Fat

Hello and welcome back to our online sport nutrition lecture series. This week’s topic is going to be fat, and there’s going to be a couple of different portions of this online lecture. The first of which will be setting up the metabolic pathways by which fat is broken down and used as a fuel source during exercise and its contribution as a fuel source during exercise.

Then, once we’ve set that up, what I’d like to do—because it’s a major topic of interest in research and even from reading your discussion forums up to this point, it’s a major topic of interest amongst people in the class—is this idea of low-carbohydrate, high-fat diets, as we typically see them used as a weight-loss diet. We’ll talk some about that later when we talk about weight management, but there are also individuals who adopt low-carbohydrate, high-fat diets because of purported performance benefits. I want to investigate whether or not there are any performance benefits or even detriments from adapting to a low-carbohydrate, high-fat diet based on the scientific evidence that we have in athletes currently.

So first, let’s talk about fat as a fuel source in terms of its availability and the sources of stored fat that we have available to use as a fuel source. This is going to be a review, but I think this is an important place to start. We talked about this in our earlier lecture of fuel sources for exercise metabolism and, in comparison to our storage of carbohydrates—which we talked about our different forms of stored carbohydrates last week—how abundant are stored fats? Hopefully, you will remember that if we think about how much fuel this is in terms of availability for exercise, we have practically unlimited fat stores to work from. This is even true again in individuals who would be considered lean—the athletes, the active individuals that we’re referring to mainly when we’re talking about sport nutrition.

Let’s take an example of a 70-kilogram individual who is pretty lean at around 10% body fat. Well, that means that that individual has 7,000 grams of stored fat available for energy. And if we break that down into calories—because every gram of fat represents about 9 calories worth of energy—that means that a 70-kilogram, 10% body fat individual has about 63,000 calories worth of energy in the form of stored fat. So to break that down in exercise terms in a few different ways, that’s enough to fuel about 207 hours of running six-minute miles—so pretty, pretty fast pace. It’s also enough to fuel about 240 hours of vigorous-intensity cross-country skiing. And if you want to make it a little bit more fun, about 385 hours of dancing—maybe an activity that we don’t always think about as exercise, but certainly one that costs energy and that is fun to do. So the point here is that we have more than enough fat to fuel most of the different types of activity and exercise that we go about in our bit, in our daily lives.

And so where do we get that fat from? Well, we get it from a couple different places, but the most prevalent place that we have it stored is in our adipose tissue. And that adipose tissue can take the form both subcutaneous—so that is directly beneath the skin. This will be familiar to you if you’ve ever done a skin-fold test where you’re taking the amount of subcutaneous fat that somebody has in different areas and using it to predict how much body fat they have overall. But of course, we also know that some store, some fat is stored viscerally. This is one of the reasons that we look at abdominal fat and its relationship to disease because visceral fat is that which is found down deeper, primarily in the abdominal cavity, found well, well a couple levels below the skin and surrounds a lot of vital organs and can be important for cushioning and mechanical protection, but also can be metabolically dangerous and increase your risk for cardiovascular and metabolic disease. And then, of course, we also store some triglycerides, or some fat, in our muscle cells themselves. These are conveniently located as sources of stored fat that can be easily broken down and used to fuel muscle contractions. And those will come back when we start looking at how IMTGs, or intramuscular triglycerides, can change in response to exercise training and can be used as a fuel source during exercise.

So if we have 63,000 calories of fat available to us as energy, why might it be difficult for us to use that fat for energy or rely exclusively upon it in many exercise situations? And the answer to that is that there are several key steps that limit fat oxidation, especially when we compare it to the number of steps needed to oxidize carbohydrate. And so we’re gonna break this figure down step by step and look at all the individual elements that make up all of these steps in fat oxidation. But to kind of review what we will go into as we dive further into this process, we can outline what some of these key steps are. And the first is the step of lipolysis. So again, we have fatty acids that are stored in adipose tissue, as you can see here. Fatty acids, as we’ve discussed previously, are primarily stored in triglyceride form, so with 3 typically long-chain fatty acids bound to a glycerol backbone. And then, of course, we also have our other stored form here, our primary stored form located within the muscle, also will also intrigue lesser ID form, and also gonna need to undergo to the process of why pollicis to get the fatty acids from their triglyceride form, whether we’re talking about adipose tissue or intramuscular triglycerides. And then, of course, there’s a very small amount that we store in our circulating lipoproteins, such as VLDLs, that don’t contribute very much as a fuel source to exercise. Primarily going to be our triglycerides coming from adipose tissue as well as intramuscular triglycerides. So the whole process with lipolysis is that we need to break those triglycerides down into fatty acids and glycerol so that those fatty acids can be removed to the fat cells and then can make their way into the bloodstream. So that’s where we are now, where they can then be transported to the actual muscle cells where we’re going to use them to fuel muscle contractions. So we have to get them from their triglyceride form, free of the fatty acids from those triglycerides, transport those fatty acids via the bloodstream to the muscle cells themselves. And then, finally, in order for those fatty acids to ultimately be used to resynthesize ATP, they’re going to need to make their way into the mitochondria of those muscle cells in order to undergo beta oxidation and full oxidative phosphorylation so that they can provide large yields of ATP. So we’re gonna go step by step in detail into each one of these and talk about some of the physiology that is implicated and allowing all of these steps to happen.

