CELLULAR NEUROPHYSIOLOGY CONSTANCE HAMMOND Chapter 6 subtitles – postsynaptic integration INTRODUCTION (1:56) This sixth and final chapter deals with the summation of presynaptic currents. Glutamate and GABA released by axon terminals have bound to their respective receptor channels and have produced inward or outward currents. Now, what is happening in the postsynaptic neuron? What are the objectives of this chapter? At the end of the chapter, you will know how to add up EPSPS and IPSPs as they propagate along dendrites and you will understand the final effect on the sodium channels located at the axon initial segment and the final effect on the presynaptic neuron activity. Can you explain in simple terms how the various postsynaptic potentials add up to influence neuronal activity? It's a little like voting for a project or against a project. If there are more votes for it, the project goes ahead. If there are more votes against it, the project does not go ahead. The project here is making an action potential. If depolarization exceeds hyperpolarization in the postsynaptic membrane, and particularly at the postsynaptic initial segment, there will be an action potential. If, on the contrary, hyperpolarization exceeds depolarization, there will be no action potential triggered. CH. 6-1 : COMPOUND PSP, PSP PROPAGATION AND SUMMATION (11:19) What do postsynaptic potentials, EPSPsand IPSPs, become once created at postsynaptic membranes? How do they propagate along dendrites? And what is their role? Here is a neuron with two afferent glutamatergic synapses, in red, and two GABAergic synapses afferent to this neuron, in blue. If now we record the activity of this neuron in whole-cell configuration and current-clamp mode to record the changes of potential, we record this compound somatic PSP. "Somatic" because we record it from the soma and "PSP" for postsynaptic potential. So this is the depolarization. Now, if we add a blocker of GABAergic synapses, it means that the blue here are blocked and we record only the effect of the red ones, we record a bigger depolarization of higher amplitude than the green one here. It means that the glutamatergic synapses depolarize the membrane and if the GABAergic synapses are active at the same time, it reduces the effect of the depolarization. Now if we do the contrary, we apply blockers of the glutamatergic transmission, APV for NMDA channels, CNQX for AMPA and kainate channels, we record a hyperpolarization of the membrane which is not big, here, and and no depolarization at all. Now the green one is in fact the sum of these two and you can see here, at the beginning, a small hyperpolarization followed by a depolarization of smaller amplitude than the orange one. If we plot all the traces at the same scale, you see here, of course, that the depolarization without GABAergic synapses is larger and the change of potential without the glutamatergic synapses, with the GABAergic synapses only, is a hyperpolarization. You see here that synapses are all along the dendritic tree and the soma here. These synapses are called axo-spinous because they're between an axon terminal and a spine. Number 2 is axo-dendritic, because it's between an axon terminal and the shaft of a dendrite, and axo-somatic for number 3, because it's between an axon terminal and the soma. Imagine that you have a lot of these types of synapses on the dendritic branches and the soma. How do they sum along the dendritic trees and, first of all, how do they propagate? Here, we record the activity of a single synapse. The configuration is called whole- dendrite because we are attached first to a dendrite and then a small hole is performed so that we have access to the whole dendrite. This is theoretical, because it's very difficult to record the activity of a single synapse, but let's suppose we can do it. We would record this. First, in current-clamp mode, we record an excitatory postsynaptic potential. I remind you that it's called excitatory because it's depolarizing and, when it depolarizes the membrane, it has a tendency to open sodium channels at the axon initial segment. So here is an EPSP and, underlying this EPSP is a current that we can record now in voltage-clamp mode. This current is an inward current due to the entry of sodium and small exit of potassium ions. This glutamatergic current is short and inward, so there is a net entry of + charges. After this entry of + charges, how do these charges propagate so that we can record an EPSP at the soma and then at the axon initial segment? I remind you that what is very important is what happens here at the axon initial segment, because this is where the sodium channels can be activated and give rise to an action potential. How does this EPSP propagate? We now record at two sites, the dendritic site here, and a somatic site here, in whole-cell configuration and current-clamp mode to record changes of membrane potential. The synapse is on the dendrite here far from the soma. This is the origin of the EPSP. If we recorded it here, we would have an EPSP of that amplitude at time t0. Then at t1, when it's here, between the soma and the dendrite, at t2, when it's in the soma, how is the amplitude of the EPSP? Here the amplitude has decreased and here, in the soma, it has decreased a lot. You can also see that the rise time here is longer. So when the EPSP propagates, its amplitude decreases and its rise time lengthens. How does it propagate? What are the mechanisms? The + charges that entered are essentially sodium ions. These sodium ions are here in the intracellular compartment where there are a lot of other + ions, the potassium ions. They repel each other, so that that they repel, they repel, they repel each other, and they go further down like that, repelling each other. That's how they can accumulate in the soma and then in the axon initial segment. But while they repel, there are also leak potassium currents in the membrane, so that now some potassium ions exit. You remember that the electrochemical force for potassium ions is to push them out, so some of them go out. So we lose + charges, while the EPSP propagates, and at the end, of course, the amplitude is a lot decreased, because there are less + charges than at the origin of the EPSP. We say that this propagation is passive, because it's a passive repulse of + charges, and decremental, because it loses in amplitude while propagating. It should be noted that + charges propagate in two directions: they don't only propagate downstream here towards the axon initial segment, they also propagate upstream, of course, because there is no reason that there be a direction of propagation. But upstream we are not interested by it, what is interesting is what it does at the axon initial segment. We just saw how one a single EPSP propagates, now what happens when you have two here (but it could be also much more than two) glutamatergic synapses on the dendritic branches. So trace 1 alone, here, is recorded in the soma and has the amplitude here in dotted lines. Synapse 2 alone, when recorded in the soma, has this amplitude, here, in dotted lines. but when the two are active close in time, they arrive at nearly the same time in the soma. How do they sum? We see here the resultant, which is exactly the geometric sum of 1 + 2. We say that there is linear spatial summation of EPSPs. Why is this sum linear? MOOC Cellular neurophysiology– Ch.6 – 2/6 It's because the two synapses are not on the same branch, so there is no interference with each other. They sum geometrically here in the soma. This is called spatial summation because the two synapses are located in different points of the dendrites, they are not located close to each other. They are on different branches so this is called a spatial summation. There is a different form of summation, which is called temporal: it happens when the same synapse is active several times in a row. Now let's record the effect of this synapse in the soma, whole-cell recording, current-clamp mode, to record the changes of membrane potential. The synapse is active once, twice, and three times, at very close times. It means that the second time it's active, the first EPSP is not yet back to normal, to control value, to basal value, and the second one and the third one also happens when the second one is still decaying. This summation is called nonlinear: you immediately see that the third one is not equal to three times the first one. It's less than that. Why? It's a question of driving force. The current underlying the EPSP is a cation current. It equals G (the conductance of the membrane) multiplied by the driving force. The equilibrium potential for cations is around 0 mV, so when the membrane depolarizes, it goes closer and closer to the equilibrium potential. If we put the equilibrium potential here at 0, and you have here the depolarization, when you depolarize the membrane, you get closer to this equilibrium potential, and the driving force decreases, therefore the current is decreased and the amplitude of the resultant EPSP is decreased. This is the nonlinear temporal summation of EPSPs.