44 action potential, the membrane potential, 02:46 the membrane becomes highly permeable
to 02:49 sodium, and as a result of that high
02:51 permeability the membrane potential 02:53 moves up towards the sodium equilibrium
02:56 potential. Well how do we test this
02:59 hypothesis? We test it just like we
03:01 tested the hypothesis that potassium 03:03 ions who are critical for the resting
03:05 potential. You vary the extracellular
03:07 concentration of sodium and if you vary 03:10 the extracellular concentration of
03:12 sodium, which is easy to do, if you vary
03:14 this, right, then you should change the
03:18 equilibrium potential. You will change
03:20 the equilibrium potential for sodium, 03:21 that's a given. 03:23 But the question is, will you change
the 03:25 peak amplitude of the action potential
03:27 in the same way? Right? And if the two are
03:30 in agreement, then you have provided some
03:32 experimental proof that this 03:34 relationship is correct. So here's the
03:37 experiment that they did. So what you
03:41 see here is a series of three different 03:43 panels: A, B, and C. The black trace
is the 03:46 normal action potential, normal meaning
03:48 that's the action potential recorded in
03:51 the normal concentration of 03:52 extracellular sodium. And what you see is
03:55 three red action potentials, and these 03:57 action potentials were recorded in the
04:00 presence of reduced concentrations of 04:03 extracellular sodium; 70% reduction,
04:06 a 50% reduction, and a 33% reduction. And
04:09 you clearly see that as the 04:11 extracellular concentration of sodium
is 04:13 reduced, so is the peak amplitude of
the 04:16 action potential. And you can plot that
04:18 on a semi-log plot. Just like we did for
04:21 the experiment with changing the 04:22 extracellular concentration
04:24 of potassium, now we're changing the 04:27 sodium concentration. We're measuring the
04:29 peak amplitude of the action potential, 04:31 that's each one of these dots is an
04:33 experimental measure, measurement at a
04:35 respective concentration. And you see
04:38 that as the extracellular concentration 04:40 of sodium is altered, so is the peak
04:43 amplitude of the action potential. Shown
04:45 also on this graph is the straight line; 04:48 that you would predict for how the
04:51 equilibrium potential changes as a 04:53 function of changing the extracellular
04:56 sodium concentration. So you see that
04:59 there's a reasonable good agreement 05:00 between the shape of these two curves,
05:03 right? But it's not perfect, and there's
05:06 over the deviation in that the peak 05:08 amplitude of the action potential is
05:10 always less depolarized than you would 05:12 predict based on the membrane that was
05:14 exclusively permeable to sodium, right? 05:18 Everyone see that? That's clear. So why do
05:21 we have this deviation? The deviation is
05:27 because the membrane is not exclusively 05:29 permeable to sodium because, remember,
we 05:31 still have that potassium permeability
05:33 that we started out with. So no matter
05:35 how high you make the sodium 05:36 permeability, within reason, there is
05:38 still going to be that finite potassium 05:41 permeability that endows the membrane
05:44 with the resting potential, right? 05:46 Here is
05:47 a schematic diagram of an action 05:49 potential, and it shows the wave form
of 05:52 the action potential that we're going
to 05:53 try to explain as we go along today,
and 05:55 it also shows a number of different
key 05:59 potentials. You see this dash line here? 06:01 This is the sodium equilibrium potential
06:03 which for this particular neuron is plus
06:06 55 millivolts. You'll also see this value
06:10 of minus 60 millivolts, that's the 06:12 resting potential, and finally you see
06:14 this value down here of minus 75 06:17 millivolts, and that's the potassium
06:19 equilibrium potential. What you see is
06:22 that the action potential bounds a 06:24 region which on one extreme is the
06:27 potassium equilibrium potential, and on
06:30 the other extreme the sodium equilibrium 06:32 potential. At rest, the membrane potential
06:35 is negative. It's near
06:38 but not equal to, the potassium 06:39 equilibrium potential. Why not? Because
06:44 there is that basal sodium permeability, 06:47 one one-hundredth of the potassium
06:49 permeability, and that makes the membrane 06:51 potential slightly more positive than
06:53 you would predict for a membrane that 06:55 was exclusively permeable to potassium. 06:58 Then you have this value, the sodium
equilibrium 07:01 potential. Note that the action potential
07:02 approaches the sodium equilibrium 07:04 potential but it doesn't get, quite
get 07:06 there. It would only reach the sodium
07:09 equilibrium potential if the membrane 07:11 was exclusively permeable to sodium. It
07:13 doesn't get there because there's still 07:15 that finite potassium permeability. So
07:18 you see, you start with the resting 07:20 potential, you can have an increase
in 07:22 sodium permeability to make the membrane
07:24 potential rise rapidly to the peak. Then
07:27 we have the repolarization phase, and the
