Ionic Mechanisms of Action Potential

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 resti

ng 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, tha

t'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 potas

sium 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? R

ight? 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 potas

sium, 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 pote

ntial 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 a

ction 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 mi

nus 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-hundred

th 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 wh

at 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 calle

d 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 t

his 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 directio

n. 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 w

hat.. 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 p

otential 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 init

iating 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 13:4

9 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 t

ake 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 intere

sting 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 1

6:13 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 y

our 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 voltag

e-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 si

multaneous 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 depolariz

ation, 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 chan

nels? 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 s

odium 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 artif

icial 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 inacti

vation 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 po

tential 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 p

ermeability 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 hyperpolariza

tion. 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. Al

so 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 identifie

d 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. 2

6:39 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 2

7:35 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-depe

ndent 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 clinica

l 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 presen

ce 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 3

0:51 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? Yes? 31:21 What would be the consequences of 31:24 ingestin

g 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 actio

n 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 co

mes 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 with

in 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 c

hange 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 sod

ium 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 amplitud

e 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 th

e 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. Al

l 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 hyperpol

arizing 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 fi

re 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 ord

er 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 4

0:16 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 close

r 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, a

nd 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 42:2

1 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.