| 00:02 | This is Lecture eight of neuroscience. where we left off last lecture was |
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| 00:11 | this very interesting structure in kinetics of voltage gated sodium channels. So if |
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| 00:19 | recall during the action potential, the that get engaged are both educated channels |
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| 00:27 | number and potential. The numbering is by leaking potassium channels. There's synaptic |
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| 00:34 | that could be coming in. So this is resting membrane potential and this |
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| 00:40 | the threshold for action potential value, resting number of potential is dominated and |
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| 00:48 | closer to equilibrium potential to potassium and cell might be receiving inhibitory inputs and |
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| 00:55 | inputs. More inhibitory inputs, more to inputs, strong inhibitory inputs, |
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| 01:02 | excitatory inputs and these hyper polarization to polarization, czar synaptic inputs that will |
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| 01:09 | discussing so that there is going to excitatory synapses causing deep polarization and neurons |
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| 01:16 | inhibitory synapses and inhibitory neurotransmitters will be hyper polarization. You recall if for |
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| 01:25 | , this number of potential DM here , reaches So that means that there's |
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| 01:33 | enough deep polarization. That means that strong enough inputs into the south, |
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| 01:38 | will activate all the non response and potential. What we discussed is that |
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| 01:46 | the rising phase this activation here, this level of the threshold fraction potentially |
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| 01:53 | voltage gated sodium channels and the more channels open, the more deep polarization |
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| 01:59 | is the more sodium channels open, more deep polarization there is and sodium |
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| 02:04 | trying to drive the number of potential its own equilibrium potential value then we |
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| 02:09 | it fails to do so. One the reasons is because the more member |
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| 02:14 | potential becomes closer to equilibrium potential to value, which is positive 55 million |
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| 02:20 | . The smaller the driving force becomes that eye. On on the other |
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| 02:25 | we said it has something to do the way this channel is built and |
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| 02:30 | kinetics of this channel. So when talked about voltage plant that was used |
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| 02:40 | Hodgkin and Huxley, we said that voltage clamp recordings we saw an inward |
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| 02:47 | followed by an outward current that the current was dominating the engine initial |
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| 02:53 | Polarizing phase of the action conference in outward current which is potassium inwards 17 |
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| 02:59 | potassium dominating the late phase of action . So if you recall, Hodgkin |
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| 03:06 | Huxley used the voltage clamp in order clamp the member and potential and different |
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| 03:13 | command potential values in order to isolate record these individual inward sodium outward potassium |
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| 03:23 | and describe what is happening during the potential. So once you reach the |
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| 03:30 | value, you open up the sodium with the sodium channels very quickly |
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| 03:36 | So the inward conductance and inward currents transient. They're fast activating the transient |
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| 03:45 | this you can see the potassium channel opening up, opening up and they |
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| 03:51 | prolonged conductance is compared to the sodium . The reason why sodium channels have |
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| 04:00 | short conductance is is because they have gates. These are the gates and |
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| 04:05 | voltage gates. That means that these are sensitive or gated by voltage and |
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| 04:12 | gates will open depending on the changing voltage. And so what's happening in |
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| 04:25 | happening in these voltage gated sodium channels that we discussed that they have six |
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| 04:34 | membrane subunits and S4 is the voltage . And as you recall, the |
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| 04:41 | of the cell relative to the outside the south is negatively charged and the |
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| 04:47 | of the cell is positively charged. so these positively charged amino acid residues |
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| 04:53 | are within the voltage sensor are actually to the negative charge and are repelled |
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| 05:00 | this positive charge. But once the deep polarization builds up as we were |
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| 05:07 | about, what happens is that once is this deep polarization, right at |
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| 05:13 | threshold level, before the action potential begins, you have basically this threshold |
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| 05:21 | build up of positive charge now on inside of the membrane and more of |
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| 05:27 | negative charge build up on the outside the membrane and now this positive charge |
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| 05:34 | repelling the voltage sensor. And this sensor literally slides up within the structure |
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| 05:43 | the protium causes conformational change causes the of the channel that allows for the |
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| 05:50 | of sodium ions. And so it that voltage gated sodium channels have two |
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| 05:59 | . One of these are called activation and another one is called inactivation |
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| 06:05 | So when the cell number and potential D polarized passed the threshold level from |
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| 06:12 | number and potential about minus 65 to 40 million bowls here you de polarize |
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| 06:18 | cell. That means you have passed threshold of action for action potential. |
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| 06:23 | have started opening voltage gated sodium channels you see that the opening is |
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| 06:30 | But this deep polarization here sustained on . However sodium channels one after |
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| 06:38 | they open open open open within 13 and they closed. And that's because |
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| 06:45 | that voltage sensor slides up the channel that voltage sensor slides up the channel |
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| 06:53 | opens these gates. Okay opens these . This ball and chain mechanism which |
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| 07:01 | in activation gate swings to close this so one arm moves away and the |
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| 07:08 | arm moves in one arm year moves , opens the channel and this ball |