07:29 after hyperpolarization phase that we 07:32 need to understand. 07:34 So the big question
07:35 for now is, “How does that switch occur?” 07:38 So what Hodgkin, Huxley, and their
07:41 colleagues suggested was that there are
07:45 two fundamentally different types of 07:47 membrane channels, that span cell
07:51 membranes. One type of channel is a type
07:54 of channel that gives rise to the 07:56 resting potential. These channels are
07:58 normally open, and they allow for 08:00 potassium ions to move out and sodium
08:03 ions to move in. Then they suggested that
08:06 there was a totally different type of 08:08 membrane channel, a so called voltage
08:10 gated or voltage regulated channels. And
08:13 these were channels that were 08:15 hypothesized to open in response to
a 08:19 depolarization. They were normally closed,
08:21 but if the membrane was depolarized 08:23 these channels would open. 08:25 Let me just
08:26 give you a diagram of what the 08:28 properties of one of those channels
08:29 might look like. So if on one axis we
08:33 plot the permeability to sodium, right? 08:36 DNA. And on the other access we plot the
08:43 membrane depolarization. 08:48 And what was suggested, was that there
08:50 are these specialized channels that are
08:52 normally closed, meaning low permeability, 08:55 but as the membrane is depolarized,
these 08:59 channels open allowing the permeability
09:01 of the membrane to sodium to increase, 09:05 right? So this is a diagram of so-called
09:08 voltage dependent permeability because 09:10 the permeability is dependent upon the
09:13 level of depolarization. So this
09:16 diagram, or this hypothesis, allowed you
09:19 to actually predict how an action 09:23 potential might be elicited. Let's say
09:26 that the membrane potential is normally 09:27 here. This is the resting potential here,
09:32 and at the resting potential the 09:35 permeability to sodium is very low. Now
09:38 let's just assume that there's some 09:40 stimulus, whether it be a synaptic
09:43 potential, or some artificial 09:44 depolarization, that moves the membrane
09:47 potential in a more positive direction. 09:49 Based on this relationship, if the
09:52 membrane is now changed from here to 09:56 here, that will tell you that the
09:59 permeability has increased from here to
10:03 here, right? Now you have to take the next
10:08 logical step forward, and ask yourself, 10:10 “What is the consequence of an increase
10:13 in the permeability to sodium?” The answer is
10:17 a depolarization. So as a result of this
10:21 increase in permeability, that will lead
10:24 to a further depolarization, and what.. 10:27 that's not.. there it is.. a further
10:28 depolarization. So this increase in
10:30 permeability will lead to a further 10:32 depolarization. What will be the
10:34 consequence of that further 10:36 depolarization? A further increase in
10:40 sodium permeability. What will be the
10:43 consequence of the further increase in
10:45 sodium permeability? A further
10:47 depolarization. So I think you can see
10:50 that once you get this thing going, you
10:52 rapidly move the membrane potential up
10:55 to these depolarized levels. Specifically
10:59 you can generate an action 11:01 potential in principle by endowing the
11:04 membrane with channels that are voltage 11:06 dependent. Okay, this was the theory; but
11:11 they went on to test this theory, and by
11:13 the way you can see this, how this would 11:15 work from the Goldman, Hodgkin, Katz
11:18 equation; which is sixty again, times the
11:22 log of the outside potassium 11:25 concentration, plus alpha, times the
11:29 outside sodium concentration, divided by
11:34 the inside potassium concentration, plus
11:38 alpha, times the inside sodium 11:42 concentration. That's pretty sloppy. But
11:44 where alpha is equal to the ratio of the
11:48 sodium permeability, to the potassium 11:50 permeability, right? So if the sodium
11:53 permeability starts getting higher 11:55 you see that alpha becomes a larger
11:58 number, so you multiply a large number 12:00 times, the sodium concentration terms,
and 12:02 therefore the Goldman equation would
12:06 predict that the membrane potential will
12:07 move closer to the sodium equilibrium 12:10 potential, right? So you can think about
12:13 it intuitively, or you can do the math 12:15 and just plug the numbers into the
12:17 Goldman, Hodgkin, Katz's Equation. Now
12:22 Hodgkin, Huxley, and Katz, and their 12:23 colleagues work one step further; they
12:26 ask the question, “Well this is a nice
12:27 hypothesis, but how do we prove it?” And
12:30 what they did was to actually go ahead 12:32 and measure the changes in membrane
12:35 permeability as a function of the 12:38 voltage. And here's an experiment, here’s an
12:42 experimental result, and they used a 12:45 technique called the voltage clamp which
12:47 allows one to clamp the membrane 12:49 potential, as I'll show you in a moment,
12:51 at various levels of membrane potential, 12:54 and then measure the sodium conductance. 12:57 The sodium conductance is an electrical
