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BIOL 4315 & 6315 Neuroscience Mon-Wed 1-2.30 PM - Neuroscience Lecture 08 Action Potential 3 and Backpropagation
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00:02 In particular things that we talked about gated sodium channel is what happens once
00:10 have depolarization and the kinetics of that . In particular, we talked about
00:17 if you have depolarization, this is voltage gated channel that contains this voltage
00:23 and S four transmembrane segment in each and that segment is positively charged.
00:29 as long as the membrane potential is on the inside, it is drawn
00:34 that negative charge. But as there a shift in the number and potential
00:38 the depolarization here, you have essentially positive charge acting as repellent,
00:46 this positively charged segments of amino acids changing the confirmation of this channel leading
00:54 the opening of this channel. But the fact that depolarization was sustained,
01:00 channels very quickly inactivate because they have gates, they have activation gate and
01:07 gate. And with the movement of voltage sensor, it opens activation
01:12 but that same movement and confirmational And the channel causes for the second
01:18 inactivation gate to close the channel. the only way we can de inactivate
01:25 release this inactivation gate, remove it the channel is by hyper polarizing the
01:31 membrane again post the resting membrane So we also talked about the fact
01:37 during the rising phase of the action , you have a very large driving
01:43 for sodium until it reduces. So more depolarize the mene potential, the
01:49 is the driving force for sodium. we said this is only one reason
01:55 membrane potential doesn't reach equilibrium potential for . And then we said the second
02:00 is the kinetics of the sodium channel that as soon as they open they
02:04 close. So you have inactivation of channels. And that's why during this
02:09 , which we call the absolute refractory , all of the sodium channels are
02:15 closed. So you cannot depolarize the further to produce more excitability, more
02:23 and other action potential. You have now rep polarize the membrane with the
02:28 of potassium e flux, which will drive the membrane potential towards its equilibrium
02:35 . Because at the peak of the potential, there's a big difference between
02:41 membrane potential and ek the ionic equilibrium for potassium. This is a very
02:48 driving force for that. Uh And also know that that this uh position
02:56 , potassium channels and particular leak channels dominating the conductance across the membrane.
03:03 this is about sodium voltage gated sodium . So we discussed two conditions uh
03:11 plus generalized epilepsy with febrile seizures pro and SME I, which stands for
03:26 myoclonic epilepsy of infancy. We talked how mutations to various parts of this
03:48 genetic mutations we call channelopathy can end in causing either GS plus or sme
03:57 severe. My chronic epilepsy of also known as Dra syndrome. And
04:14 also discussed this concept of febrile seizures we talked about high febrile seizures.
04:20 the most common type of seizures, in developing brains that they can be
04:25 with high levels of hyperthermia. So the temperature goes up, typically 100
04:30 four °F and stays there for a amount of time. Um Having one
04:36 two febrile seizures especially does not qualify as being a person with epilepsy or
04:43 with epilepsy. So you have to um multiple seizures typically and unprovoked seizures
04:51 there are different types of seizures. we talked about generalized, right.
04:55 talked about loss of consciousness and we about gus plots. Uh And then
05:00 said that there's so many different expressions seizures. Some of them have a
05:07 epilepsy and status epilepticus with full collapse loss of consciousness, what we
05:14 tonic clonic behavior of muscles, contracting relaxing in a tonic kind of a
05:21 and then walking out for a So there are all of these different
05:26 and different epilepsies really. And the is that some of the uh seizures
05:32 not even have a motor component I just have an um emotional component or
05:38 can have a loss of consciousness with a blank stare also not necessarily having
05:43 contractions or any motor component. So is all of the information that's really
05:50 . Now, neurotoxins and NAV we have talked about how there are these
05:58 toxins. We talked about how these toxins, in particular, when we
06:04 at the uh all of the techniques Roderick mckinnon used in order to solve
06:11 structure of the potassium channel. And later, if you remember he used
06:16 ray crystallography and then to visualize that , we've mentioned several different techniques.
06:24 we also mentioned that amongst those techniques the fact that he was using
06:30 He was using scorpion toxins. He using spider toxins and these toxins are
06:37 to interact with the types of channels we're discussing voltage gated sodium channels,
06:44 gated potassium channels. There's a variety receptors and protein channels and protein
06:52 So the G protein coupled and Ln channels and voltage gated channels and all
06:58 them have their own conserved across the . It's within one family like potassium
07:05 gated potassium channels. So have conserved , but those sequences are gonna be
07:10 from family voltage gated. So channels will have a variety of different subtypes
07:16 channels with in the family with some sequences, but also different from the
07:22 channels and so on and so And so the fact of the matter
07:26 different substances the way they interact with channel. So there are ses
07:30 it's imagine it's like a door that multiple locks on it. And you
07:36 to have a precise key in order open a lock. And some of
07:42 locks will just one key, one will open the door wide. And
07:48 lot of these are referred to as or something that opens the channel is
07:53 to as agonist. Sometimes it can a full agonist because it immediately opens
08:01 channel. Some of the substances will a partial effect, partially open the
08:08 . Some of the substances will keep channel open longer but will not really
08:13 the opening. It's just once it's , it will keep the channel
08:17 others will close the channel faster and that are antagonists. If agonists opens
08:25 channels, antagonists sometimes also used interchangeably blockers, they will essentially block channel
08:41 . So we are talking about in voltage gated sodium channels and toxins here
08:50 , coming in this case from puffer are used as experimental tools. So
09:03 loves Simpsons, right? What? not opening exactly what we're talking
09:25 Let's see if I get good. on it. Hey, hey,
09:29 this fugu? It is blue. I should warn you that one fo
09:39 . They say this cat shaft's a mother but I'm talking about chef
09:46 He's a complicated man. But X she's here. Cover for me.