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| 07:18 | the chain swings and closes in activates channel. So in this situation the |
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| 07:24 | is inactivated. Now in order for channel to be open again, you |
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| 07:32 | have to d enacted it. So have to d inactivation means you have |
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| 07:38 | remove inactivation gates. Deon activate. as you deign activate and the voltage |
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| 07:49 | slides down back in the protium caused closure of the channel again. To |
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| 07:55 | that. You have to have hyper Because if you have hyper polarization then |
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| 08:04 | gonna have build up of negative charge the inside again. Once again pushing |
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| 08:11 | voltage sensor to stay in this position caused the closure of the gates. |
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| 08:19 | you cannot go 1231. Because from uh you have to you cannot go |
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| 08:28 | close from inactivated. You have to to Deion activated. You have to |
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| 08:33 | it then closed and when you're closed you're open. And as soon as |
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| 08:37 | open you get inactivated. So you to hyper polarize the member and remove |
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| 08:44 | to reposition the voltage sensor again to it was before the deep polarization. |
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| 08:54 | is during the polarization, reposition it in the protein to what it was |
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| 08:59 | the re polarization. And that again conformational change allows for the protein gates |
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| 09:07 | . So this is another reason why ions tried to drive the overall membrane |
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| 09:16 | value to the equilibrium potential value for . And when I speak of |
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| 09:21 | I want to remind you of This , not this one. This particular |
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| 09:31 | that you have. What we're talking the equilibrium potentials. We're talking about |
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| 09:37 | potential and we're talking about driving forces the difference between the membrane potential and |
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| 09:43 | equilibrium potential for the ion. so in this case, what you |
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| 09:51 | here is you have basically sodium driving the influx stays the overall VM toward |
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| 10:04 | E. N. A. Which a quite liberal potential but is failing |
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| 10:09 | do so despite this positive feedback loop polarization. More sodium decolonization. More |
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| 10:15 | deep polarization. More sodium for two . Once the number of potential is |
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| 10:20 | polarized, the driving force for sodium not that great. And also the |
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| 10:28 | channels open up and they very close . Close get inactivated. So that's |
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| 10:35 | second reason why it never reaches that potential value for sodium. And uh |
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| 10:44 | have the at that point you have channel inactivation. Sell Okay, so |
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| 11:02 | , the structure of the voltage gated channel, the four subunit six trans |
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| 11:08 | segments. As for voltage sensor, poor lupus selectivity filter. Now, |
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| 11:13 | do we how do we do these of recordings where we can record from |
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| 11:18 | channels or how we can record from selves? And we're gonna talk for |
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| 11:23 | few minutes. Next about the techniques are commonly used to study action potential |
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| 11:30 | in modern day and electrophysiology. So of inserting very large electrodes inside very |
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| 11:37 | accents, we can target small We use very small electrodes. We |
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| 11:44 | target specific areas of the neuron dendrite axon specific locations because we can |
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| 11:53 | the cells pretty well using infrared we can isolate little patches of the |
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| 11:59 | with these micro electrodes with these Thats what they call pipettes or micro |
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| 12:05 | because you will be using electrical current electrical current through them. And so |
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| 12:13 | pipettes can record from small patches of membrane and they can even isolate activities |
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| 12:19 | single channels. And that's why we're of recording activity through single channel |
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| 12:26 | So we can also use other different to isolate the currents apart from uh |
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| 12:33 | clamp. And in particular pharmacology and recordings that you can do with typically |
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| 12:42 | is called the patch clamp are several ways of which you can record. |
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| 12:49 | if you look first of all, you bring the pipette to neuron on |
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| 12:54 | top, it says if you apply suction. So it's really just just |
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| 13:02 | suctioning onto the plasma membrane and forming is called the tight contact between type |
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| 13:10 | . And membrane. And this kind a configuration is called cell attached configuration |
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| 13:17 | cell attached recording. Now it's it's interesting way to record activity through the |
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| 13:26 | . Uh but it's somewhat limited but is one way in which you can |
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| 13:31 | activity thing to sell the more common . Once you form that cell attached |
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| 13:41 | , you produce a much faster and suction that ruptures of plasma numbers. |
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| 13:50 | now the cida plaza, the south this neuron becomes continuous with the interior |
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| 13:57 | the pipe at. And so it's important that whatever you have inside the |
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| 14:03 | that we call the pipette solution, journal solution matches exactly what we now |
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| 14:08 | inside the cells because these neurons are 10 nm. Uh sorry 10 micrometers |
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| 14:17 | diameter. And these pipe pads you know, their tips are one |
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| 14:23 | meter in diameter. But the size the whole pie pad is that you |
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| 14:27 | take it with your fingers and that's the order of interest which means it's |
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| 14:33 | reservoirs. If you have too much the acquis like water in your solution |
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| 14:40 | , you would basically ruin the You had too concentrated of the ions |
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| 14:45 | your solution. It also would not good for the south. But once |