12:59 measure of permeability, we'll use the 13:01 two interchangeably. So here's the first
13:03 experiment that they did; they changed 13:06 the membrane potential from its normal
13:08 level of minus 60 millivolts to a new 13:11 level of minus 35 millivolts. The
13:13 importance of this technique by the way,
13:14 it's called a clamp, is because you can
13:16 change the membrane potential to a 13:18 depolarized level and force the membrane
13:21 potential to stay there. So you don't, you
13:24 prevent, the cell from initiating an 13:26 action potential. Because, if you have an
13:28 action potential, then all the 13:29 permeability changes are going to happen
13:31 and be over within one millisecond. So
13:33 you have to prevent that from happening. 13:34 So the voltage clamp proposal, the
13:38 voltage clamp approach, allows you to do
13:39 that. So what you see is that when you
13:42 change the membrane potential from minus 13:44 60 to minus 35, there is a change in
the 13:47 sodium conductance or sodium
13:49 permeability. If now you repeat this but
13:51 instead depolarize the cell to minus 20
13:54 millivolts, you see that there is an 13:57 additional change in sodium conductance. 14:00 And if you change the membrane potential
14:01 to plus twenty millivolts the change in
14:04 sodium permeability is even greater. So
14:07 what this experiment clearly shows is 14:08 that the greater the depolarization,
the 14:11 greater is the permeability. So you could
14:13 take this value, and then you could take
14:15 this value, and you can take this value, 14:17 and you could plot it on this curve. 14:21 Put your points here, oops,
14:24 put your points here, and you could 14:27 reconstruct this kind of relationship
14:29 between depolarization and permeability. 14:31 So this experiment then gave strong
14:33 experimental evidence that there are 14:35 these voltage dependent changes, these,
14:38 these, channels of the membrane that 14:40 respond to voltage, channels that are
14:43 normally closed, but in response to a
14:45 depolarization, open allowing sodium ions
14:48 to move across the membrane. There's
14:52 another interesting aspect of this 14:54 experimental result, and you see it
in 14:57 the traces. You see that there is a very
15:00 rapid increase in the sodium 15:02 permeability as you depolarize the cell. 15:05 But here's something very interesting;
15:07 note that despite the fact that the 15:09 membrane potential continues to be
15:10 depolarized for this entire period of 15:13 time, four milliseconds, you see that
the 15:15 sodium conductance increases, but then
it 15:19 spontaneously decays back to where it
15:21 started. 15:26 That's a process called inactivation. So
15:28 despite the fact that the membrane 15:30 channels open in response to that
15:32 depolarization, even if you maintain the
15:34 depolarization, they close down. 15:36 They don't like to stay open. They just
15:38 open for a several millisecond period of
15:42 time. So is this just an interesting
15:46 experimental curiosity, or does it have 15:49 some importance? What might be the
15:52 importance of inactivation? Okay, I think
15:57 everybody, even though this is really 15:58 sloppy everybody, like this positive,
this 16:00 is a positive feedback regenerative
16:02 cycle. Once you start the cycle going the
16:05 sodium permeability increases, more 16:06 depolarization, and you rapidly move
the 16:09 membrane potential up to a very
16:11 depolarized level, that's associated with
16:13 a very high level of permeability. So I
16:16 think everybody was comfortable with how
16:18 this could explain the initiation of the
16:20 action potential, but what if there was
16:23 no inactivation, there would be no 16:26 repolarization. So you need something
16:29 once those sodium channels open. 16:31 You need something to close them back
16:33 down again. Otherwise you would only have
16:36 one action potential; you would only have
16:38 one twitch of the muscle, one beat of 16:39 your heart, one thought, one sensation. 16:42 That would be it, and then all your
nerve 16:45 cells would be permanently depolarized. 16:47 Not good. So there needs to be a way of
16:50 returning the membrane potential, back 16:51 down to the resting potential, after
you 16:54 have the peak value. And inactivation is
16:57 one of the mechanisms that contributes 16:59 to that. But is that the only mechanism? 17:06 Somebody's shaking their head saying
17:08 no. 17:09 What's the other mechanism? The
17:12 sodium-potassium ATPase. That is not
17:17 correct. Sounds good, and it is an
17:22 important mechanism. I'm going to get to
17:24 that at the end of the lecture. So you're
17:26 saying you need to activate the pump. 17:29 To, well, because all this sodium came
into 17:31 the cell, and we need to activate the
17:33 pump, to pump it back out again. 17:34 That's essentially what you're saying. 17:36 I'll explain later why that's not
17:38 correct. Okay, your partner is saying a
17:43 voltage-gated potassium. So, yes, in
17:46 addition to membrane channels that are 17:49 voltage dependent, that are closed and
17:51 when they open they allow sodium to come