09:58 to go. Not fugu if it cut improperly it, yes.
10:03 It is poisonous. Potentially fat. if sliced properly, it can be
10:08 tasty. I must get the Grab that one. Your hair
10:18 Um Master. You are needed in kitchen. I said color dart but
10:24 , we need your skilled hands. skilled hands are busy. You do
10:31 . Mhm. Poison, poison, . Concentrate. Who? Mm F
10:51 tasting beautiful language, isn't it? god's sake. Don't eat another
11:09 I couldn't bur the Mr Simpson, . I shall be blunt. We
11:13 reason to believe you have eaten poison . What should I do? What
11:19 I do? Tell me quick? need to panic. There's a map
11:22 the hospital on the back of the . Try something. New homer.
11:28 it hurt you? Homer? I heard of a poisoned pork chop.
11:33 wife agreed that I should break this you. No need doc. I
11:36 read Marge like a book. it's good news, isn't it?
11:42 , Mr Simpson. If in you've consumed the venom of the blowfish
11:46 from what the chef has told me quite probable you have 24 hours to
11:52 24 hours. Well, 22. sorry. I kept you waiting so
11:57 . Oh, I'm gonna die. gonna die. Well, if there's
12:02 consolation it's that you will feel no at all until sometime tomorrow evening when
12:07 heart suddenly explodes. Now, a death anxiety is normal. You can
12:12 to go through five stages. The is denial. No way because I'm
12:16 dying. Second is anger you after comes fear, it's fear. What's
12:22 ? Fear? Bargaining doc? You get me out of this. I'll
12:25 it worth your while. Finally. . Well, you all gotta go
12:29 time, Mr Simpson. Your progress me. I should leave you two
12:33 . Perhaps this pamphlet will be So you're going to die.
12:48 There's another uh link that hopefully works us. Yeah, introducing the
13:01 But the marine world has other more delicacies in store known to hardly anyone
13:06 the West. For example, the , the poisonous puffer fish of which
13:12 are about 100 species worldwide. You a license to sell puffer fish in
13:18 . But as a buyer, you one too, Okamoto has a fugu
13:28 and of course a license. He's here to buy the increasingly popular non
13:33 farmed fugu, which can be recognized its shorter fins. Nor is he
13:39 in the smaller species caught in the from Japanese waters? This true connoisseur
13:45 only looking for one thing, toxic fugu as fresh as possible. And
13:51 means Tora Fugu tiger puffer fish, Kobe beef of Fugu cuisine,
14:06 Made a single specimen of this species is only found in the sea of
14:15 may well cost €100. One of historic districts is located around Asakusa
14:31 Most wild Fugu restaurants are to be here. There are about 3000 restaurants
14:37 in Fugu in Tokyo. Today. the outside, they're usually easy to
14:43 and they're always highly specialized. One them is Rizo Okamoto restaurant where sometimes
14:53 Prime ministers drop by. Wo say its name the pure fish place.
15:00 also need a license to prepare The poison in Fugu is tetrodotoxin.
15:07 1000 times more potent than cyanide and is no antidote. The poison paralyzes
15:13 victims but leaves them fully conscious. preparation is critical. The skin and
15:21 of the fish are poisonous and they not contaminate the non toxic meat on
15:26 Mussels. High concentrations of highly poisonous are found in the innards, especially
15:42 liver and ovaries. Special disposal is takes about 10 minutes to neatly
16:01 A tour of most commonly, the is cut into thin slices and
16:06 Roper fish is unspectacular rather bland. U Ozai restaurant exclusively serves its guest
16:14 captured in the wild in small The poison of the Fugu fish triggers
16:22 in the mouth and is intoxicating. here they do not cater for guests
16:29 might be eager to try out tiny of this poison. What we serve
16:42 is 100% non toxic. If we this law, we're ruined. Just
16:49 me myself, a real fugu meal of at least six courses. This
17:03 in gourmet pleasures is mainly enjoyed in when there is something to celebrate.