So let’s dive right in and start with the first step here, which is lipolysis. And the primary enzyme that regulates the rate of web pollicis & adipocytes is going to be known as hormone-sensitive lipase. And this is the enzyme that is again primarily responsible for splitting that triglycerides into fatty acids and glycerol so that those fatty acids are now available to be transported in the bloodstream to muscle cells. A similar isoform of this enzyme exists in vascular walls, and that’s primarily responsible for the small amount of fatty acids that are broken down off of our VLDLs and circulating kyra micron. Again, those contribute very little to energy expenditure during exercise, but the few fatty acids that are cleaved off of these forms of fat are also achieved via a similar enzymatic pathway. And there’s a specific type called lipoprotein lipase that accomplishes that job. But hormone-sensitive lipase exists in both active and inactive forms, and as the name suggests, its activity or inactivity is tightly regulated to circulating levels of hormones, specifically epinephrine and insulin. And we can see that relationship depicted here graphically. And what we can see here is that epinephrine—as we know, epinephrine levels and circulating epinephrine levels are tightly related to sympathetic nervous system activity—and what happens when there is more epinephrine circulating is that more epinephrine will bind to these receptors on the outside of adipose cells. And when that happens, it facilitates the conversion of more hormone-sensitive lipase into an active form versus the inactive form, which ultimately facilitates further breakdown of fatty acids and release of fatty acids into the bloodstream so that they can be available for energy production.

Now, to counter that, while epinephrine levels will stimulate hormone-sensitive lipase to express more of an active form, insulin is actually counter-regulatory. So when, when epinephrine levels are high, usually insulin levels are suppressed, so we have less insulin binding to fat cells, and that also facilitates the effects of epinephrine on hormone-sensitive lipase and increasing hormone-sensitive lipase activity and fatty acid breakdown. But when more info is circulating, what’s gonna happen is less of this hormone-sensitive lipase is going to be converted into the active form, and fewer fatty acids are going to be cleaved off their triglyceride form and available for energy production. So if we think about this for a moment, you should go ahead and pause the video, and I want you to take a minute, and I want you to think about why this relationship physiologically between hormone-sensitive lipase and these two hormones, insulin and epinephrine, is actually an intuitive design. Now, hopefully, after you have had a chance to restart the video and pause and think about this for a minute, you’ll arrive at the conclusion that if we think about what happens when lots of insulin is around, it’s usually indicative of the fact that we have plenty of carbohydrate available to, to rely upon. So when we want to increase fat breakdown, we want to increase the hormones that are going to stimulate hormone-sensitive lipase in a fashion that’s going to facilitate further lipolysis and subsequently further fatty acid oxidation. But when we have a lot of insulin circulating, it also usually means that we have carbohydrate available to us as well. And so when insulin is around in higher proportions because we are shifting our reliance more towards, towards carbohydrates, this means that we have a negative feedback loop that occurs with these two hormones that allows our lipase activity to increase and then able to enable us to rely more on fat and rely less on carbohydrate when we’re able to do so, and vice versa. So this hormonal triggers really help to gauge and regulate our enzyme activity, which allows us to incur some of the shifts and fuel selection that we see happening when shift intensities of exercise and subsequently shift our reliance on different fuel sources.