17:54 in to the cell, there are also voltage 17:56 dependent potassium channels. So these
17:58 are channels that are permeable to 18:00 potassium and, but, are normally closed,
18:04 but in response to a depolarization they
18:07 open allowing potassium ions to move out
18:09 of the cell. This next experiment shows
18:13 you a simultaneous measurement of both 18:17 the changes in sodium conductance, or
18:19 permeability, and the changes in 18:21 potassium permeability, in response
to 18:24 different changes in depolarization. So
18:27 here’s where we're changing the membrane 18:28 potential from minus 62 minus 35. Here
18:32 are the changes in sodium permeability 18:34 we saw previously, and now what you
see 18:37 below is that there are also changes
in 18:39 potassium permeability. If you give a
18:41 larger depolarization, you get a larger 18:43 change in sodium permeability, and you
18:45 get a larger change in potassium 18:47 permeability. If you give even a larger
18:49 depolarization, there is a larger change 18:52 in sodium permeability, and a larger
18:54 change in potassium permeability. So the
18:56 same kind of relationship applies to the
19:01 potassium channels, the voltage defender 19:03 potassium channels, as we saw for the
19:04 voltage dependence sodium channels. Okay? 19:08 Now, despite the fact that this channel,
19:12 and this channel, are voltage dependent, 19:16 there
19:17 are several critical differences between 19:19 the two. One obvious difference of course
19:21 is that this channel is permeable to 19:23 sodium and this channel is permeable
to 19:25 potassium, but there are two other major
19:28 differences. One that is really important. 19:33 And you see it right before your eyes? 19:35 You see the two differences, what are
19:36 they? The latency, so by that you mean
19:41 that whereas the sodium channels open 19:46 very rapidly in response to the
19:48 depolarization, there is a delay in the
19:54 opening of the voltage dependent 19:55 potassium channels, that's the latency
19:57 you're referring to, right? Okay, so big
20:02 deal. So one's fast, the other's slow. Let me
20:08 just think of this for a moment. 20:10 What if the potassium channels didn't
20:14 have that latency? What if the potassium
20:16 channels opened just as quickly as the 20:19 sodium channels? Would that be good? That
20:24 would be bad, that would be real bad 20:26 because, whereas these channels are
20:30 trying to depolarize the cell, opening 20:32 these channels are trying to
20:33 hyperpolarize the cell. So the two
20:35 processes would be working against each 20:37 other, and you probably wouldn't get
any 20:39 action potential. So this latency is
20:43 absolutely essential of, for, the potassium 20:46 channels to contribute to the action
20:48 potential. 20:49 Now they do open, but with a delay,
and 20:53 why is that important? Because it gives
20:54 time for the sodium permeability changes 20:58 to move the membrane potential up
21:00 towards the sodium equilibrium potential, 21:03 do their job, and now the voltage
21:06 dependent changes in potassium can kick 21:08 in and help repolarize the membrane,
21:10 along with the inactivation of the 21:12 sodium channels. So there's two processes
21:16 that are contributed to the 21:17 repolarization: the process of sodium
21:21 and activation, the intrinsic process, and
21:23 the delayed increase in potassium 21:25 permeability. So the nervous system
21:28 is investing a lot in 21:31 bringing that action potential back
down 21:33 to the resting potential as soon as
21:34 possible. 21:35 And why do you want to do that? You want
21:40 to have a short duration action 21:41 potential. Why do you want to have a
21:43 short duration action potential? So you
21:45 can have a whole bunch of them per unit 21:47 time and so that your nerve cells can
21:50 participate in this coding of 21:51 information. 21:54 Okay this next slide,
21:56 actually a series of slides, is just 21:58 going to step us through in a systematic
22:01 way the changes in membrane potential, 22:04 sodium conductance in red, and potassium
22:07 conductance in blue. So let's begin. We
22:11 have some stimulus, let's not worry about
22:12 exactly what it is from right now. It
22:15 could be a synaptic potential, it could 22:16 be an artificial depolarization, but
some 22:19 initial stimulus. This depolarization
22:21 here leads to an increase in the sodium 22:23 permeability. As a result of that
22:25 increase in sodium permeability there is
22:28 a further depolarization. So we entered
22:30 this positive feedback cycle such that 22:32 increase in sodium leads to a
22:34 depolarization, depolarization leads to
22:36 greater sodium in permeability, and we
22:38 rapidly move the membrane potential up
22:40 to the peak value. Note that while the 22:43 sodium changes are occurring very
22:45 rapidly, there is relatively little 22:48 changes in the potassium conductance. But
22:51 now, when we get to the peak of the 22:53 action potential, you see these two
22:55 processes occur. One is that the membrane
22:57 potential starts to decay and that's due