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| 14:50 | have a whole self configuration you can all of the currents or whole cell |
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| 14:56 | of approach into the cell activity Now, instead of rupturing the |
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| 15:03 | you can also after forming this cell mode. Instead of rupturing the |
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| 15:09 | you can try to shake it off little bit and draw it out to |
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| 15:13 | electrode. That allows you to isolate axis of the membrane that will contain |
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| 15:20 | channels of interest receptor channels of And in this kind of a configuration |
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| 15:29 | you have done is you have exposed inside cytoplasmic part of this protein to |
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| 15:39 | outside environment which is your experimental So for the substances for example that |
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| 15:49 | pass through plasma membrane and they may molecules on the inside pharmacological you may |
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| 15:57 | to apply these molecules and see what they have on conductance is if they |
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| 16:01 | be found inside the cells of certain and how they would affect it because |
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| 16:07 | a great way to test any molecules could be potentially binding on the side |
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| 16:13 | plas mix side of this channel, if their membrane and soluble and membrane |
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| 16:21 | . So that way you can test a fact molecules would have on the |
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| 16:27 | because you're exposing the inside outside experimental in another way and configuration is once |
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| 16:37 | draw this patch of the membrane then you apply very fast and strong |
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| 16:46 | , rupture the membrane and allow for membrane to Rien il. And once |
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| 16:53 | re enables itself, you're lucky will the outside of the program exposed to |
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| 17:01 | outside environment. Now you have very uh small piece of the membrane with |
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| 17:08 | channels of interest and you're studying the that would be not passing through the |
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| 17:14 | membrane but rather binding on the outside this channel and seeing how binding of |
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| 17:20 | substances to this channel are going to the flux of ions through this |
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| 17:25 | Okay, so this configuration was called out in this case the inside of |
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| 17:33 | protea um is exposed to the outside . And here the outside extra cellular |
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| 17:40 | of this podium is exposed to the environment. And so these are all |
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| 17:46 | and common configurations that will allow us use voltage clamp that will allow us |
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| 17:53 | record single channel activity, whole cell and uh that's how we know and |
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| 18:02 | more and more about the brain every . These are still techniques that are |
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| 18:06 | every day in neurophysiology, electrophysiology So patch clamp technique and voltage clamp |
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| 18:17 | . Patch clamp was developed a little later. But if you recall uh |
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| 18:25 | clamp became a very important tool in individual ionic conductance is and studying individual |
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| 18:33 | conductance is like sodium n word potassium . And so uh we're gonna talk |
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| 18:44 | dr Toshio Narahashi in the second but we're gonna watch a little bit of |
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| 18:52 | so you have to also um forgive of the um if if you know |
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| 19:03 | Simpsons, the Simpsons can be offensive everybody and themselves at some point during |
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| 19:08 | series. And uh so don't take of this in any way personally or |
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| 19:15 | racially or ethnically because we can watch episode and everybody gets gets their |
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| 19:25 | Oh she's here for me. Not if it is cut in property it's |
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| 19:33 | . Yes it is poisonous potentially But if sliced properly it can be |
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| 19:38 | tasty. I must get the Oh miss crabapple master you are needed |
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| 19:51 | the kitchen. I said cover for . But master we need your skilled |
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| 19:57 | . My skilled hands are busy. do it poison poison. Tasty fish |
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| 20:13 | , concentrate mm. Fan fugu Oh beautiful language isn't God's sake. |
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| 20:38 | eat another bite. Couldn't possibly Mr ? I shall be blunt. We |
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| 20:44 | reason to believe you have eaten poison . What should I do? What |
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| 20:49 | I do? Tell me quick. need to panic. There's a map |
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| 20:52 | the hospital on the back of the . Try something new homer. What'll |
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| 20:59 | hurt you? Homer? I never of a poison pork chop. Your |
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| 21:03 | agreed that I should break this to . No need. I can read |
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| 21:07 | like a book. It's good isn't it? No. Mr |
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| 21:13 | If in fact, you've consumed the of the blowfish and from what the |
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| 21:17 | has told me, it's quite probable have 24 hours to live 24 |
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| 21:24 | Well, 22 I'm sorry. I you waiting so long. I'm gonna |
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| 21:29 | . I'm gonna die. Well, there's one consolation is that you will |
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| 21:34 | no pain at all. Until sometime evening when your heart suddenly explodes |
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| 21:39 | A little death, anxiety is You can expect to go through five |
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| 21:44 | . The first is denial. No . Because I'm not dying. Second |
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| 21:47 | anger you. Little after that comes . Fear. What's after? Fear |
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| 21:53 | ? You gotta get me out of . I'll make it worth your |
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| 21:56 | finally, acceptance. Well, we gotta go sometime. Mr Simpson. |
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| 22:00 | progress astounds me. I should leave two alone. Perhaps this pamphlet will |
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| 22:05 | helpful. So you're going to But Mhm. It's here. |
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| 22:21 | C. Penney Beauty in select stores at JCP dot com slash beauty. |
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| 22:26 | the marine world has other more exotic in store. Known to hardly anyone |
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| 22:32 | the west, for example, the , the poisonous puffer fish of which |