17:08 , the chances of being poisoned at fugu restaurant are practically zero. Thanks
17:13 the high demands placed on the I see what, 50 years
17:28 more than 100 Japanese died each year fugo poisoning. Today, there are
17:34 three a year, all victims of amateur and recreational cooks. I'm not
17:47 at all. I ate a Fugu a child and it always tasted very
17:51 to me. Fu is a delicacy eaten in Japan despite earthquakes, tsunamis
18:03 nuclear disasters. Rizo, Okamoto will run out of work and in the
18:08 too, it's likely to remain quite . I did go ok. All
18:20 . So this is what we're talking . So every time you're gonna think
18:26 Simpsons or puffer fish, you're gonna tetrodotoxin and tetrodotoxin uh was found of
18:35 and isolated in Japan because of the aspect of it, people consuming food
18:42 . So I guess there's a little of conflicting kind of a description in
18:47 video like we serve 100% nontoxic and the narrators says that they don't cater
18:55 doses here. They get large So it's a little bit confusing.
18:59 I guess that uh the way it toxic. And the reason why hundreds
19:05 people used to die from it is it's delicacy. But the way it
19:09 toxic is if you don't eliminate the that they were mentioning that contain
19:18 And it's in this puffer fish that the toxin, it's actually not synthesized
19:23 puffer fish. It's bacteria that inhabit puffer fish and produce this toxin.
19:31 so the liver that he was mentioning particular. OK. So buffer fish
19:38 a lot of other uh uh a of other animals that may contain these
19:47 , they don't synthesize them. And certain puffer fish will be completely tetrodotoxin
19:53 . And there's certain subtypes of puffer that are susceptible to the bacteria to
20:00 that bacteria and producing high levels of . So the toxicity comes, if
20:07 preparation methods are not correct, you to eliminate that from the actual meat
20:13 you're eating. But it is suggested definitely is that the meat itself will
20:20 contain very low levels of TT And that's a whole thrill that they
20:27 . You get slight numbness in your . And that's because it blocks uh
20:34 acts as an antagonist as a blocker voltage gated sodium channels that we're
20:42 Uh also remember. So this is , deadly. It goes through the
20:48 chain. It is. So if ingest it, it's now part of
20:52 food chain. Uh these uh voltage sodium channels are not only located in
20:59 C MS, they're also located in parts of the body and it can
21:05 paralyzed from, from and death can about due to lack of breathing and
21:13 up of the diaphragm and inhibition of proper contractions of the diaphragm.
21:20 it is uh what we call a reversible antagonist because it can be washed
21:29 . It doesn't form a covalent bond the channel, so it can be
21:35 away. So, although there, no antidote if you get a
21:39 you know, they got poisoned with X, by the way, do
21:43 know what happens here in Texas? you get bit by strange insect,
21:47 haven't seen a snake. Um, spider. No. Well, there's
21:59 control apparently and there's poison control number that's what you're supposed to do.
22:05 the reason for that is because they're familiar with what might have happened to
22:10 and the steps that you may need take as opposed to you calling a
22:15 or doctor or, or, or of the uh medical treatment facilities
22:21 er, room. So before you there or if you call them,
22:25 just say come in, they're not in that. So the, it's
22:28 something and something poison control of But I'm sure if you google poison
22:33 Texas spider bite or uh some other , it will, it will show
22:38 that information. I didn't know that last year, my instinct would have
22:43 to call 911 to seek medical But, uh, I heard this
22:48 the radio program, public radio Uh, and, uh, they
22:53 that you have to call the poison because a lot of times 911,
22:58 won't be able to walk you through to do kind of. And it
23:01 be, you know, a matter minutes with some of these toxins and
23:06 also. So be careful out We have a lot of stuff crawling
23:11 the Gulf coast. And um we rattlesnakes actually in the marshes here all
23:19 the Gulf coast. Uh So there's, there's all sorts of bugs
23:24 carry these things that part of the chain. Again, the the fish
23:27 not synthesize this toxin, that is part of the bacteria that is found
23:33 certain species will not be susceptible to bacteria. Now, recall, we
23:39 about the voltage clamp and we said this voltage clamp setup allows you to
23:44 the command potential. And so that can essentially control the number and potential
23:51 of passively recording. Oh, it up, it shifted down, you
23:56 now lock and hold or control the and potential at the desired value with
24:02 voltage plan technique. So doctor Toshi Narahashi that was working with these
24:09 Actually, in the seventies, not people were talking about channels and things
24:15 that uh he was one of the of kind of a channel understanding channel
24:22 if you may and he had this . But what he did is that
24:27 saw that it blocks action potentials. so he postulated that it blocks action
24:33 because it blocks the rising phase of action potential by blocking voltage gated sodium
24:41 . So, right, if you a substance and you have a passive
24:45 , you have an action potential. you apply the substance and action potential
24:49 gone. What you have learned about potential from Hutchin and Huxley in the
24:54 and sixties that sodium goes in potassium out. So if the action potential
24:59 form, you would suggest sodium cannot in there's no rising phase of the
25:04 . OK. And that was, what you thought. But in order
25:09 demonstrate it experimentally, in order to it experimentally, right, I always
25:13 that there's calculations, there's observations from from the recordings, but you want
25:19 have an experimental proof. And this that when finally Toshi Nahai took him
25:26 couple of years to take a Well, not a couple of
25:29 he flew with this vial to United full of toxin. It took him
25:34 couple of years to get to the clamp reported. So if you're talking
25:39 uh eighties, you had these setups they were pretty sparse, maybe larger
25:46 would have one of these setups that expensive. It was still fairly new
25:51 that time, the voltage clamp. so what he did is that he
25:56 voltage clamp, he depolarized the plasma . And he saw this very strong
26:01 current, which we already know this the inward sodium current that was followed
26:06 this prolonged outward current. And what know is that this is an early
26:12 current that has the properties. We early activation, fast activation, but
26:17 fast inactivation within a millisecond or so current followed by later activated potassium outward
26:27 , which is sustained and prolonged. so he added TT X on this
26:36 . And what we saw is that we we have the same depolarization in
26:41 presence of TPX, almost 100% blocked or sodium purpose. So that was
26:49 experimental proof. It's not just blocking potential. So proving it is blocking
26:56 voltage gated sodium conductance and it does affect voltage gated potassium conductance. In
27:05 presence of depolarization, you still got outwardly activated part because potassium channels are
27:13 are open, there's no antagonist, no blocker like the sodium channels.