23:00 in part to the inactivation process, 23:03 although the inactivation process itself
23:05 contributes to the repolarization of the
23:08 membrane. But we have the inactivation
23:10 Illustrated here. And then at this point
23:13 in time, you also see that the potassium 23:15 permeability is beginning to increase
23:17 dramatically. A slightly later time, the
23:20 potassium permeability is even greater, 23:22 the membrane repolarization is even
23:24 greater, and a slightly different longer 23:27 time now we see the membrane potential
23:29 has returned back to the resting 23:31 potential. 23:36 This is an interesting point in time
23:38 because the membrane potential is back 23:40 to the resting potential. The sodium
23:43 permeability is back to where it started. 23:46 But look what the packed potassium
23:50 permeability is doing, look what the 23:52 potassium permeability is doing. Just as
23:55 these potassium channels are slow to 23:57 open, they are also kind of slow to
close. 24:02 So what's the consequences of the
24:04 situation at this point in time where 24:07 the sodium permeability is back to
24:09 normal but the packed potassium 24:10 permeability is still elevated? You will
24:16 have the undershoot or hyperpolarizing 24:18 after potential, because the membrane
24:19 potential will be more permeable to 24:22 potassium than it was at rest. Remember
24:24 at rest 24:25 it was 0.01, the alpha value was 0.01
because 24:28 of the resting potassium permeability. 24:31 Now there's going to be this additional
24:33 voltage dependent of potassium 24:35 permeability, and that's going to
24:36 contribute to the after 24:38 hyperpolarization. Eventually the
24:41 membrane channels that are permeable to
24:43 potassium, the voltage dependent channels, 24:47 will return to their basal levels and
24:51 the membrane potential will return back 24:55 to the resting potential. So this then is
24:58 the entire sequence of events that 25:00 underlies the initiation, the
25:03 polarization, the peak value and 25:07 the overshoot, the repolarizing
25:09 phase, and the after hyperpolarization. 25:11 So you need to know all these different
25:14 phases of the action potential. 25:16 You can
25:17 see why it, by itself, cannot contribute; 25:22 it cannot explain the later phases of
25:25 the action potential. If you just look at
25:27 the changes in sodium permeability by 25:28 themselves, you see they extend out
to 25:31 four milliseconds the action potential
25:33 is much narrower than that, so you need 25:35 help. The sodium and activation process
25:37 needs help, and the potassium late 25:40 increase in potassium helps out. Also if
25:42 the action potential was just based on
25:44 these changes in sodium permeability, you
25:46 could have an action potential, and it
25:47 would repolarize, but you wouldn't have 25:49 an after hyperpolarization,
25:50 because in order to get after 25:52 hyperpolarization you have to have the
25:54 net potassium permeability greater than 25:57 it is at rest. 26:01 Now there's some
26:02 interesting pharmacologic work that 26:06 supports this theory. Two drugs have been
26:11 identified that have been useful 26:12 experimentally. One is called
26:15 tetrodotoxin. What tetrodotoxin
26:19 does is illustrated in the next slide, 26:21 using the voltage clamp approach. So this
26:25 is just the changes in voltage dependent 26:26 sodium potassium permeability that are
26:29 elicited with different polarizations, 26:31 and what you see here is, in the presence
26:33 of tetrodotoxin, or TTX, the voltage 26:36 dependent changes and sodium
26:37 permeability are completely abolished. 26:39 What tetrodotoxin does, we know now,
didn't 26:42 know when these experiments were first
26:43 done, is it plugs up the core of the 26:47 sodium channel. The sodium channel
26:49 tries to open. Tetrodotoxin goes in. It
26:51 blocks it up so sodium can not pass 26:54 through. Then there's another interesting
26:57 drug TEA or tetraethylammonium, 27:02 and what you can see TEA does is that
27:06 it selectively blocks the 27:07 voltage-dependent
27:08 potassium channels but not the voltage 27:11 dependent sodium channels. So in a sense
27:13 it's like TEA, it also is an open channel 27:16 blocker. It gets into the channel. It
27:18 plugs it up so potassium ions can't go
27:20 through. So if you had any doubt about it
27:23 before, this clearly shows that these two
27:26 channels are separate. Whereas they're
27:28 both voltage dependent, and they have 27:30 different permeability, as you can see,
27:31 you can selectively block one but not 27:33 the other, so it's not as though one
27:35 channel is the incoming permeability 27:37 permeable to sodium and then later
27:39 becomes permeable to potassium. It's two
27:42 separate channels, and as you'll learn 27:45 throughout the course and the physiology
27:47 there's many, many tens, if not hundreds 27:51 of different types of membrane channels
27:54 that all have a unique role in 27:57 contributing to the excitability of
28:00 nerve cells. We're just going to talk
28:02 about them very generically 28:04 here: the voltage-dependent sodium