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| 22:37 | are about 100 species worldwide. You a license to sell puffer fish in |
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| 22:43 | , but as a buyer you need too is Okamoto has a fuego restaurant |
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| 22:54 | of course a license. He's not to buy the increasingly popular nontoxic farmed |
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| 23:00 | which can be recognized by its shorter . Nor is he interested in the |
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| 23:04 | , all of these species caught in wild from japanese waters. This true |
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| 23:10 | is only looking for one thing toxic as fresh as possible and that means |
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| 23:17 | fugu, tiger puffer fish, the beef of fuego cuisine. Okay, |
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| 23:36 | single specimen of this species, which only found in the sea of Japan |
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| 23:41 | well cost €100. One of Tokyo's districts is located around Asakusa Temple. |
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| 23:57 | wild fugu restaurants are to be found . There are about 3000 restaurants specializing |
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| 24:03 | Fugu in Tokyo today. From the they're usually easy to recognize and they're |
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| 24:09 | highly specialized. One of them is Oh tomatoes restaurant where sometimes even Prime |
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| 24:19 | drop by war says its name, pure fish place, you also need |
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| 24:26 | license to prepare fugu. The poison Fugu is tetrodotoxin. It's 1000 times |
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| 24:33 | potent than cyanide and there is no , The poison paralyzes its victims but |
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| 24:40 | them fully conscious proper preparation is The skin and entrails of the fish |
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| 24:47 | poisonous and they must not contaminate the toxic meat on the muscles. High |
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| 25:03 | of highly poisonous tetrodotoxin are found in innards, especially the liver and |
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| 25:09 | Special disposal is necessary. A seasoned Chef like Okamoto San takes about 10 |
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| 25:30 | to neatly gut. A tour of commonly the fish is cut into thin |
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| 25:36 | and eaten raw as Sashimi. In the taste of unprepared puffer fish is |
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| 25:43 | , rather bland. The rose I exclusively serves its guests captured in the |
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| 25:52 | in small doses, the poison of fish triggers numbness in the mouth and |
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| 25:57 | intoxicating. But here they do not for guests who might be eager to |
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| 26:04 | out tiny doses of this poison. , What we serve here is 100% |
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| 26:18 | toxic. If we break this law ruined, let's go. A real |
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| 26:27 | meal consists of at least six. there are just three a year. |
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| 26:37 | victims of unlicensed wild is a child it always tasted very good to |
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| 26:48 | Wild fugu is a delicacy still eaten Japan despite earthquakes, tsunamis and nuclear |
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| 26:57 | . Ricky's Okamoto will not run out work and in the future too, |
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| 27:01 | likely to remain quite exhausting. So now if you think about |
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| 27:27 | you're gonna think about puffer fish. what you heard uh and you saw |
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| 27:34 | the video is that it contains a called tetrodotoxin here, it's abbreviated as |
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| 27:42 | T. T. X. And why I have mouthwatering tales of |
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| 27:47 | So what's the thrill of eating fresh boo is I guess little remnant uh |
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| 27:56 | the meat that makes it your tongue they said. So it's a little |
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| 28:00 | of a thrill seeking experience. I love to try it. You have |
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| 28:06 | have a shaft that's uh certified. think they take a number of years |
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| 28:11 | train in order to remove the organs are concentrated with tetrodotoxin. Because if |
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| 28:20 | cut it improperly it leaks into the or leaks into the meat and can |
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| 28:26 | poisonous. So the fish itself doesn't or synthesize this toxin. The fish |
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| 28:34 | that toxin. And these toxins are by microorganisms that are found in uh |
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| 28:43 | organs of these animals. And similarly toxins such as Saxon toxin which can |
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| 28:53 | found in clams mussels during what is red tide. So when the temperature |
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| 29:00 | hot and you will also experience here the Southern States that you're not recommended |
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| 29:08 | eat certain animals like oysters or certain during really hot periods of the year |
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| 29:19 | the temperature and water goes up. is a lot of algal formations and |
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| 29:25 | also a lot of microorganisms and they invade the clams and mussels and carry |
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| 29:35 | the toxin. So again it's the that will produce these toxins. It's |
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| 29:42 | that the clams or mussels or puffer synthesized these toxins themselves. They carry |
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| 29:49 | Now in 1959 the story goes back Toshio Narahashi who actually discovered what tetrodotoxin |
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| 30:01 | . And Toshio Narahashi in the fifties talking about channels and people just started |
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| 30:09 | about channels in the late fifties. is that interesting? So people didn't |
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| 30:17 | know about channels. We'll talk about roderick Mackinnon used X ray crystallography and |
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| 30:22 | was in the nineties to visualize the . And so this is a lot |
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| 30:26 | time that passes. About 30 40 of a lot of studies that happened |
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| 30:31 | the channels and uh has this toxin is isolated from puffer fish in |
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| 30:40 | And he takes a vial of that him to United States and his maybe |
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| 30:46 | pocket or something like that. So he knows is that it's paralyzing. |
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| 30:52 | know that it can block the contractions can block action potentials. But he |
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| 30:59 | know the precise mechanisms of action of substance tetrodotoxin. So he finds his |
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| 31:08 | to the United States and he uses cloud, he uses voltage clamp. |
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| 31:18 | so he At that point you have realize in the 60s early 60s we |
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| 31:26 | about how instrumentation improved and people like and Huxley started using action potential, |