27:19 , in this situation, if you another blocker, in this case,
27:25 Tetra elei, you abbreviated as te tetra ey ammonium te A. If
27:36 apply tetra Thami tetra thy ammonium does affect inward sodium currents at all,
27:44 it is rather specific to voltage gated currents. And it blocks potassium
27:52 So we have NTE A or tetra is a, is a chemical
27:58 It's not a natural toxin. And we have toxins agonists, antagonists and
28:06 of those that come from nature You have insects, you have fish
28:14 them, uh they are part of venom and in some animals and then
28:22 have also chemical synthesized or chemical substances will be very specific to either voltage
28:30 sodium channels, voltage-gated potassium channels and on. So you should have probably
28:38 . So if these mutations cause these forms of epilepsy, what happens to
28:45 function of this channel? Does it the function of the channel? Decrease
28:51 function of the channel? Most of mutations with gaps and dismay that will
28:56 the function of the channel. But of these channels like voltage gated sodium
29:02 . So precise subtypes will be expressed both subtypes of cells, ex excited
29:08 inhibitory. So other therapies and medications target voltage gated sodium channels. We
29:16 talked about the toxins and how toxins bind and block them. But then
29:21 said there are agonists that can help and promote activity in these channels.
29:26 indeed there are substances and pharmaceutical medications target voltage gated sodium channels in trying
29:35 repair the dysfunction that is happening in voltage gated sodium channels. But it's
29:42 about the balance of excitation and an . So if you are affecting.
29:48 this case, you have too little and in this, in these conditions
29:54 plus and sme I so a lot epilepsy drugs will actually tries to raise
29:59 levels of inhibition. But we'll talk this when we talk about GABA transmission
30:04 we'll discuss some of the gaba uh that are used for controlling seizure activity
30:12 , and Children including gas since many . Yeah. Uh you talked about
30:16 seizures and how they're not like a for a genetic disorder. But his
30:24 is like the febrile seizures that you're about that are normal or is it
30:28 disorder? So what it it is total, it is a disorder.
30:32 generalized epilepsy that has a component of seizures. And uh if in normal
30:42 brains, if you have a temperature , let's say 100 4 °F,
30:46 could lead to febrile seizure in humans have mutations in these channels. A
30:54 rise in temperature from normal body physiological to change to let's say 99 °F
31:02 degrees. It's, it's enough to much higher sensitivity to, to hypothermia
31:09 much lower threshold for having hypothermia induced . So, ok, so think
31:17 it from all sorts of perspectives, using toxins to solve the channel
31:24 We're using toxins to demonstrate how it the currents through the channel. Are
31:29 blocking the currents antagonists? Are they ? Are they opening the channel more
31:36 they interacting, that means they're If you have mutations in different
31:41 the same way you'll have different toxins are interacting with different parts of sometimes
31:48 the same channels. So we have variety of toxins that affect sodium
31:53 Uh We have red tides and as global temperature goes up on average,
32:01 gulf waters in particular temperature in the waters is, is rising and during
32:07 summertime, it's like a warm It's not even refreshing sometimes to,
32:12 swim down here. But also that an environment for a lot of bacterial
32:20 . And quite often during really hot of the summer, especially if the
32:27 have remained still without too much rainfall uh changes in the, in the
32:34 of the current movement or tides or . You can have a build up
32:39 this bacteria and this bacteria which uh essentially will form itself within the
32:52 for example. And you will have uh often warning that uh uh sometimes
32:59 the months during which you're not supposed consume shellfish in the southern climate.