Okay, so now we have our fatty acids removed from the adipocyte itself, broken down from its triglyceride form, and now we have fatty acids that are available to move into the bloodstream. And because this is a critical step, blood flow to adipose tissue is going to be a critical limiting factor in getting those fatty acids that have been released for their triglyceride form into the bloodstream so that they can be transported. And the question then becomes, once those fatty acids have been released from their triglyceride form and once they then are into the bloodstream, are they freely transported into the bloodstream, or do they need some help? And the answer to that is that they’re not freely transported. The transport of fatty acids in the bloodstream is dependent upon a carrier protein known as albumin, which is what is depicted in the figure over to your right. So we’re kind of adding on to this. We started in the adipose cell itself. We now have undergone lipolysis via hormone-sensitive lipase, and now we have fatty acids that can move from the adipocyte into the bloodstream. But once they’re there, in order to be transported, they rely on this protein known as albumin. And albumin is actually the most abundant protein found in blood. As we can imagine, fatty acids being one of our major two fuel sources for most things that we do, we need a decent amount of this carrier protein in the blood in order to transport those fatty acids. And so one of the things that limits this step in terms of the removal of fatty acids being limited by the amount of blood flow that is being presented to the adipose tissue, we’re also going to be limited by the amount of human, or the concentration of circulating albumin in the blood, and the number of sites on each albumin molecule that are open and are able to accept a fatty acid for transport. So if we look at how much albumin we have around, interestingly, we’ll see that a normal plasma albumin concentration for a human being is about 0.7 millimoles per liter. And that number becomes useful when we start to think about what normal circulating plasma fatty acid concentrations are like at rest but also during exercise. And so if we look first at rest, we can see that in comparison to how much albumin we have around at rest, we have about 0.2 to 0.4 millimoles per liter of circulating fatty acids in the plasma. So thinking about the fact that this albumin is there to carry these fatty acids, we have more than enough albumin at rest to meet the demand of resting plasma fatty acid concentration. So at rest, we have no trouble transporting all of the fatty acids that are in blood to their target tissues. But when we start to look at what happens during exercise and we look at peak rates of circulating fatty acid or circulating plasma fatty acid concentrations, we can see that number rise up to 2.0 millimoles per liter—so more than twice the concentration of circulating albumin. So what this tells us is that during exercise that allows for peak fatty acid circulation, we pretty, we more than double the capacity of albumin to bind these fatty acids. So during exercise, while we are freeing up a lot more fatty acids, we don’t necessarily have enough albumin to transport all of the fatty acids that are being released to where they need to go. So we more than exceed our capacity to do so. So therefore, not all of those circulating fatty acids are going to be able to reach their target tissues. So this is definitely one of the limits of fatty acid metabolism is our limited ability to transport the fatty acids that are being freed to their targets due to the limited capacity of albumin.

Okay, so now let’s say we have a fatty acid; it’s bound to albumin; it’s transported via the bloodstream to the muscle now; and now what we need to do in order for it to be fully oxidized and used as fuel, the first thing, or the next thing that needs to happen, is it needs to get across the sarcolemma, or the membrane, the cell membrane of the muscle cell. And so we will now start to think about what might be necessary for this transport to occur. And if we take a look at what’s going on here, let’s take our fatty acid; it now is moving out of the vasculature, and it needs to get across this semi-permeable muscle cell membrane or sarcolemma. And we have a few different proteins that help facilitate the translocation of these fatty acids across this membrane or the diffusion of these fatty acids across this membrane. So this is going to be facilitated by these different proteins, and we have them located in a couple of different places. And the first type of protein that we have is this FABPPM, and what that stands for is fatty acid binding protein that is bound to the plasma membrane. So this set of proteins, these FABPPM proteins, are located actually embedded within the sarcolemma itself. So via this protein, some of this membrane permeability will be altered in a fashion that will allow some of our fatty acids to then diffuse into the sarcoplasm or cytoplasm of the muscle cell itself. We also have a protein known as FAT/CD36 that is found actually in the sarcoplasm itself, and these function in a very similar fashion to how GLUT4 proteins, which are also found in the sarcoplasm, translocate to the membrane. And once they get to the membrane, in a similar fashion to the proteins that are bound to the membrane itself, it also further facilitates the transfer of these fatty acids across the sarcolemma by altering that membrane permeability. And in a fashion similar to GLUT4, what’s interesting is just how GLUT4 translocation is facilitated via muscle contraction independent of the activity of hormones like insulin, FAT/CD36 translocation is also stimulated by muscle contraction. So during exercise, more of these proteins translocate from the sarcoplasm to the sarcolemma and facilitate further uptake of fatty acids out of the vasculature and interstitial space and into the, the muscle cell across the sarcolemma. So both of these proteins ultimately serve to facilitate the diffusion of these fatty acids across the membrane, across the sarcolemma; they just do it in slightly different fashions—one is bound to the membrane, and one is found in the sarcoplasm and needs to translocate to the membrane before it can ultimately alter membrane permeability. So these are going to be our primary two proteins that are going to get these fatty acids from the vasculature into the muscle cell themselves across the sarcolemma.