28:05 channel, the voltage-dependent potassium 28:07 channel. So within these two drugs, what
28:16 would you expect them to do to an action 28:19 potential? With TTX you wouldn't have an
28:22 action potential, and I don't think 28:25 there's any reason to show
28:27 that because you would, it would block 28:29 the voltage-dependent sodium channels. 28:30 You can't have an action potential
28:32 without voltage-dependent sodium 28:33 channels. By the way, is there any
28:35 clinical application for a drug that 28:37 would block a voltage dependent sodium
28:41 channel, or is this just an experimental 28:43 curiosity or tool? Remember the first
28:46 slide I gave you a bunch of neurological 28:48 disorders. Is there any one of those
28:52 neurological disorders that might be 28:53 amenable to treatment with a drug like
28:56 tetrodotoxin? It obviously'd have to be
29:00 very careful about the dose. Like
29:04 epilepsy? Not like epilepsy? Yes, epilepsy. 29:07 So epilepsy is a disorder associated
29:10 with abnormal neuronal discharges, right? 29:13 So you want to quiet or dampen them
down. 29:16 So one way to do that, I mean it's very
29:18 crude, but is to block some of the 29:20 voltage dependent sodium channels, right? 29:22 But again, you'd have to be very careful
29:23 with it, with it, the dosage. Now tetrodotoxin
29:26 is not used, but there is a drug 29:28 that is used. What's the name of the drug
29:29 that's used to treat epilepsy? I'm thinking
29:36 of Dilantin. There may be others; that's
29:39 one. Okay, TEA, what would TEA do to an
29:46 action potential. Tetrodotoxin is
29:47 going to block them, but with TEA do? 29:55 Someone in the front is saying it would
29:56 make the action potential longer. 29:59 Everybody agree. Anything else? 30:06 You wouldn't have an undershoot. 30:09 Absolutely. Here's the experiment that
30:11 shows that. So here's two action
30:13 potentials: a normal action potential 30:15 and then one recorded in the presence
of 30:18 TEA. You see the initial phase of the
30:21 action potential in both cases looks 30:23 very similar but this action potential
30:26 is much broader and it doesn't have an
30:30 after hyperpolarization. You have an
30:35 action potential that repolarizes, how is
30:37 it possible that this can repolarize? The
30:43 closing of the sodium channels. The
30:46 inactivation of the sodium channel. So
30:48 here, this clearly shows you what the 30:49 inactivation, this is a neat experiment
30:51 in a way, because it shows you what the
30:54 inactivation process by itself does. It
30:57 does, you can have an action potential. 30:59 But the action potential's based solely
on 31:02 the sodium activation process is
31:04 going to be somewhat longer in duration 31:06 than would an action potential be that
31:09 uses the combined processes, processes of
31:13 sodium inactivation and the delayed 31:15 increase in potassium. Okay? <illegible audience question>Yes? 31:21 What would be the consequences of
31:24 ingesting some TEA? It would be really
31:28 pretty bad. So... So...Action potentials, let's
31:36 just talk about muscle cells. You're
31:37 going to learn about muscle cells in 31:39 physiology if you haven't already. So
31:41 muscle action potentials are just like 31:43 nerve action potentials. You have voltage
31:44 dependent changes in sodium permeability 31:46 and voltage dependent changes in
31:48 potassium permeability that contribute 31:50 to the repolarization, and when you
have an 31:52 action potential in the muscle cell,
that 31:53 produces a twitch, right? 31:55 So you mentioned spasticity. 31:56 So what
31:57 would happen if you were to injest a 31:59 bunch of TEA, you would have major
32:01 contractions of all the muscles or the 32:02 skeletal muscles in your body, and your
32:05 nervous system would be pretty 32:08 excitable also. So, you couldn't get
32:14 multiple at, well you could still get 32:16 multiple action potentials, but they
32:17 wouldn't be as... you 32:18 couldn't get as many for what's a given
32:22 period of time, but the whole neuron 32:25 would be more excitable. Okay, I want to
32:29 return to the issue of the sodium 32:31 potassium pump, because it's related
to 32:34 this other issue about the amount of
32:38 sodium that comes into a cell with the 32:40 action potential and the amount of
32:43 potassium that leaves the cell. Now it's
32:46 very easy to get the impression, and I
32:48 must say I've been guilty of it to a 32:52 certain extent, and it's a lot of people
32:54 with guilty to it about it a lot of 32:56 people are guilty in the text books
you 32:58 read, in that you get the impression
32:59 that when you have an action potential 33:00 is this gush of sodium that runs into
33:03 the cell, that moves into the cell. And
33:05 also there's discuss your potassium that
33:07 leaves the cell. You get the impression
33:09 that that could change the concentration 33:11 of sodium inside the cell, because of
all 33:14 that sodium that comes in from the
33:15 outside to the inside, right? Is that what you
33:17 think? You get easily to think that and
33:21 some sodium does come into the cell, but