33:07 I was explained that all monks that have r are safe to consume
33:16 are not safe to consume shellfish all that don't have R. So January
33:23 our February has our March, has April has R May June,
33:29 R, July, no, August, no, R.
33:33 And that's also related to the climate and to the temperature changes. And
33:37 September is not still gonna be not safe. These bacteria can multiply and
33:42 conditions. They can inhabit themselves within shellfish uh with a toxin that is
33:49 sait toin. So it at like x like tetrodotoxin. Saxitoxin is a
34:00 channel blocking toxin and Bato Toin that shown here. Bato Toin is found
34:10 frogs and it's found in dark frogs the Amazon jungle. Some of the
34:19 tribes will use these dark frogs to poison the dark tips with the poison
34:28 the frogs. And those dark tips will use to hunt other animals or
34:35 themselves from, from the intruders. it's a Ditro co toin that comes
34:42 a frog and it blocks inactivation. channel remains open. Ok. So
34:51 toxins are blockers of the channel. no flux of current. In this
34:59 , we're talking about blocking inactivation. three never happens if you don't have
35:09 activation, what happens in this If once there's depolarization it opens,
35:14 contains being it sustained, opens, opening, that's, that's not very
35:20 because you have so much excitability and in the brain. Oysters on the
35:25 shell is one of the favorite dishes in the South. Redfish on the
35:31 shell is really fantastic too. Uh have puffer fish. I'd love to
35:38 to Osaka Temple area and this is of those 3000 restaurants uh, not
35:46 big fan of going to get hit dark frogs anytime soon. But what
35:53 have is different toxins. As we when we talked about Roderick mckinnon,
35:59 have different binding sites. And remember talked about side directed muta genesis and
36:06 is because he was di di di , he was directing muta genesis and
36:15 to a specific site along these OK. Maybe you're targeting S
36:24 maybe you're targeting S five, maybe targeting the um the hairpin loop region
36:31 different substances will bind to different parts the channel. And this is an
36:36 of that how it's the same channel one toxin or two toxins tt X
36:43 will block the channel and Bacot toin keep the channel open longer. It
36:50 not open the channel by itself, it will keep the channel open
36:56 And that's because they have different binding . And this is how in the
37:01 days you could derive the three dimensional . So the three dimensional structure,
37:06 particular of these voltage gated sodium lidocaine that was in the movie that
37:14 watched uh I believe uh two lectures , we watched a little movie about
37:21 squid John T Song. And there a mention that there was some an
37:26 that was discovered and that an aesthetic used routinely every day and in the
37:32 setting. And that's li OK. that binds also has its own distinct
37:38 side. In this case, on S six, the 76 segment S
37:46 of right here of the transmembrane as of the sub units and lidocaine by
37:56 sodium channels has anesthetic properties. So quite often used in anesthesia or minor
38:06 , dental anesthesia, uh minor anesthesia, local anesthesia typically.
38:17 Now how do we probe these We already talked about how we record
38:21 channel currents if we withdraw a patch the membrane. But this is more
38:29 of how these recordings are done. of all, there are some recordings
38:32 are called cell attached recordings, other . If you essentially attach this electrode
38:40 a practical terms, what this looks is that you have an electrode under
38:44 microscope and you use this, what call Piazza controlled very accurate manipulator.
38:50 put it inside the cell or you it on the membrane of the cell
38:55 here and you apply very mild suctions your cell attached. And then how
39:02 you apply more suction? You actually the electrode that has solution connected to
39:08 little tube and that little tube comes and it's connected to a little syringe
39:15 an experimenter like me, an electro who take their syringe, put it
39:20 their mouth and go and when we that, you break into the cell
39:28 that is called whole cell recordings. your cytoplasm becomes continuous with the inside
39:35 the P path and that's important. then you should say like,
39:40 wait a second. So then you to have really the same composition in
39:44 pet as you have in the Because you know basic rules. If
39:48 have too much water, the cells , they shrink all of these things
39:53 happen based on the concentrations. And size of your neuron is 10 micrometers
40:00 diameter. The size of the electrode several centimeters. So what you have
40:06 when you attach your electrode and you this membrane for what we call whole
40:11 recording, you have this massive not electrical recording but this massive fluid reservoir
40:22 to the size of the neuron that's and this continuous with the cytoplasm of
40:27 neuron. This massive reservoir is this pet. It's giant compared to the
40:33 of the actual cell. Now, some instances, you apply mild suction
40:41 you say a prayer or two and have other, you know ways of
40:46 these complicated experiments. If you shake table, maybe you shake the electrode
40:53 . But if you're lucky, you excise a little patch of this
40:58 And that's what we talked about already . Then you can have channels in
41:02 patch of the membrane, you can single channel activity, you can uh
41:06 composite if you have multiple channels within patch of the membrane. And this
41:11 important from pharmacological perspective because all of sudden what you have done by withdrawing
41:20 piece of the membrane is you have not only have a piece of the
41:25 with the channels, but you have exposed this inside the cytoplasmic domain of
41:33 channel. You made that accessible. not just to air, you made
41:39 and expose it to any solution of choice, any of your drug of
41:43 choice. And that's important, certain toxins or whatnot that target receptors.
