Okay, well, I think we're now ready to move on to consider the opposite form of plasticity that we see in the cerebral cortex. Long-Term Depression. And this too was first studied in, and its mechanisms well understood within the hippocampus of the rodent model system. So, we're going to use the very same Schaffer collateral connection from CA3 to CA1 in order to demonstrate Long-Term Depression. Only now the critical difference is the frequency of stimulation that's applied in order to induce plasticity. Whereas, previously, we had a short high frequency burst of action potentials as our stimulus. Now, what we have is a more extensive period of low frequency stimulation, as our stimulus that induces Long-Term Depression. And indeed, what is seen is, when there is a period of low frequency stimulation, 1 Hz, as opposed to 100 Hz, or even 500 Hz. What we find is, a reduction by nearly 50% and the amplitude of the excitatory post synaptic potential at that CA3 to CA1 shave are collateral synapseh So this is in fact Long-Term Depression. Like Long-Term Potentiation, Long-Term Depression can be measured for very long periods of time. beyond the hour or so that is used here in this experiment. So Long-Term Depression can last for days, for weeks, for months, as best as we have observed in our animal model work. we think like Long-Term Potentiation. This is a very important means of modifying synapses across a life span. Well, let's consider the molecular mechanisms of how Long-Term Depression is induced, and how it's sustained over time. Long-Term Depression, like long-term potentiation, require the activity of both the presynaptic element and the postsynaptic process. So, there is a requirement for activity in order for the synapse to be depressed. But what's different is the nature of that activity. And I've already suggested what might be different. For Long-Term Depression there is low frequency stimulation. That stimulation is sufficient to cause the release of glutamate course. And glutamate binds to AMPA receptors and NMDA receptors. And the amount of depolarization that the low frequency stimulation provides this postsynaptic spine is sufficient to expel magnesium from the pore of the NMDA receptor channel. However, the amount of depolarization that leads to depression is not so great that we have a rapid high level increase in post synaptic calcium. Rather the amount of calcium that rises within the spine is relatively modest compared to what we saw with the stimulus that gives rise to Long-Term Potentiation. So what we have is a slower, lower level increase in postsynaptic calcium. That is critical, that lower level increase in postsynaptic calcium activates not protein kinases now but phosphatases. So these protein phosphatases then dephosphorylate target proteins. And that seems to have the opposite effect on the proteins that traffic AMPA receptors between vesicular pools in the postsynaptic spine and the postsynaptic membrane. With the activation of these protein phosphatases, what we see is the internalization of AMPA receptors. So the removal of AMPA receptors from postsynaptic membrane means that we cannot generate as much current density at that postsynaptic membrane when Glutamate is Is released, and that, in effect, is the weakening of the postsynaptic process. So remember, both long-term potentiation and Long-Term Depression require activity. They both require activation of AMPA receptors and glutamate receptors. The key difference is the level of postsynaptic calcium and the rate of rise of postsynaptic calcium. When calcium rushes in to that post-synaptic membrane, then one might anticipate the activation of kinases and the insertion of new AMPA receptors leading to the potentiation of that synapse. But if the rise of calcium is slow into a much lesser level. Then we expect the activation of protein phosphatases and the internalization of AMPA receptors. Thereby depressing the strength of that sypnatic connection. Okay we're going to shift gears now and talk about plasticity in a different part of the brain rather than the cerebral cortex. We're going to think about plasticity of the cerebellar cortex. And there the dominant form of plasticity that seems to shape the function of the circuitry of the cerebrum is Long-Term Depression. And there are some important differences in the details of Long-Term Depression in the cerebellar cortex that I want you to know. And when we come back and talk about the cerebellum, I think this consideration of plasticity in this circuit will make much more sense to you. Now, the cerebellum has some of the most beautiful architecture of any set of neural connections found anywhere in the brain. So, I won't possibly attempt to do it justice or to introduce you to all that right now, but rather so that you can understand the basic players, I'll just highlight a few of them here. The principle neuron that's found in the cortex of the cerebellum, it's not a pyramidal cell but rather its called a purkinje cell. So the Purkinje cell is really just an amazing cell, it has a huge cell body. It grows a beautiful dendritic arbor that has a very interesting geometry and we'll come back and talk about that. And the geometry of this dendrite allows it to receive perhaps as many as 100,000 or more inputs from tiny axons that run through this, this dendrite. So these axons are called parallel fibers and as they pass through the dendrites of the purkinje cells they make synaptic connections. So, the principal