Now, finally, what we also have is our third class of fatty acid binding proteins, which are also found in the cytoplasm or sarcoplasm itself. These are FAPC, or cytoplasmic fatty acid binding proteins, and what these do is once these fatty acids have been facilitated across the sarcolemma and are now in the sarcoplasm, well, they now need to be carried to the mitochondria where they can undergo further breakdown and ultimately undergo oxidation in order to produce energy. So these fatty, these fatty acid binding proteins that are found in the cytosol or sarcoplasm are the proteins that, once the fatty acids are actually inside the muscle cell itself, that’s what’s going to pick them up and carry them to the mitochondria where they can undergo the rest of the metabolic processes needed. Now, obviously, that process of getting fatty acids from the bloodstream across the sarcolemma into the muscle cell is really specific to fatty acids that are coming from things like adipose tissue or sources of fat that are outside the muscle cell itself. But we know that we store some fatty acids within, or some triglycerides within, the muscle itself, or intramuscular triglycerides. And if we think about our different types of muscle cells, our different fiber types, we may begin to ask ourselves the question, are there differences in intramuscular triglyceride content across fiber types? And the answer, not surprisingly, is that our type 1, or are highly oxidative fibers that have the metabolic hardware and circulatory hardware to both deliver and metabolize large amounts of fatty acids and prefer to work that way energetically, have the highest storage of intramuscular triglycerides. And if we take a look at a picture from an electromyograph of a skeletal muscle cell, this is actually the inside of the muscle cell itself, and what we can see here, this dark circle, this mi, this is the actual mitochondria itself. So outside of the mitochondria, this is all going to be out in the sarcoplasm of the muscle cell; we see these lipid drops. So we can kind of see this one depicted here, which is a pretty large lipid drop. And what we might notice is, first of all, that triglyceride is inside the muscle cell itself, so we’re not going to need to worry about transporting those fatty acids across the sarcolemma after they’re broken down from intramuscular triglycerides. But we also noticed that it’s very conveniently located directly adjacent to the mitochondria, and that makes a lot of sense to reduce the distance that those fatty acids need to travel before they ultimately go into the mitochondria for oxidation. And if we look at intramuscular triglycerides in the muscle cells of trained versus untrained individuals, while almost in all individuals IMTGs are, as I just said, found adjacent to the mitochondria in these droplets, one of the things that happens as in response to training is that a greater proportion of these IMTGs are found adjacent to the mitochondria compared to untrained muscle. So if we were to take a look at this electromyograph before a period of training, so in an untrained individual, this lipid droplet might be a little bit farther from the mitochondria itself. But once we trained, what we would see is that these triglycerides actually start to relocate and move even closer to the mitochondria themselves. And this is one of the reasons why endurance athletes are better able to oxidize IMTGs as a fuel source during exercise. We also know that these lipid droplets, these IMTGs, contribute substantially to exercise because something else we could do is look at the size of these lipid droplets before and after about of the exercise, and we would undoubtedly see that following a bout of endurance exercise, the size of these lipid droplets would actually decrease, indicating that they are indeed an important fuel source during exercise. So training allows us to use IMTGs better because it facilitates the location of those lipid droplets to the actual mitochondria itself; they get closer to the mitochondria. In addition, we’ll also see that endurance athletes typically have overall higher absolute storage of IMTGs in type 1 fibers specifically compared to untrained individuals. So you have more fatty acids conveniently located closer.

To the mitochondria where they can be used for fuel. And the way that these are broken down, as I've alluded to previously, is again through hormone-sensitive lipase. It functions in the same way as it does in adipocytes inside of skeletal muscle cells, ultimately to release those fatty acids from their IMTG's.

In terms of the carrier proteins, now that we're thinking about inside the muscle cell, because the process of lipolysis is occurring inside the muscle cell itself, we don't need to rely on fatty acid binding protein that is in the plasma membrane—so FABP p.m. or FABPcd36—to help get those fatty acids across the sarcolemma. They're already there. But what we do still need to happen is those fatty acids still need to be carried by cytoplasmic fatty acid binding proteins, or our FABPCs. Because, if we'll remember, those proteins are necessary to carry the fatty acids that are going to be released from these IMTG's into the mitochondria—albeit maybe a short distance to travel—but once they're released, they are indeed in the cytoplasm and are going to need some help getting to the mitochondria itself.