41:50 of them are somehow get inside the . So we don't know how they
41:56 . And we don't know if they on inside of the cells. We
41:59 actually know some of the substances that passed through the plasma B, we
42:04 know if they're lipid soluble. So don't know if they're gonna act on
42:08 outside of the channel somewhere inside the , on the, on the,
42:13 the cytoplasmic side, on the inner or the extracellular side. And so
42:19 allows you now to expose the cytoplasmic of the channel to different substances to
42:25 chemicals. And this is a very pharmacological neurop pharmacological technique that is applied
42:31 used because if you have a substance is hard to cross through the
42:37 but let's say it does to a small extent. But you want to
42:40 once it crosses what effect it has good way to do that is to
42:45 a lot of that substance or and that by exposing that substance to the
42:51 of the channel by using this inside reporting it's inside out because the inside
42:58 the channel is exposed to the outside . And this recording is another
43:05 So we drive the electrode there, uh suction on mildly onto the neurons
43:14 then we retract and we apply more on the pipe hat. And what
43:23 is that you get a patch of membrane out. And as that patch
43:28 a membrane is sitting in the high , you break that patch of a
43:33 and it's possible lipid bilayer. So happens is that if you're lucky
43:37 you do whatever rituals you do to this, you get your memory to
43:44 again. And the way we and is in this case, you have
43:49 outside part of the channel exposed to choice of solution or chemicals or
43:57 And that you could study to, be more effective and also to isolate
44:02 out of the system. Now, have isolated channels of interest, you
44:07 voltage clamp. So you can isolate currents. That's what voltage clamp
44:12 You have toxins. You can block currents and understand how they affect action
44:18 or flux through sodium and potassium And uh you can study various pharmacological
44:26 to see their effect on the cytoplasmic the extracellular domain of of of these
44:34 of interest or in this case, would be receptor channels. Uh if
44:39 applying certain agonist or antagonists uh or gated channels because they will also have
44:45 sides to what we talked about agonist antagonist also. So action potential gets
44:52 in the axon initial segment. Here have a spike initiation zone. For
44:58 most part, we will be talking neurons for the sensory neuron. The
45:02 initiation zone is different. It happens the sensory nerve ending. If you
45:08 that sensory neurons is also to gain inside, which is gonna be on
45:12 exam pseudo in the cell, it's be in the exam has a peripheral
45:17 that runs into the muscle fibers and and has the central axon that communicates
45:23 sensory information through the dorsal part of spinal cord. And neurotransmitters releases,
45:30 know, and then you also have cells that's more classical kind of a
45:36 initiation spike initiation is action potential uh zone in the axon initial segment.
45:45 these are shown as membranes with high of voltage, voltage gated sodium
45:53 So that tells you that most of voltage gated sodium channels will be expressed
45:59 that axon there surely will be some the soma and dendrites, but they
46:04 be dominating the axon and the and and the densities and numbers of the
46:11 . So once this action potential gets here, OK. How does it
46:18 generated, you know how it gets . You have excitatory input that comes
46:27 . So you have the cell that sitting at mine was 65 millivolts,
46:36 , minus 65 millimal. This is resting number of the 10. Then
46:43 said the cell will depolarize and it hyperpolarize and linger around until it reaches
46:49 threshold for action potential generation. So does it get to this threshold?
46:56 already talked about it. It gets this threshold because remember, the cell
47:01 be having excitatory and inhibitory synopsis. the excitatory glutamate synopsis will depolarize the
47:10 in the absence of another neuron. could be a stimulus like a visual
47:14 , auditory stimulus, touch stimulus that that depolarization, initial depolarization stimulus,
47:23 ? It could be intrinsic signals, repetitive stimulus that we have circuits in
47:29 brain that don't need a stimulus. just once like you have the circuit
47:34 your heart, it's once it's it doesn't really stop. If it
47:42 it's all good, right? So is the, this is a cycle
47:46 the heart will beat to, to, to, to,
47:50 to, to, to all of life. Nothing is stimulating it.
47:55 has this uh rhythm generation on its . So some neurons will have this
48:04 intrinsic ethnicity of active states and inactive . Uh Of course, activity can
48:11 influenced by chemicals too. We talked how if you increase extracellular potassium,
48:17 number in potential is potentially gonna go to the threshold to action potential.