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| 31:34 | doing action potential recordings. The voltage were started being to develop. So |
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| 31:38 | the 60s there were foreign feuding between voltage clamp setups were maybe a few |
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| 31:44 | the whole United States for example. it was a rare type of instrumentation |
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| 31:52 | an advanced type of experimentation at the to do. So he comes to |
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| 31:57 | United States and he uses voltage clamp as if you recall if you have |
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| 32:02 | deep polarization it will have a significant current which is sodium voltage gated sodium |
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| 32:10 | and sodium influx ng followed by potassium opening up potassium the flexing and he |
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| 32:19 | tetrodotoxin during the recording and he shows tetrodotoxin specifically blocks both educated sodium channels |
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| 32:31 | it does not affect potassium channels. that's important because if you had the |
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| 32:39 | even in Japan you had a way record action potentials and apply tetrodotoxin and |
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| 32:45 | action potentials. You don't know if just blocks voltage gated sodium channels. |
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| 32:50 | if it blocks both voltage gated sodium and potassium channels as well too. |
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| 32:56 | . So this gives you a definitive that this substance T. T. |
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| 33:02 | . Is a selective antagonist for sodium and the way that it was described |
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| 33:13 | uh in uh one of the movies I have uh you can find these |
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| 33:20 | very easily. The Simpsons episode and is that you get paralyzed. So |
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| 33:28 | was toxin and the vault educated sodium that we're discussing in neurons in the |
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| 33:34 | . There are also present in other that are present in the heart. |
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| 33:38 | educated sodium channels are present and muscle and the diaphragm and typically it's muscle |
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| 33:47 | and the diaphragm that give up with with tetrodotoxin uh and it can be |
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| 33:53 | . So in Japan they mentioned in movie there used to be a lot |
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| 33:57 | poisoning tetrodotoxin until they started regulating the around around strict licenses for the chefs |
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| 34:07 | can prepare puffer fish and serve puffer and the restaurants that have the registrations |
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| 34:12 | licensing to do so and so on . So if this is a blocker |
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| 34:18 | an antagonist antagonist, the blocker is same thing which means it closes the |
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| 34:25 | . There are also specific blockers are for outward currents for potassium currents that |
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| 34:33 | not affect one such substances cetera, , ammonium, members of ammonium or |
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| 34:41 | . E. A, which selectively potassium currents. The outward potassium codes |
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| 34:48 | does not affect the inward sodium So now we have a lot of |
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| 34:55 | at hand. We have voltage We can clamp the potential command the |
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| 35:03 | we desire with voltage plan we can individual currents and sodium currents potassium currents |
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| 35:11 | currents chloride currents focusing on sodium and mostly during the action potential. We |
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| 35:18 | use pharmacology and specific substances and specific or chemicals not necessarily even toxins. |
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| 35:27 | is a natural toxin that comes from . There's a lot of chemical agonists |
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| 35:32 | antagonists, antagonists or blockers agonists would something that opens and activists activates |
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| 35:41 | So by having now the voltage clamp the pharmacology you can block all the |
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| 35:48 | channels and just address the potassium cars use the voltage cloud and be very |
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| 35:55 | and it gets even more sophisticated. are subtypes of voltage gated sodium channels |
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| 36:00 | subtypes of voltage gated potassium channels. some of these subtypes have very selective |
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| 36:07 | specific blockers and antagonists and agonists as . So uh another toxin by atraco |
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| 36:23 | from Colombian frog, it says over or inactivation. So if you recall |
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| 36:31 | protean, the sodium protium is a dimensional structure and it has activation and |
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| 36:40 | gates in different substances may target different of this protein channel. Some toxins |
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| 36:48 | target inactivation gay. And once the gets inactivated it stays inactivated for a |
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| 36:56 | , very long time. Maybe that don't want to start getting the activation |
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| 37:03 | and the gate doesn't open very It needs a lot more deep polarization |
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| 37:09 | past minus 45 minus 40 maybe minus to start opening. So it's it's |
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| 37:16 | maybe the the toxin or another substances affecting that gate. So you can |
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| 37:25 | activate channels to so you can over channels and that means that the channel |
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| 37:34 | channels will stay open for a long action potential is going to become very |
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| 37:39 | and the cells are going to get over time because that is a normal |
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| 37:45 | for them to stay in. So you recall roderick Mackinnon used different toxins |
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| 37:54 | deduce three dimensional protein structure. And is because different substances will bind to |
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| 38:02 | parts of these programs. Having pharmacological like that allows us to study individual |
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| 38:13 | currents, their dynamics, their reversals equilibrium potentials. And we have to |
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| 38:20 | ourselves that nature's potent. So some these toxins for um substances that come |
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| 38:31 | bites or venoms like snake venoms for can be quite powerful. Uh So |
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| 38:40 | just learned recently on the radio that supposed to call the poison control center |
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| 38:46 | you get bit by like a spider snake out here and the nature on |
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| 38:54 | gulf coast and the marshes. There's lot of different critters and creatures. |
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| 38:59 | that's something to keep in mind. don't have the number but uh You |
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| 39:05 | didn't know that who to call if get bit by a snake you know |