So now we have our fatty acid at the mitochondria, and we need to transport the fatty acid into the mitochondria itself. And in order for that to happen, and these fatty acids are ultimately going to have to cross both the outer mitochondrial membrane into the inner membrane space and then cross the inner mitochondrial membrane so that they can enter the mitochondrial matrix where beta-oxidation and oxidative phosphorylation are going to occur. There are a few different enzymes and steps involved here. And the first thing that needs to happen is once these fatty acids are in the cytoplasm, they have to be activated to form this fatty acyl CoA complex. And they do that via the activity of this enzyme that is bound to the outer mitochondrial membrane, known as CoA synthetase. Basically, what this does is it combines the fatty acid with some free coenzyme A to form this fatty acyl CoA complex. And that fatty acyl CoA complex is then able to move across the outer mitochondrial membrane and into the inner membrane space.

In this space, what's going to happen is the fatty acid is now going to bind to a compound known as carnitine. And what's going to happen when this fatty acid binds to carnitine is it's going to form this acylcarnitine complex. And the coenzyme A—that CoA portion of that molecule—is now going to be released and it's going to be pumped back outside the outer mitochondrial membrane so that that coenzyme is now free to form more fatty acyl CoA complexes. This conversion of fatty acyl carnitine is the first step; the binding to carnitine is the first step in the process of transporting an activated fatty acid into the mitochondria itself. So this step is limited by the activity and concentration of carnitine in the inner membrane space. This process is also catalyzed by an enzyme. The binding of fatty acids to carnitine and creation of this fatty acyl carnitine complex—free coenzyme A is again released—and this creation of acylcarnitine and release of free coenzyme A is facilitated by another membrane-bound enzyme known as carnitine palmitoyltransferase 1. And so the name of that enzyme tells you a lot of what it does: it's able to transfer its fatty acids from the fatty acyl CoA complex to make a fatty acyl carnitine complex and ultimately move that remaining free coenzyme A back out into the outer mitochondrial—outer outer mitochondrial membrane—so that it can pick up more fatty acids and continue to facilitate the process of getting fatty acids across that first membrane and into the inner membrane space.

So now we're in the inner membrane space, and what we ultimately need to do is get across this second membrane, the inner mitochondrial membrane, into the mitochondrial matrix so more of the metabolic magic, so to speak, can happen. And what allows this to occur is this fatty acyl carnitine complex. So again, these are the fatty acids combined with these carnitine compounds; it now needs to be brought across this membrane, and it's brought across that membrane by an enzyme bound to the inner mitochondrial membrane known as translocase. Translocase allows for this acylcarnitine complex to move across the inner mitochondrial membrane and into the mitochondrial matrix where ultimately now it can pick up some more free coenzyme A and can be converted back into a fatty acyl CoA complex, which is going to be necessary to undergo the process of beta-oxidation.

So that's not the entire story here. We can see now, okay, we have now recreated a fatty acyl CoA complex which can then move forward for beta-oxidation. But remember, this was a fatty acyl carnitine complex when it moved across the membrane via the activity of the enzyme translocase. So if we can think about what now needs to happen, is we also now need to get this carnitine back into the inner membrane space because remember the presence of carnitine in the inner membrane space is critical for getting fatty acyl CoA complexes across the outer mitochondrial membrane. And so in order for that to happen, any carnitine that gets pumped across the inner mitochondrial membrane ultimately needs to get back into the inner membrane space so it can pick up more fatty acids and continue to further undergo this process. And so this carnitine is actually able to diffuse back across the inner mitochondrial membrane, out of the mitochondrial matrix and back into the inner membrane space via the activity of another enzyme known as carnitine palmitoyltransferase 2. So call it carnitine palmitoyltransferase 2, CPT-2, is the enzyme that's going to allow for this carnitine, once it has been removed and this fatty acyl CoA complex is once again formed via the combination with some free coenzyme A, we then can now take that free carnitine and bring it back across into the inner membrane space so we can pick up more fatty acids.