33:24 it's like a minut amount compared 33:27 to the normal intracellular
33:30 concentration, which is in the millimolar 33:32 range, the amount of sodium that crosses
33:34 the membrane is in the picomolar range. 33:36 So whereas some sodium comes into the
cell, 33:38 it produces no net change in the
33:41 concentration of sodium within the cell. 33:43 The change in potential is across the
33:46 microscopically thin cell membrane. 33:48 That's where the charge distribution
33:49 occurs. There's not a big change in the number
33:52 of positive sodium ions inside the cell 33:55 or a change in the number of potassium
33:57 ions. It's all the changes are across the
34:00 thin surface of the membrane. And that's
34:02 why the voltage dependent that's why the
34:04 sodium potassium pump is not essential, 34:06 because there's no change in sodium
34:08 concentration, so you don't really need 34:10 the sodium potassium pump to repolarize
34:12 the membrane. Okay now the sodium
34:16 potassium pump is important though, in
34:18 the long term. 34:20 An interesting experiment was done many
years ago 34:23 and there's an agent that's used to
34:25 block the sodium potassium pump. What is
34:26 it? 34:30 Somebody saying it. It's kind Ouabain
34:35 Ouabain. Yeah. Ouabain is used to block the
34:38 sodium potassium pump. Actually has some
34:39 clinical utility. 34:41 What's the clinical utility of Ouabain? 34:46 Somebody said, "heart." It's used to treat people
34:54 with congestive heart failure. You'll learn
34:57 about the mechanism for that shortly in
35:00 physiology, but what you can do is you 35:03 can treat a nerve axon with Ouabain
35:06 which blocks the sodium potassium pump 35:09 and in the presence of that Ouabain
you 35:12 can fire more than 500,000 action
35:15 potentials in the presence of that 35:18 sodium potassium pump with there being
35:21 no change in the amplitude of the action 35:22 potential and no change in the resting
35:24 potential. So that experiment clearly
35:26 shows you that the sodium potassium pump
35:30 is not necessary for the action 35:31 potential to action potential sequence
35:34 that we see in the nervous system. Now
35:37 with prolonged treatments of Ouabain, 35:39 however, there is a change in the resting
35:41 potential, there is a change in the peak
35:43 amplitude of the action potential. That's
35:45 because that small amount of sodium 35:47 that comes in with every action
35:49 potential eventually builds up to the 35:52 point where it does produce a
35:54 concentration change, but you need a lot
35:56 of action potentials before that happens, 35:59 So you can think of the sodium potassium
36:01 pump as the generator in your car, right? 36:04 You have a battery in your car and
36:07 the battery needs to be charged. The
36:09 battery can run the lights and a lot of
36:11 the electrical equipment for a long 36:13 period of time. You can leave your lights on for
36:16 24-hours, right, without the engine 36:17 running? That's based on the battery. 36:19 Eventually you got to charge the battery. 36:21 That's what the generator does. So what
36:23 the sodium potassium pump does is charge 36:26 the nerve cell batteries. And there's two
36:29 nerve cell batteries that are important: 36:31 there is the battery for sodium
36:33 (equilibrium potential 36:33 for sodium), and there's the battery
for 36:36 potassium (equilibrium potential for
36:38 potassium). So you maintain the sodium
36:41 equilibrium potential battery by 36:42 maintaining the sodium concentration
36:45 inside the cell low, and you maintain the
36:47 potassium equilibrium potential battery 36:49 by maintaining the concentration of
36:51 potassium inside the cell high, right? So
36:54 the sodium potassium pump makes the 36:57 batteries, charges the batteries, and
36:58 then the action potential uses those 37:01 batteries for its generation. Okay, is
37:06 that clear? Okay we're going to do one
37:11 more one thing, more aspect of the action 37:13 potential, and that's something called
37:16 the absolute and refractory periods. And
37:19 so the absolute refractory period, this 37:22 is a period of time after you initiate
37:24 one action potential where it's 37:27 impossible to generate another action
37:29 potential no matter how much you 37:31 stimulate the cell. All right, let's say
37:34 you initially need a depolarization of
37:36 15 millivolts to get an action potential, 37:38 if you initiate that action potential
37:41 and then try to initiate an action 37:42 potential very soon thereafter, even
if 37:45 you depolarize the cell by 100
37:46 millivolts, you can't get another action 37:48 potential. Then there's this other period
37:51 of time called the relative refractory 37:53 period, where it's possible to initiate
37:56 another action potential but only with a
37:58 greater stimulus. That's relative. So let
38:01 me make a little diagram to show you 38:03 these two different features. So let's
38:08 just say that we have a cell at 38:13 -60 millivolts, and we have the threshold
38:17 of -45 millivolts. Right. 38:26 So, we depolarize the cell to -45