synapse in question when we consider Long-Term Depression it's the synaptic connection between the Parallel Fiber and the Purkinje cell. Now, there's another important connection that I need to tell you about that makes plasticity in the cerebellum possible. And it is the connection that comes from a nucleus in the brain stem that has a very nice name. It's called the inferior olivary nucleus [SOUND]. And if you've looked at the outside of the medulla, you'd see something that we call the olive. And the olive is a bulge in the ventral-lateral surface in the medulla caused by the growth and development of this nucleus. Well, this nucleus contains cells that grow out these very special axons, that enter the cerebellum cortex, and wrap around the primary dendrites of these cerebellar Purkinje cells. And because of this intimate relationship between these growing axons and Purkinje cell dendrites, these axons are called climbing fibers. Now these climbing fibers make an incredibly strong synaptic connection with the dendrites at the Purkinje neurons. Perhaps the strongest synapse that we know about in the brain. It's right here in the cerebellum between the climbing fiber and the Purkinje cell. So we've got these two synaptic inputs onto the Purkinje cell, that we're going to consider. And see how their concurrent activation leads to the depression of the connection between the parallel fiber in the Purkinje cell. So again we're thinking about the cerebellum. Just want to emphasize that point, not the cerebral cortex. So let's see what the phenomenon actually is. So, the pairing of activity between the climbing fiber and the parallel fiber is what causes the depression in the strength that the connection between the parallel fiber in the Purkinje cell. So what we're going to measure here is the amplitude of the excitatory postsynaptic potential that is induced by stimulation of the parallel fiber. Now the test phase is the pairing of the activation of the climbing fiber and the parallel fiber. And what we see is that after that test phase there is a reduction in the amplitude of the excitatory postsynaptic potential. So that this is now depressed [SOUND] by about 50%, following this pairing of climbing fiber and parallel fiber activation. So, this is Long-Term Depression in the cerebellar cortex and it's mediated via the internalization of AMPA receptors. So the synapse between parallel fiber in the Purkinje cell is a glutamatergic synapse. Gluatamate's released, it can interact with receptors for glutamate. Only here, notice that we do not have NMDA receptors, or at least NMDA receptors are not contributing in an important way to this form of cerebellar plasticity. Rather, what we have are metabotropic receptors for glutamate. So, these metabotropic receptors, as you may recall, are receptors that activate G proteins that are associated with their cytoplasmic face and those G proteins then can then go on and activate other effector systems within the postsynaptic process. In this case those effector systems involve Phospholipase C and the IP2 pathways that leads to the generation of IP3 or Inositol trisphosphate and DAC glycerol. These second messenger systems can cause the release of calcium from richer cellular stores. Cell calcium is flooding into this postsynaptic spine with the activation of this metabotropic glutamate receptor. Meanwhile, calcium is flooding into this post-synaptic spine because the climbing fiber has been concurrently activated. And as I mentioned, the climbing fiber gives rise to one of the most powerful synaptic connections in all of the brain. This synaptic connection induces significant depolarization into this, dendritic tree. That depolirization opens up voltage aided calcium channels allowing calcium to flood into the cell. So calcium coming from outside, as well as calcium coming from the intracellular stores, results in a significant and a rapid rise in intracellular calcium in the post-synaptic spine of the Purkinje cell. This sudden and high-level rise in intracellular calcium interacts with calcium-sensitive mechanisms that internalize AMPA receptors. So, in this model system, protein kinase C phosphorylates target proteins that leads to the internalization of AMPA receptors. So AMPA receptors are being drawn from the post-synaptic membrane of these spines into the visicular compartments within the post-synaptic process. Removal of AMPA receptors is depressing this synaptic connection. Now, let's be clear: In the cerebral cortex, high levels of calcium lead to potentiation, but in the cerebellar cortex, high levels of calcium leads to depression. And the difference has to do with the second messenger cascades, and the consequence of activating those cascades. In the cerebral cortex, those cascades end up inserting AMPA receptors into the post-synaptic membrane. But here in the cerebellum a different cascade leads to the internalization of AMPA receptors and the, the depression of that synaptic connection. Now we've come to the conclusion of this tutorial and we're ready to begin to apply these concepts and think about how synaptic plasticity might play out. In the context of ongoing patterns of activity in the brain rather than those induced by stimulating with microelectrodes in experimental situations. And this brings us to a fairly new discovery in our growing understanding of the mechanisms of synaptic plasticity. So our next tutorial will be about spike timing dependent plasticity. I'll see you then.
