Neuro: 9:00 - 10:00Scribe: Molly Clark

Friday, January 22, 2010Proof: David Davis

Dr. LesterSynaptic TransmissionPage1 of 8

Introduction [S1]:
So we’re going to accept both cranial nerve 7 and 8 for #26, and we’ll try to find some extra information on identification. But because you can’t see both of them, it’s ambiguous.
And then #33, the last one, is going to be a bonus. The answer was hippocampus. Sounds like some people got it right anyway. It’s a bonus, so if you didn’t get it right, you don’t lose anything.
Ok, so hopefully a lot of what we’re going to do today will be a review. I think most people have some familiarity with synaptic transmission and receptors and transmitters and things like that. And in fact, when we go through transmitters again, you’ll see that we recap a bunch of the stuff that Dr. Banos talked about in the brainstem. We talked a little bit about it in cytology too. So, hopefully this will be pretty painless. I have to talk a little bit about ion channels again, and try to keep that to a minimum.
How is the signal transferred? [S2]
Ok, so this is by way of introduction. So when you have two neurons, and they want to talk to each other, how can they do that?
And actually, before people knew anything about synaptic transmission, it was proposed that maybe just the current crossing the membrane of the terminal would be able to induce a current in the next cell and then regenerate a voltage change and transfer the signal.
But again, remember water plumbing analogies. The resistance of the membrane is relatively high compared to the extracellular space. So any current leaving this neuron is going to be dissipated in the extracellular space just as if you were trying to get water to go from this pipe to one that was very narrow, it would just all flow outside and around the cell.
So even though people proposed that, it wasn’t very satisfactory.
How do synapses work? [S3]
So a bunch of guys, notably Katz and Eccles, who were working in Australia at the time, debated for a significant number of years about the nature of synaptic transmission. And it came down to whether one was a pharmacologist or one was a physiologist.
And the physiologist believed in electrical transmission. There were like, “why would you design a system that could propagate action potentials really quickly, and then rely on the slow release of a chemical to get the signal across the cleft?” That just seemed backwards to them. And they liked physiology and ion movement.
Learning Objective #8 [S4]
So we’re not going to talk much about electrical transmission, but you need to know that it does exist. And it exists more between glial cells, particularly astrocytes, to allow coupling. And it does exist between certain types of neurons, but we’re not going to memorize those. So those neurons can function quickly as a group. And you tend to see it more during development than at later times in life.
So, these are gap junctions. Electrical synapses are called gap junctions.
The membranes of two cells come very close, and they form these hemichannels. And then there’s a path directly between the cells.
And these channels, called connexins, are big enough that molecules like cAMP can go through. So not only do they pass ions, they can pass other things as well. So up to like a molecular weight of around 1000.
So, that’s all we’re going to talk about for electrical synapses.
Learning Objective #1 [S5]
So, again, as we talked about for how action potentials and axons and myelination come about to allow very efficient propagation, again the way that I remember the synapse and how it’s organized is that the structure directly predicts the function.
And for the neuromuscular junction, which is a prototypic synapse, we know that we want it to function in a particular way, right.
So, when we have a motor command, a voluntary motor command or whatever, we want to be able to move rapidly and contract those muscles very rapidly.
We might also want to repetitively activate those groups of muscles. So, we have to have high fidelity synaptic transmission as well.
So we might want to, if we’re playing the piano or something, we’re using the same sets of muscles over and over again, so transmission has to occur. Things have to go back to rest without inactivating or desensitizing it. And then it’s ready to go again.
So that’s how we want to think about how the synapse is designed both pre- and post-synaptically. So pre-synaptically, how do we get the transmitter out efficiently? Post-synaptically, how is the receptor for the transmitter designed to do its job very well? So we’re going to talk about those few things.
nmj structure – anatomy overview [S6]
For those of you who are not familiar, this is the neuromuscular junction. We have an axon coming down with its myelin sheath. Then it forms a number of boutons called the motor end plate on the muscle fiber itself.
If we take a cross section through one of these synaptic bouton contacts, you can see that there are active zones, and the active zone refers to where the vesicles are concentrated, directly opposed to the post-synaptic membrane where the acetylcholine receptors are present.