38:30 millivolts, its a reasonable value for the
38:33 threshold, that then would lead to the 38:35 initiation of this all-or-nothing action
38:39 potential and we would have the 38:42 hyperpolarizing after potential, right? 38:47 Let's talk about the relative refractory
38:49 period. What if we, what if we tried
38:59 to deliver to deliver what if we try to
39:02 deliver now a second stimulus. Remember
39:04 this was a depolarization of 15 39:06 millivolts, right? Fifteen. What if we delivered
39:09 that same 15 millivolt stimulus here? It
39:16 would be sub threshold. It wouldn't reach
39:19 threshold, so the same stimulus that was
39:21 used here would not be sufficient to 39:22 fire an action potential here. However, if
39:24 this stimulus was made somewhat larger, 39:27 than we could initiate another action
39:30 potential, right? So in a sense the
39:32 relative refractory period is due at 39:34 least in part to the after
39:36 hyperpolarization. Now the absolute
39:39 refractory period is this period of time
39:41 right there after an action potential 39:42 when no matter how much you depolarize
39:44 the cell you cannot generate another 39:46 action potential. In order to understand
39:50 the absolute refractory period, we have 39:51 to return to this process of
39:53 inactivation, and that's shown here. So
39:57 here you see two voltage clamp pulses. 40:00 This is just like the one I showed you
40:01 earlier, it's just a different time 40:03 compression. You depolarize the cell from,
40:06 what's this, -60 up to about zero 40:08 millivolts, and then you measure the
40:10 changes in sodium permeability. Just like
40:14 what you saw, there's the process of 40:16 activaton, or opening of the channel,
and 40:18 then there's the inactivation process. If
40:21 you give a second pulse, a second 40:22 depolarizing stimulus, several
40:25 milliseconds later, you see that you get
40:28 a change in sodium permeability just 40:31 like the change that you saw the first
40:33 time. But now the interesting result is
40:35 here and that is when you deliver this 40:38 second pulse at times less than this
one, 40:43 if the second pulse comes closer and
40:46 closer to the first pulse, you see that 40:48 the change in sodium permeability is
40:50 less than produced by the first pulse. And
40:52 if the second pulse comes very soon 40:54 after the first, you see the second
pulse 40:56 produces absolutely no change in sodium
40:59 permeability. isn't that pretty weird. So what
41:03 this shows you, what this tells you, is
41:06 that when you open and inactivate a 41:09 channel, it takes time to recover from
41:12 that 41:13 inactivation. The channel opens, and then
41:15 it closes, but it will not allow a 41:18 membrane depolarization to open it again
41:20 until it has a little bit of a rest 41:22 period. You can think of it that way. 41:24 So this slow process of recovery from
an 41:29 activation endows the cell with the
41:31 absolute refractory period. Why? Let's say now
41:33 we're talking about action potentials 41:34 and nerve cells. Let's just say this is
41:36 the first depolarization producing an 41:38 action potential If you tried to
41:39 depolarize the cell again, right here 41:43 you're not going to be able to get
41:44 another action potential because the 41:46 channels have not recovered from their
41:49 inactivation. Right? It's like you've
41:53 taken the sodium channels away. Is there any
41:59 value to the absolute refractory period? 42:08 Is it good for anything? Or is it just... 42:10 actually you might think it's bad, you
42:12 think it's bad because it prevents you 42:14 from getting more action potentials. So
42:21 what if this was the nerve, and you have
42:22 action potential propagating down to 42:24 the end of the nerve, right? What's going
42:28 to happen when it gets to the end? Is it
42:36 going to go back? What stops it from
42:40 going back once it gets to the end? It
42:44 can't go back because the membrane 42:47 channels here are inactivated so even
42:50 though there's a depolarization at the 42:52 end of the axon, that depolarization
is 42:54 spreading backwards, but it can't
42:56 initiate another action potential here 42:58 because the membrane channels that are
left 43:01 behind are inactivated. So this process of
43:04 inactivation which seems kind of silly, 43:06 actually, is absolutely essential for
43:07 existence because without it, once we had
43:10 an action potential in one part of an 43:12 axon, we would have action potentials
43:14 going back and forth continuously 43:15 forever. That wouldn't be very good, right? 43:17 So inactivation is a really essential
43:20 part of the our existence. Okay, and that
43:25 actually leads me to the final topic, and
43:30 that is, once you have an action 43:32 potential at one part of a nerve cell,
43:34 like the cell body, how does it propagate 43:36 to the distant regions of the synapse? 43:38 That distance of course can be many
43:41 meters in length. So that's the process
43:44 of propagation of the action potential, 43:46 and we will pick up with that when we
43:49 return in ten minutes 43:51 Thank you.