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| 39:09 | it's it's poison control and there is number for for Texas poison control. |
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| 39:13 | understand they know that uh creatures are what to do. Uh calling 911 |
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| 39:22 | ambulance may not be as useful. tetrodotoxin we have tetra cell ammonium, |
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| 39:31 | have cocaine, cocaine is also going buy the voltage gated sodium channels. |
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| 39:37 | also antagonist channels. So this tells that this is something that's the nature |
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| 39:44 | popular fish. It's also can be in newts and frogs. So |
|
| 39:48 | T. X. Is not just to puffer fish because it comes from |
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| 39:55 | and cocaine which comes from a plant also bind to multiple gated civilian |
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| 40:04 | And it is a illicit substance. Saxon toxin comes from Dina flag allies |
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| 40:14 | red tide. As we discussed. you will hear that don't eat clams |
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| 40:20 | maybe paralytic clams And Northeast especially with and child fish are very popular in |
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| 40:26 | summer months. They may advise you consuming them. And then of course |
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| 40:31 | have also uh potassium specific uh low tetra methyl ammonium antagonists. Uh low |
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| 40:43 | versus high affinity affinity means uh if there's low affinity for ammonium that means |
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| 40:52 | need very little of that substance to an effect on the channel. Yes |
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| 41:05 | and uh what's on it seems counter the stimulant channel and companies doing this |
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| 41:18 | a stimulus. So you think that would be. Yes but it would |
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| 41:23 | to specific subtypes of both educated sodium in the specific areas of the |
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| 41:28 | And there would be other neurotransmitters and modulators such as dopamine involved. That |
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| 41:33 | be doing uh different things also. it's uh it's but it's a known |
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| 41:42 | and in fact cocaine was also used a local anesthetic because it blocks firing |
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| 41:51 | certain voltage gated sodium channels. So certain places and it can affect the |
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| 41:58 | rate very much so also by affecting voltage gated sodium channels. Let's see |
|
| 42:05 | is for example lidocaine lidocaine will buy a binding site Right here you can |
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| 42:14 | Lidocaine has a binding site on six trans membrane segment. So little |
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| 42:22 | will also block sodium channels, block potentials and block the perception of |
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| 42:32 | Right? So but uh now As mentioned cocaine was also in the |
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| 42:41 | Used as a as a local around . So these three dimensional protein structures |
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| 42:47 | have many different binding sites to lidocaine is a local anesthetic and you |
|
| 42:55 | find it in C. B. . And you will find it |
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| 42:59 | Okay we'll go to dentist office and will do a root canal. You'll |
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| 43:05 | it there too. So now okay you recall we talked about the ivy |
|
| 43:13 | or current voltage plots. So each actually has its own current voltage |
|
| 43:25 | And different cells have the overall current plot. So these I. |
|
| 43:29 | Plots, we now know that ionic is dependent on the potential difference across |
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| 43:36 | number. We're talking about ions, flux of ions. The direction and |
|
| 43:42 | strength will depend on the member in . So for example at this number |
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| 43:49 | potential you have strong inward current zero balls. That positive 52 million |
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| 43:57 | You have no inward current and no current at all. So the ionic |
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| 44:03 | is the strength of those conducting The direction of this conductance. This |
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| 44:07 | dependent on the member in potential And the changes in this member in |
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| 44:12 | will also affect the strength and maybe the direction of these currents. |
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| 44:20 | so if you have a voltage clamp you can clamp the potential that your |
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| 44:28 | values minus 15 minus 25. 0 50. You can record currents through |
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| 44:37 | through receptor channels or wholesale full sell . And so this for example shows |
|
| 44:45 | you have this current flexing here at million balls cost ratios, current flux |
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| 44:52 | here -25. This current reduces to million goals. What happens to zero |
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| 44:57 | lights? zero move also has no at all. So Positive 25, |
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| 45:06 | happens in this current? It changes direction is not in the opposite direction |
|
| 45:12 | the further away it is from zero larger amount that you can get. |
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| 45:18 | we already talked about that, I that you can do these injections into |
|
| 45:22 | south with instrumentation the square wave like talked about resistant capacity to properties and |
|
| 45:28 | membrane response of the south. And talked about these I. D. |
|
| 45:34 | . So if you had a situation for each basically change in membrane potential |
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| 45:45 | 25 million balls change you always in direction or the other direction, positive |
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| 45:52 | negative change always cause the change same in the current in PICO amperes. |
|
| 46:02 | becomes a linear plot or on that . So it's a linear relationship between |
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| 46:11 | and the current and the reversal or potential. We studied equilibrium potentials. |
|
| 46:18 | when you talk about channels, certain , maybe single ion conductance is and |
|
| 46:27 | will represent equilibrium potentials for those And other receptor channels you learn later |
|
| 46:34 | of course may conduct sodium and potassium even calcium. And so they're reversal |
|
| 46:44 | values are gonna be influenced not just one, will not be just selected |
|
| 46:48 | one, but maybe by two. they may show a reversal of these |
|
| 46:54 | at zero million volts. Now this for acetylcholine. So this actually shows |
|
| 47:02 | acetylcholine receptor channel which is going to sodium and potassium will have the reversal |
|
| 47:10 | . That zero million volts here. come back to this diagram in the |
|
| 47:15 | . In reality each channel Both educated channels that we're talking about, potassium |
|
| 47:24 | and single neuron may contain 12-15 different of voltage gated channels to three sodium |
|
| 47:36 | three potassium to three calcium. so they will contain a variety of |
|
| 47:45 | volt educated channels and a lot of channels will not have the atomic or |
|
| 47:55 | curves ivy currents. Instead they would the nonlinear curren's. You recall the |
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| 48:06 | versus outward. Okay, maybe we do the following thing. I can |
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| 48:13 | the screen a little drawing. So is negative. 100 millionV -50. |