And you can see that the receptors are packed in very densely. If we look down on the muscle, these little things are the receptors, and you pretty much can’t get anything else in there. So we’ve optimized the number of post-synaptic receptors ready to receive transmitter.
nmj – physiology overview [S7]
So, the first thing that we know about the neuromuscular junction is (and this is going to contrast later with central synaptic transmission) that we always get a large post-synaptic potential. It’s large enough to get the muscle fiber across threshold for generating an action potential. So the muscle always contracts. If we want to walk, we don’t want to rely on the fact that sometimes our muscles will contract and sometimes they won’t. So, this is a high safety factor. We have a big post-synaptic response, and we get an action potential and contraction.
Now, you can separate the synaptic response from the action potential. Remember we talked about this on Wednesday. The two different potentials that we need to consider are synaptic potentials and action potentials. With a synaptic potential, and if we reduce the synaptic potential with curare, which comes from a plant and blocks the post-synaptic nicotinic receptors, you can get a smaller response.
And this is approximate time frame for these synaptic responses. They’re going to occur for the most part in the millisecond time range. So we’re going to get a synaptic response be relatively brief, and then if it gets to threshold we’ll get an action potential. So it has this rising phase and decaying phase.
And the reason people put curare on is that obviously they wanted to study the synaptic response independently from the action potential.
Origin of the EPP [S8]
So here’s one way we talked about briefly, it is going to contrast with action potentials right.
So, remember if this was an axon, and we generated an action potential here, it would regenerate itself the entire length of that axon. It’d be the same size.
But synaptic potentials are only generated/initiated at the site of the synapse. So, the channels open there, we get a synaptic response, it’s going to be maximum amplitude right at the synapse, and then if we measure the size of that synaptic response further and further away, it will get smaller. So it’s going to decay away.
And that’s because, as the current here is the largest, some of that current is going to leak out of the membrane and encounter internal resistance. And so there will be less current to depolarize the membrane the further we get away. Because we’re not opening new ion channels like we were for sodium channels and action potentials, we’re just relying on whatever current was generated here to carry on depolarizing the membrane. So again, this is the leaky pipe analogy right. We start the flow of water at one end at one point, and then it’s going to go away from the synapse in both directions, and some of it’s going to leak out. So there will be less and less the further away you get. So we call synaptic potentials graded potentials or passive. And they undergo passive decay.
nmj – acetylcholine receptor [S9]
So let’s go to the post-synaptic side very quickly. The acetylcholine receptor is initially purified from this guy, which is the electric ray, because it has a huge number of acetylcholine receptors in its electric organs. And so he’s great for isolating the receptor, and we use the aid of a toxin from this snake, called alpha bungarotoxin, which has a very high affinity for the receptor.
So then you can create an affinity column that has alpha bungarotoxin and get your purified, torpedo, electric organ preparation. And then you can get the alpha bungarotoxin to pull off all the receptors and purify it. And they did that.
And our current model of what a ligand gated ion channel looks like, typified by the acetylcholine receptor, is like this.
So it’s a protein that is formed by a number of subunits. In this case five.
And it has a central pore that, when an agonist like acetylcholine binds to a binding site, there’s a conformational change of the gate, and ions can move in and out.
Learning Objective #4 [S10]
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Acetylcholine receptor channel [S11]
So we need to know a little bit about which ions move in and out here. And don’t worry about all the details. Focus on this side right (left column).
So, here is the channel in the closed state, waiting to bind agonist. It binds agonist, and the channel opens. And notice that both sodium and potassium move through the same channel.
And we know from the driving force, or from the electrochemical gradients, that sodium is going to come in when we open an ion channel that will let sodium through. And potassium is going to move out.
And what this does is, if we looked for the equilibrium potential for this channel, it’s going to be intermediate between sodium and potassium. There’s going to be no net current flowing through this channel. Around 0mV ok. That’s sort of where you can imagine that sodium and potassium balance each other exactly.
Multi-ion channels [S12]
Again, if we use our little diagram showing the membrane potential, so sodium is up here and potassium is down here. It turns out that around 0 is where they’re going to reverse. Where you’re going to have no net current flow. The response will go from being in one direction to going in the opposite direction. There’s an inward movement of sodium ions close to resting potential right.
(Looking back at slide 11) So there’s very little driving force on potassium at resting potential, so sodium comes in and depolarizes a membrane so it gets towards threshold for generating the action potential.
So all excitatory, ligand-gated channels and synapses work in a similar manner. They all let mainly sodium in at the resting membrane potential because there is very little opposing driving force for potassium to leave. So, that’s the main point. At resting membrane potential, mainly sodium comes in, and little potassium goes out. If we have positive ions coming in, that’s depolarization.
