So what we've been talking about so far are the molecular structures that constitute ion channels and this is a way of discharging those gradients that are essential for generating electrical signals. Now, let's turn to the molecular mechanisms that are responsible for establishing those gradients in the first place. I mentioned when we first introduced this topic that those gradients are established by ion pumps and ion transporters. And here we see a variety of pumps and transporters. Probably the most important that I should emphasize is right over here on the left hand side of figure 4.9. It's the Sodium-Potassium ATPase. This is, arguably the most important molecule in the nervous system. It seems a little ridiculous to even make such a claim, but if it were possible to identify one molecule of supreme importance this would be it Sodium-Potassium ATPase. It consumes a significant portion of all of the energy that is burnt up in the function of a nervous system. And it's one of the principal reason why the brain requires such a constant and highly regulated flow of blood to deliver oxygen and glucose the principal substrates for oxidative metabolism in brain tissue. There are other pumps besides the Sodium-Potassium ATPase. As we see here there are pumps for calcium. There ion exchangers that move sodium and calcium and protons. There are co-transporters that can take advantage of a concentration gradient for one ion to transport another and we don't need to get into all of this complexity at the present time. Rather, what I want to do is consider in a little bit more detail, the function of. Our principal pump, the Sodium-Potassium ATPase. Now, the story of how this function works is the result of a lot of empirical research that had been done in various ways one can imagine based on our prior consideration of the action potential that one can study this pump by removing either sodium or potassium from the solution and asking how these manipulations might affect pump activity. One might also gain some insight by depriving that pump of its supply of energy to find out what's happening. So these kinds of experiments, were done. and what's shown here in figure 4.10 is an experiment where a, tagged sodium molecule was allowed to flux across the plasma membrane, due to the activity of this pump. So sodium was loaded up within a cell and the efflux of sodium which represents the activity of the pump is measured over time. So as this experiment unfolds there is some discharge of sodium ions that can be measured experimentally. And if one removes potassium from the medium outside of the cell, now there's a sudden drop in the activity that's being indicated by the efflux of sodium. So this suggests that there must be some kind of codependency between the Influx of potassium ions into the cell, and the efflux of sodium. Well, when potassium is restored, and the fluid's around the cell. Now, the rate of discharge of sodium is restored. But now, a key experiment is imposed upon this model system. the energy that's required to drive this activity is depleted by adding a compound which blocks the synthesis of adenosine triphospate which is the main energy currency for active cells. And, as this metabolic, poison, if you will, is introduced, there is a precipitous decline in the efflux of sodium ions providing strong evidence for the energy requirement of this transport. When ATP is restored, then there is a somewhat rapid rise again of efflux of sodium, back towards the rate at which this experiment was run. So taking these two phases of the experiment together, we know that there's some kind of co-dependency between the influx of potassium and the efflux of sodium, and we also know that this process is energy dependent. So these kinds of experiments have led to the following model. There is some kind of integral membrane protein, that constitutes a pump for sodium and potassium. And this integral membrane protein has enzymatic activity that can cleave ATP and consume energy in the form of breaking that high-energy phosphate bond. So, we have good reason to believe that there is a configuration of this protein that allows for the binding of sodium ions to the exposed cytoplasmic domain within the central region of this integral membrane protein. There seems to be a particular stoichiometry to the transport of sodium and potassium. We think there're likely three binding sites for sodium ions that are available in this configuration. And then with the consumption of energy, in the form of cleaving this high-energy phosphate bond from ATP, there is a conformational change, such that this protein appears to flip within the membrane. We don't know exactly how this happens. But it seems to change conformation allowing these three sodium ions now to be discharged. On the extracellular face of the membrane, and at the same time, this conformational change has exposed now binding sites for potassium ions. Now, with the binding of these potassium ions, the inorganic phosphate is lost, and the conformational change occurs. And we are restored to the starting configuration of this protein. And this allows for these potassium ions now to be discharged and enter the intra-cellular space. So now this protein is ready to return to its starting position. In this schema, where sodium ions can aggregate. So with the consumption of a high-energy phosphate bond, there is a cycle of this pump that transports 3 sodium ions from inside the cell to outside the cell. While at the same time allowing for 2 potassium ions to travel from outside of the cell to the interior. So as this pump churns and burns energy we establish a concentration gradient with higher sodium ions outside of the cell compared to inside, and higher potassium concentrations inside the cell compared to outside. All of this requires energy. Now, notice the stoichiometry. There's more positive charge leaving the cell than entering the cell with each turn of a cycle. From this point of view, we say that this pump activity is electrogenic, that is, it has the potential to generate a small current. That current typically is negligible unless we're talking about a very small compartment where the concentrations of ions might actually change during the course of a high frequency burst of action potentials. Such compartments exist in the smallest of our axons we find in peripheral nerves. So, in such situations as ionic concentrations might be altered by a burst of action potential activity, pump activity will increase, more positive charge will leave the cell as a result of the pump, and the inside of the cell will hyperpolarize. So that would be the impact of this electrogenic nature of pump activity. Again, it's typically negligible, except for the very smallest of axons, or the very tightest of compartments where pump activity can have an impact on the resting memory potential. Well we can summarize and have an opportunity to review these mechanisms of pump activity by viewing a animation from chapter four you can follow this animation. by navigating to the website that supports our textbook or by clicking on the hyperlink that you have available to you at the bottom of your tutorial notes. Well, I hope this has been helpful. Our next tutorial will focus on the means by which electrical signals can propagate along an axon. And we will return to some of these concepts related to the structure of ion channels and pumps when we consider synaptic neurotransmission in forthcoming tutorials.