48:23 this depolarization can happen due to synaptic or it can happen due to chemical
48:31 such as potassium extracellular concentration goes up changes the number of potential. Other
48:40 , you can have some toxins or things that also influence and open the
48:45 channels. For example, depolarize the . So once this information is received
48:51 the cell, what the cell does , these cells can receive sometimes tens
48:56 thousands and even hundreds of thousands of , excitatory and inhibitory and different ratios
49:03 different numbers. The cell has to all of that information. It has
49:10 calculate. Did I receive enough of excited input? Or am I getting
49:17 polarized a hyper polarizing input that's pushing further away from firing an action
49:23 If I get depolarized and not from synaptic inputs or what we just talked
49:28 chemical changes, I will then produce massive action potential, which I'm gonna
49:34 here. We're gonna return here with potential. It's all or none of
49:39 . So the synaptic inputs, these will be graded, some of them
49:45 be smaller, some of them can larger, both directions, depolarizing,
49:50 polarizing, it's degraded input sensory inputs synaptic inputs are graded. These depolarizations
49:56 graded versus the action potential which is or none. Because once you reach
50:03 threshold value, what happens? Voltage slides up and opens all of the
50:10 channels. OK. So you activated channels and this is all or non
50:16 , you cannot stop it. You drive the number into minus 30 0
50:20 come back to minus 60. Once pass that threshold, you open sodium
50:26 , floodgates are open. Boom, initiated a spike line. Yeah.
50:31 this spike once it gets initiated, gets initiated by very high density of
50:37 gated sodium channels that will be found the Axion initial segment here. And
50:42 will also have voltage gated potassium And they say segments, this action
50:47 will in what we call orthodromic So this is what we discussed,
50:54 Kal, we talked about Ramon Kal up with this principle of dynamic polarization
50:59 he said that all inputs came in from dendrites, they get processed by
51:03 sound and he drew axels. He these are the outputs going in that
51:08 . So this was his uh principle dynamic polarization. There are poles,
51:12 receding Pope there. Output poem and the flux of information is in this
51:17 . OK. So this is referred as orthodromic signal. Once you've produced
51:23 action potential act in the initial it will travel through this insulated
51:32 It's insulated with my and CNS, will even sides. And in each
51:39 of wrong beer was is shown here the same for gated sodium and potassium
51:45 . Each nodes of wrong beer. have high densities. Again, of
51:51 same voltage gated sodium and potassium channels will regenerate or reproduce the action potential
51:59 each note of brown deer. So by the time and remember some axons
52:05 be very long. Ok. Think Dwight Howard. Think about any of
52:11 seven plus feet guides that have axons run from their spine into their
52:19 That's a far distance. So what does is it assures that it travels
52:25 distance fast in an insulated fashion. by the time it reaches the terminal
52:30 of the axon, it produces the amplitude and shape depolarization, the same
52:38 action potential at the terminal end as produced at the axon initial segment.
52:45 so this movement of the action potential what we call forward propagating or or
52:51 drama. But what we'll discuss is there are also not cases and not
52:58 experimental but also cases of information that from Axion visual segment back into the
53:05 and back into the development. And is referred to as an dr.
53:10 you can have experimental stimulation of the to induce the flow of the signal
53:16 the opposite direction to what Raonic Cajal . But we do have some
53:22 It's not exactly as Raonic Cajal thought it was uh dynamic polarization in one
53:28 , but rather we have a little of bipolar information flux here from one
53:33 to another with a major action potential reaches here is responsible for neurotransmitter release
53:40 depolarization and we'll talk about it in second section of the class. And
53:45 you take a midterm, we'll see this week, you'll have the whole
53:49 section material of the class available to . And we'll start talking about the
53:54 between neurons. And so this this of a flow of information is forward
54:00 as a drama. And this kind information is that propagating that they gaming
54:08 anti drama. OK. Typical conduction . Oh Sorry, this is or
54:14 ro this is anti draw anti in opposite direction. That's how the easiest
54:21 remember. Typical conduction velocity, 10 a second cast, uh typical life
54:27 action potential. Uh Well, two uh that's the typical duration of the
54:37 potential. That's not the typical length an ax to duration of about two
54:41 or so, can vary between one or less even to few milliseconds in
54:49 in muscles of cardiac tissue. Those potentials are much, much longer on
54:54 order of hundreds of milliseconds with neurons the fastest tissues. OK. Propagation
55:02 the action potential. Remember that these the factors that will influence propagation.
55:08 myelin or insulation. Uh you have of myelin that is wrapped around and
55:14 preserves the charge until it allows it break out essentially and reproduce itself with
55:20 node of round deer have oligodendrocytes and Schwan sauce and PNS. This type
55:28 a conduction jumping and regenerating action potential referred to as saltatory conduction of the
55:36 potential. And it happens of nodes Ron deer. And as we
55:42 you will need the same channels to first action potential action. The initial
55:46 , we will need the expression of same channels of each node of Ron
55:50 in order to reproduce that action So this this this explanation, experimental
55:58 and theoretical explanation came out. Now 15 years ago, it was written
56:04 two of my colleagues and one of friends, Doctor Chris Dula, who
56:09 a professor of Neuroscience at Tufts University Boston. And Chris and John Huard
56:17 propose the mechanism. How can you action potentials one moving forward and one
56:24 propagating. So they came up with explanation that was also based on the
56:31 of course. And they published an called who let the spikes out and
56:39 they explained. And what they saw is that if you have this very
56:45 depolarization that comes in to the SOMA in the Axion initial segment, you
56:52 have two subtypes of voltage gated sodium . NAV 1.2 which are located closer
57:01 the SOMA and NAB 1.6 which are further away from the SOMA but still
57:07 that same Axion initial segment. So have higher density of NAV 1.2 here
57:14 higher density of NAV 1.61 0.2 or stands for a different subtype of voltage
57:21 sodium channel and a sodium V voltage subtype 1.2 versus 1.6. So this
57:29 depolarizing stimulus actually enters into the axon it fails to trigger N ad 1.2
57:39 1.2 is referred to as high threshold gated sodium channels, high threshold.