Efficiency of the EPP [S13]
So, how is the receptor designed to do its job really well? And again, this is how do we optimize the signal? (Student question: I have a question about the channels. Why is it that potassium is not leaving as quickly?) Because there’s not the same amount of driving force at resting membrane potential. (looking at slide 12) So figure it like this, the difference between the membrane potential and the equilibrium potential is the driving force. So that’s about 30 or 40. If this is -60, that’ll be 38mV driving force. But you know it’s about 100mV for sodium. So that’s the potential that each ion has. It’s just the bigger battery for sodium than for potassium, so it’ll drive more current. Now, when you get towards zero, they start to have a pretty similar driving force, and they’ll completely oppose on another. And then obviously, if we have the membrane potential up here, it’ll be mainly potassium leaving and very little sodium entering.
So remember when we said we have all these receptors. A high density of receptors, so now what do we need? We need a high concentration of transmitter that’s going to activate a lot of those receptors. And the reason for that is that reaction is diffusion limited, and if we think about chemistry it’s mass action. The products of transmitter and receptor will be proportional to the concentration of those reactants. So the more receptors we have, and the more transmitter we have, the faster that reaction will proceed.
So we get a very fast reaction when we have a lot of transmitter. So, once a transmitter is in the cleft there is very rapid binding.
Then there’s very rapid opening of those channels. Once they have 2 molecules of acetylcholine they open almost instantaneously. In fact, they open with a rate that is almost beyond the limitations of our equipment to measure it. So the rate is 100,000 per second. If you flip that over and take the inverse, that means the channels are opening in less than 10 microseconds once they’re bound. So incredibly fast. That’s not going to be a limitation when we talk about action potentials in the millisecond range.
Then the channel doesn’t stay open very long, only about a millisecond or so. All it needs to do, as long as there are a lot of channels opening, is to provide enough synchronous depolarization to get to threshold.
Agonists unbinds very quickly, so it tends to just drop off. It doesn’t hang on very long.
And it’s degraded and diffuses away.
And the receptor recovers without desensitization. We know that for most receptors, if you keep an agonist around too long they’ll go in to some sort of inactive mode. And we don’t want that to happen. That might be a protective thing if we do have an excess of transmitter, but under normal conditions you don’t want to see that. So the receptor recovers without desensitizing.
So what this means is, at that synapse, once that signal gets at the end of the terminal and wants to release transmitter, it releases a lot of transmitter and we get very rapid binding, opening of channels, and a very brief response, but a lot of channels makes a big response. And then that recovers very rapidly, so it’s ready to respond again. So that’s what I mean about hi-fidelity synaptic transmission.
Learning Objective #3 [S14]
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mEPPs quantal hypothesis [S15]
Ok. On the pre-synaptic side of things. So, we’ve done the job on the post-synaptic side. We’ve made a very fast, brief response. Enough to activate sodium channels and get an action potential. What happens on the pre-synaptic side? Well we need to learn a little bit about two things.
The only things that I want you to have a good grasp of is that transmitter is packed in these things called quanta. And so we have the quantal hypothesis.
And also know that the main triggering signal for release is calcium. So calcium is important for release. And we’ll just walk through those couple of things.
So, these are what experiments were done to prove this case.
And, for those of you that know something about synaptic vesicles and transmitter release and everything, you have to bear in mind that when they did these experiments there wasn’t a good enough electron microscope to actually see the vesicles. They deduced the vesicle theory of synaptic transmission from the physiological methods.
The way they did that was, first of all they noticed when they had an evoked synaptic response or a big synaptic response, they’d occasionally see really tiny spontaneous events that were really small.
And then the second thing they did was to reduce synaptic transmission. So, they kind of knew at this point that calcium had something to do with synaptic transmission, so they reduced the transmitter release by lowering calcium, and they could get an incrementally smaller and smaller response. So remember that synaptic response is graded. Part of the reason it’s graded is the quantal theory that maybe you’re releasing one or more vesicles so you can get a graded response that gets bigger and bigger and bigger. But it does so in a stepwise manner. So you can reduce the response down to nothing, but what they saw was there were little increments in the response that seem to be statistically small all-or-none unites that they called quanta that they then said we’ll probably discrete packets of transmitter.
We stimulate here or here we get no response, and we get one vesicle. Here we may get two. Maybe this is one too. Maybe this is four. Anyway, they analyzed that and showed that that was the case.
So for efficiency at the neuromuscular junction. I keep meaning not to put this up because it confuses people.