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| 48:35 | a year. Just positive 15, , Positive 100 million balls. And |
|
| 48:50 | example, what is the negative This is paris or current? And |
|
| 49:06 | negative value. So this is negative in Paris. This is by convention |
|
| 49:16 | inward current. Okay, inward current positive charge, moving inside the |
|
| 49:26 | So on this side we have inward on this side, we have outward |
|
| 49:46 | . Okay, this is the nana this is our high current access on |
|
| 49:53 | line. So let me just put million bowls here, for example |
|
| 50:04 | And let me draw something like this ask you a question. This is |
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| 50:15 | current here And there is no current -80. And my question is, |
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| 50:24 | you think that this is a representation the ideal relationship for potassium or |
|
| 50:36 | Mhm. Let me ask you another . So, now you see that |
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| 50:55 | 55 little balls. This is zero . Right? Everybody agrees that current |
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| 51:03 | zero value here. Right, on Y axis. Yeah. So which |
|
| 51:12 | is this is this sodium or is potassium? Why is that sodium |
|
| 51:26 | positive 55 is equilibrium potential for Therefore, there is no net flux |
|
| 51:33 | sodium channel. Right? So it's current. zero net current. And |
|
| 51:40 | that's for potassium. Yeah. So is going out here, right, |
|
| 51:50 | is out. It's the flexing, ? And if these potentials you can |
|
| 51:59 | this is this is this is resting of potential right here, resting number |
|
| 52:03 | potential is -65 million volts. And what is sodium doing at at |
|
| 52:11 | threshold potential here -45 threshold potential is going to go inside the cell when |
|
| 52:20 | sodium channels open? Yes. So charge going inside the cell is inward |
|
| 52:28 | it reverses until sodium reverses Right here about positive 52 positive 55 goals. |
|
| 52:47 | the other thing to note is Visa or linear channels and certain channels prefer |
|
| 52:58 | conduct outwardly. You can see that the same change here. So this |
|
| 53:04 | 50 million ball change. This is million balls. And this is positive |
|
| 53:12 | millones. You can see that this under the current. It's much bigger |
|
| 53:24 | the current is much higher in the direction. Alright, so that means |
|
| 53:33 | this channel preferring to conduct or the rectification ions have a preferred direction of |
|
| 53:45 | . So, these ions are pathetically ions we're looking here will be conducting |
|
| 53:52 | outwardly than inwardly. Maybe it's a of multi educated uh or ligand gated |
|
| 54:05 | that conduct sodium and potassium in but it prefers to conduct in the |
|
| 54:10 | direction. So, we'll talk about channels for example, in glutamate |
|
| 54:15 | you have glutamate, the separate channels will be rectifying. Okay, do |
|
| 54:23 | remember that? We talked about the electric behavior up the cells and we |
|
| 54:34 | that some neurons will fire pretty constant slow patterns of action potentials. Some |
|
| 54:46 | will produce very fast frequencies of action . And these were our excitatory cells |
|
| 54:53 | we discuss. And we said that variety in the frequency and the patterns |
|
| 55:00 | firing comes from the inhibitory cells Right. We said that that is |
|
| 55:07 | happening. The different dialects come from , inhibitory cell subtypes. So how |
|
| 55:20 | these different cell subtypes are capable of these different patterns of action potentials? |
|
| 55:27 | the action potential is both educated sodium potassium channels. How come some are |
|
| 55:32 | of firing so fast and they must a very short relative refractory period following |
|
| 55:40 | action potential and others cannot fire that . How come? Some have continuous |
|
| 55:46 | frequencies on the dialect that's continuous and are in burst or delayed or |
|
| 55:55 | How does this come about? It about because each cell subtype, one |
|
| 56:02 | subtype will express a subtype of voltage channels that will have these kinds of |
|
| 56:10 | for example. And these are gonna the 12 channels that are expressed by |
|
| 56:16 | self. And these are gonna be other cell that's gonna look something like |
|
| 56:22 | for example a little bit different, gonna have something like this. And |
|
| 56:30 | these cells will have different subtypes of and because these channels will have different |
|
| 56:41 | kinetics and therefore ivy flocks, some them linear are those knobs. These |
|
| 56:48 | are built to be able to produce patterns of action potential. Remember |
|
| 56:55 | What makes a different subtypes of neurons the fact that they express a slightly |
|
| 57:01 | subset of molecules I. E. is 12 different subtypes of voltage gated |
|
| 57:07 | channels. This suddenly express 1 to . This sound expressed subtype 267. |
|
| 57:15 | one they express 77 78 or So now the conductance of course we |
|
| 57:27 | depends on numbering potential and ionic concentration . But so you can start envisioning |
|
| 57:34 | cells will express different subtypes of local sodium channels. Those local gated sodium |
|
| 57:40 | will have their own I. Properties and these I. D. |
|
| 57:46 | will determine the dialect in which these can speak and produces frequencies and patterns |
|
| 57:53 | the action potentials especially in the inhibitory because that's the diversity in the behavior |
|
| 58:02 | also in the processing complexity comes traumas the inhibitory cells. Okay this is |
|
| 58:10 | a few slides left. Uh The we're probably going to finish everything |
|
| 58:16 | This is very short to note that when you're looking to study something in |
|
| 58:23 | they're very small. You have to sophisticated microscopes and microscopy, small pie |
|
| 58:29 | , instrumentation is expensive and you have patch of the membrane and you maybe |
|
| 58:34 | one or two channels and there are few channels. It means your signal |
|
| 58:39 | week. So sometimes you cannot determine those recordings where they're like oh I |
|
| 58:45 | captured it. Is it really doing ? So you may want to go |
|
| 58:49 | amplify that. And you would use systems where you would express, for |
|
| 58:55 | single channels and over express them in choose sides and frog eggs. So |
|
| 59:01 | you have a lot of these channels and dominating in this egg. This |
|
| 59:06 | is one millimeter mountain micro meter. you can use large pie pads. |
|
| 59:11 | don't need microscopes to record currents and you can study for example, potassium |
|
| 59:18 | in a very simple system here in frog blue side. Once you determine |
|
| 59:23 | of the kinetics and dynamics of this expressed target channel in which in this |
|
| 59:29 | potassium channel now you can go back that patch of the membrane where your |
|
| 59:36 | is weaker and the system is more , you have other channels involved in |
|
| 59:42 | patch of the membrane. And now can very clearly sort based on your |
|
| 59:47 | of the kinetics and the dynamics of current through specific channel. You can |
|
| 59:51 | use that apply that knowledge to isolate the currents and more complex systems and |
|
| 59:57 | patches of neuronal membranes. Okay, what's happening in neurons as we studied |
|
| 60:06 | neurons will receive excitatory inputs that are be d polarizing inhibitory inputs. And |
|
| 60:13 | are going to make a decision whether going to produce an action potential. |
|
| 60:20 | and if they produce an action potential going to get produced in the excellent |
|
| 60:25 | segments and it's going to get regenerated each note of run beer. So |
|
| 60:32 | initial segments will be loaded just with types of channels, voltage gated, |
|
| 60:38 | densities of voltage gated sodium channels and gated potassium channels. And so knows |
|
| 60:44 | wrong beer is where action potentials are to regenerate as they travel down the |
|
| 60:52 | of the accent, which could be lengthy and at the terminal location there |
|
| 60:57 | the excellent terminal the amplitude of that potential is preserved the same as it |
|
| 61:04 | at the axon initial second. from the early drawings of ramon |
|
| 61:10 | he talked about how he thought that dendrites and Soma were receiving the information |
|
| 61:16 | that all of the information was going the Soma through the axon into Jason |
|
| 61:22 | . So we called this and he this the principle of dynamic polarization, |
|
| 61:28 | there's only one polarity and this is input processing and there's output and nothing |
|