57:45 means they need a lot of depolarization have a high threshold. A lot
57:50 current needs to cross the threshold in for these channels to. So,
57:55 the meantime, depolarization passes through this and it hits NAV 1.6 those
58:03 voltage gated channels. And NAV 1.6 referred to as low threshold channels and
58:08 don't need as much of depolarization. so this current passes through here first
58:14 NAV 1.6 and NAV 1.6 generate the propagating action potential. And that's the
58:22 potential that propagates and regenerates and the propagating action potential is going to cause
58:29 release. And that's the function of forward propagating action potential. But what
58:35 a little bit later is that this depolarization through NAV 1.6 that produces the
58:44 propagating action potential. This depolarization that from this spike of this explosion.
58:53 explosion of the charge here is now to 1.2 channels, 1.2 channels and
59:03 still have the excitatory input and they additional depolarization from already open N A
59:13 . And now it's high enough of threshold for these channels to open NAV
59:20 they start conducting sodium. And you say, OK, then sodium is
59:24 move down the line to where the or forward propagating action potential happened and
59:30 won't because this area is very It could be at zero or plus
59:36 millivolts. But this area near the could be at minus 45 minus 50
59:44 be further away. And that provides opportunity for a small sodium charge,
59:50 charge to propagate that back propagating action , which is very small, it's
59:59 a fraction of depolarization but it travels into the Selma and into the
60:07 So you have orthodromic forward propagating spike synaptic neurotransmitter release and you have the
60:15 propagating spike that is going to be back into the SOMA and the
60:21 And at this point, your question be, why do I need
60:27 OK. Why do I need Why do I need a back propagating
60:32 what's its function? So if the propagating causes neurotransmitter release, what's the
60:38 propagating do? And the back propagating potential is very important for synaptic plasticity
60:46 what what we call binding the pre input that the cell receives? Acknowledging
60:53 there was a response from the if there is an input coming into
60:59 cell and there's no response from the or the response is taking too
61:05 that communication can be irrelevant between There's this concept of spike counting dependent
61:13 . We don't have time to get it. But timing for neurons is
61:16 everything and timing within milliseconds. That's neurons really form strong connections and communication
61:24 them. It's just like in human , you say hello to somebody and
61:29 typically expect for them to say how are you when you say hello
61:34 them? And they just look at . I don't know. How long
61:37 you look at them back, you , for five seconds, 10 seconds
61:41 kind of a lead. If somebody hello, two minutes later, you're
61:45 , what is that? Is that me, is that information relevant?
61:48 ? It's, it's, it's for , it could be minutes or hours
61:51 day sometimes like in human communication, ? For neurons within milliseconds. So
61:57 this neuron is putting all the it has to have a way that
62:03 active synapses know that the cell is . And that way of knowing is
62:08 back propagating action. So for the , what you should know is you
62:14 know that the action potential gets The axon initial segment gets propagated,
62:23 , orthodromic forward propagating cause neurotransmitter release . Propagating is involved in synaptic plasticity
62:33 cellular activity here, uh synoptic and you have two types, subtypes of
62:40 gated sodium channels. The low threshold forward propagating and high threshold produce back
62:49 spot. So it's, it's a bit of information, but the question
62:54 be too, too difficult. All . So now we're finishing up on
62:59 , we're talking about the neurons signal they produce action potential, resting memory
63:06 . And then we're gonna talk about they communicate, how they communicate with
63:12 other, how you have all of interconnections. But one very important thing
63:17 we already learned is that action potentials the code. So their frequency,
63:25 pattern is what codes neuronal activity. codes essentially your perception, sensitive
63:32 your thoughts and your motor outputs your functions and your emotions and everything else
63:40 the brain is responsible for. So we come back in the second
63:44 after midterm, we will talk about transmission, uh how neurons communicate.
63:50 before that, we are kind of going to withdraw ourselves into a larger
63:56 of the brain. Remind ourselves how brain comes about and the major parts
64:00 functions of the brain before we go network communication. Good luck on
64:08 If you have any concerns during the of the exam with technical issues,
64:14 you don't or other issues, medical , I will be available uh by
64:21 . I'll be checking the email Good luck. It shouldn't be too
64:25 uh for this midterm one. But study
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