| 61:35 | back from this axon back into the . Uh This is again illustration of |
|
| 61:43 | of ranveer this uh principle of dynamic has been challenged in the last couple |
|
| 61:51 | decades. Now, action potentials will produced and they will get produced in |
|
| 61:58 | areas because these areas will have very densities and there will be very sensitive |
|
| 62:03 | deep polarization. So the threshold and much deep polarization you need an excellent |
|
| 62:10 | segment. Maybe very little that you in order to generate action potential |
|
| 62:15 | But if you were communicating to distill right, you would need to have |
|
| 62:22 | very strong input and you would need have multiple excitatory input so that that |
|
| 62:28 | would travel into the acts of initial and stimulate the action initial segment to |
|
| 62:37 | the action potential. So the closer are to soma, the closer you |
|
| 62:43 | to the Acts of the initial the more influence you have on the |
|
| 62:49 | properties of the cell. You can whether the cells fire action potential. |
|
| 62:54 | your inhibition you can block it. you have a citation you can easily |
|
| 62:58 | this area with just a little bit deep polarization and producing action potential and |
|
| 63:05 | your threshold will have to be on deep polarization. Will have to be |
|
| 63:08 | high because part of it is going be lost. These are nonviolent dated |
|
| 63:13 | insulated processes that are going to be and these inputs are going to be |
|
| 63:18 | part lost before they reach that accident segment. So strategically to control the |
|
| 63:25 | properties and firing properties, output properties this neuro cells and the synapses would |
|
| 63:32 | to be located disclosed to the soma these paris somatic and these accident initial |
|
| 63:38 | regions. So how the uh principle dynamic polarization has been challenged was through |
|
| 63:49 | discovery that there are two subtypes of gated sodium channels in the axon initial |
|
| 63:56 | where this action potential gets produced. is N. A. D |
|
| 64:03 | So the nominal culture is N. . Is sodium V. Is |
|
| 64:08 | So that tells me it's a voltage sodium channel. 1.2 is a |
|
| 64:14 | As I mentioned, there are multiple of voltage gated sodium channels, potassium |
|
| 64:19 | other calcium channels. And this cell the accident initial segment expresses to N |
|
| 64:26 | . B 1.2 and N A. 1.6. And so it happens that |
|
| 64:32 | you staying for these channels you will that N A. V 1.2 are |
|
| 64:40 | closer right here, closer to the in this purple region. And N |
|
| 64:48 | . D 1.6 are located in this region which is a little bit further |
|
| 64:54 | from the soma. They're both an initial segment. But maybe 1.2 is |
|
| 65:00 | to the soma and maybe 1.6 is away. Now, what we talked |
|
| 65:06 | is typically a lot of the synopsis the Selman I was telling you that |
|
| 65:12 | lot of the influence of integrative properties firing properties of the south will be |
|
| 65:18 | by neurons and connections that project right into these per somatic regions that a |
|
| 65:25 | of inhibitory cells will target these somatic and a lot of the excitation will |
|
| 65:32 | coming in the more distal excitatory That's just the way it is. |
|
| 65:37 | an ambition sometimes has a stronger control what's happening. There's remember a lesser |
|
| 65:44 | of inhibitory cells but a greater variety the subtypes of the inhibitory cells in |
|
| 65:52 | hippocampus as we discussed and also another circuits. So now if you have |
|
| 65:58 | lot of deep polarization coming in, synapse activation is not going to |
|
| 66:04 | this neuron is not going to produce action potential here and action initial |
|
| 66:09 | So you have to activate many different . Excitatory synapses. They have to |
|
| 66:16 | the inhibitory zone here where you'll have synapses and this d polarizing signal. |
|
| 66:24 | this green arrow is going to enter strong enough. The deep polarization is |
|
| 66:30 | to spatially enter into this axon initial . Then what happens next is it |
|
| 66:40 | into this N. A. 1.2 purple zone, But it doesn't |
|
| 66:46 | N 81.2 channels because these channels turn to be what we called high threshold |
|
| 66:54 | . That means they require more current more deep polarization, high levels of |
|
| 67:00 | in order to open. So this polarization is passing through here and then |
|
| 67:08 | hits this yellow area that is populated N 81.6 and we call these low |
|
| 67:16 | both educated sodium channels that means that don't require that much voltage in order |
|
| 67:22 | open up and once the voltage and polarization enters into this area it causes |
|
| 67:29 | explosion that causes the opening of the gated sodium channels positive feedback loop generates |
|
| 67:39 | action potential and number three, that a forward propagating action potential that travels |
|
| 67:46 | the axon and regenerates each note of beer until it reaches its terminal destination |
|
| 67:54 | axon terminal. So we call this forward propagating. It's propagating. It |
|
| 68:02 | being propagated and it's being regenerated and note of round dear. I guess |
|
| 68:09 | each note of run beer again is to be loaded with a lot of |
|
| 68:14 | gated sodium and potassium channels. We reproduce this action potential, regenerated and |
|
| 68:22 | its amplitude all the way to its destination. So now what happens |
|
| 68:32 | Now you have deep polarization that's coming the synapses into the soma axon initial |
|
| 68:40 | . And now you have d polarized area and maybe 1.6 even more because |
|
| 68:44 | produce an action potential here These channels now going to sense not only the |
|
| 68:52 | polarization that's coming in, but also eruption and deep polarization in the nearby |
|
| 68:59 | that happened because of the opening of M- 81.6. Now they have high |
|
| 69:05 | threshold high enough of the voltage to activated. But the current that gets |
|
| 69:12 | here goes the opposite direction. It's back propagating action potential. It cannot |
|
| 69:19 | into this direction because here you have higher default organization and this deep polarization |
|
| 69:25 | , click is not as strong to back propagating spike invades back into the |
|
| 69:33 | and back into the dendrites of the . So that's why we call this |
|
| 69:38 | propagating action potential because instead of forward and causing the release of the |
|
| 69:45 | the function of the action potential in axles is to cause the neurotransmitter release |
|
| 69:53 | the synopsis here. But the back action potential has a different function. |
|
| 70:02 | doesn't travel, it doesn't propagate, doesn't regenerate, it actually travels back |
|
| 70:12 | a smaller, much smaller currents. action potential is on the order of |
|
| 70:16 | kilovolts back propagating spike is on the of a couple of million volts that |
|
| 70:22 | back into the cell. So therefore serves a different function. The forward |
|
| 70:27 | action potential is neurotransmitter release deep realization external terminal and neurotransmitter release. The |
|
| 70:34 | propagating action potential serves a different And uh in particular I think I |
|
| 70:43 | have had this thing that I should showed you when we talked about that |
|
| 70:53 | electric behaviors of neurons. This is the following uh slides. But this |
|
| 71:01 | good to know. It's a different will express different channels and different locations |
|
| 71:07 | because of that they'll have these very patterns and dialects and in particular the |
|
| 71:14 | comes from the inhibitory cells. and then we delved into the synaptic |
|
| 71:22 | . So now you wanna know, do I care about this back propagating |
|
| 71:28 | potential and what its function if the propagating action potential serves the function of |
|
| 71:35 | your transmitter down the axon. The propagating action potential is very important. |
|
| 71:41 | plasticity. Spike timing dependent plasticity in spine, plasticity in excitatory de polarizing |
|
| 71:53 | reinforcement and summation as more deep polarization be coming in. And I have |
|
| 72:00 | challenge of the day. Do you it's preferred to property deep polarization forward |
|
| 72:04 | backward directions? But we will probably that for the second section of the |
|
| 72:10 | . So we will formally end this section of the course here today. |
|
| 72:17 | you very much. And I will everyone online on zoom on Tuesday. |
|
| 72:23 | will send you the link via email we'll post it on blackboard as |
|
| 72:28 | Good luck studying and preparing yourselves and your questions for the midterm review. |
|
| 72:34 | |
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