Now, once inside the mitochondrial matrix, we can now undergo the process of beta-oxidation. And to sort of review what's going on here, remember now we have a fatty acyl CoA complex, and what we need in order to run each turn of the Krebs cycle is we need these two-carbon acetyl CoA molecules. And so what ultimately happens to this fatty acid is that fatty acyl CoA is split into these two-carbon acetyl CoA molecules, and each of these two-carbon acetyl CoA molecules can then be oxidized in the citric acid or Krebs cycle.

And so let's review what the primary purpose of oxidizing these two-carbon acetyl CoA molecules is in the Krebs cycle. And we can remember that the primary purpose of this is to: one, generate a very small amount of ATP on its own; but primarily what it does is it generates carrier molecules of NADH and FADH2, ultimately going to carry electrons to the electron transport chain and allow for the large amounts of ATP resynthesis that we can see when breaking down long-chain fatty acids, and specifically when there is enough oxygen available to pick up those final electrons at the end of the electron transport chain. For every NADH molecule that we create, we will produce 3 ATP, while for every FADH molecule we create, we will produce 2 ATP. So this is physiologically, again, one of the main reasons why we're able to get large amounts of ATP from a fatty acid. Again, like a typical long-chain fatty acid that's found in the human diet or an adipose tissue would be about 16 carbons long, so we get a substantial amount of ATP from all the acetyl CoAs that we can form from breaking down—at beta-oxidizing—one of those long-chain molecules.

So now let's take a look at what happens to fat oxidation during exercise. And if we ask ourselves the question of what happens to the rate of fat oxidation as someone exercises longer at any given intensity of exercise, well, we'll generally see that at a given intensity of exercise, as duration increases, fat oxidation also increases. Now this is both a function of—as duration gets longer, we have more time for all of these steps involved in fatty acid breakdown and metabolism to occur—so we're able to ramp those metabolic pathways up at a point at which they can make a significant contribution to overall energy demand. But we also know that this increase in fat oxidation is going to occur as our carbohydrate stores become potentially more limited.

So if we take a look at fat oxidation then as a function of exercise intensity, we can see on the figure here we're looking at exercise intensity on the x-axis as a function of percent of VO2 max, and then on the y-axis we're looking at the overall rate of fatty acid oxidation in grams per minute—so how much fat is being oxidized and used to supply the energy needed to perform at each one of these given exercise intensities. And what we can notice when we take a look at this figure is that when we go from light to moderate intensity exercise, as these metabolic pathways have time to ramp up and make significant contributions and reach peak levels of oxidation in their contribution to energy expenditure, we see that generally fatty acid oxidation increases as intensity goes from light to moderate. But once we shift from moderate to more vigorous intensity exercise, we see that as the rate of ATP demand increases, well, we're not able to wait on those fat pathways for quite as long in order to adequately meet that rate of demand. So once we start to get up in trained individuals, we see this rate of fatty acid oxidation start to peak somewhere around 62 or 63 percent of their VO2 max. This would also be somewhat close to their ventilatory and lactate thresholds; there might be a little deviation there, but again, if we think about what's going on, once we push past that point, we start to increase the rate of energy demand to a level where we need to recruit other energy sources—primarily carbohydrate—in order to make up the difference, which is why we then start to see the peak rate of fat oxidation decrease as we go above that level. And then when we get into much higher intensities, somewhere around about 75 percent of VO2 max, we see a really sharp decline in the rate of fatty acid oxidation. And again, this is because we are having to rely more on forms of energy like carbohydrate that can be broken down at a fast enough rate to make up that energy deficit and continue to supply ATP at a rate that is commensurate with the demands of the exercise session itself.

Now, a couple of weeks ago, we read an article suggesting that when an individual undergoes a period of endurance training, they ultimately will be able to oxidize fat at higher levels at any given intensity of exercise compared to pre-training. So they might be able to have higher peak rates of fat oxidation following training and may be able to shift some of that peak compared to where it was pre-training. And so this implies that there are metabolic adaptations that occur in response to training that allow more fat to be exercised or oxidized during exercise itself. And so if we take a look at what these are, there is evidence to suggest—I'm going to review an overview of some of the findings from years and years of research on what allows this to occur—but some of the metabolic adaptations that occur that allow for higher rates of fat oxidation in endurance athletes are mechanisms such as increased mitochondrial density and oxidative enzyme activity. So by having more mitochondria per unit of muscle tissue—this is primarily occurring a lot in your more oxidative fibers—and by increasing the activity of the enzymes that are involved in the mitochondrial breakdown of fat to resynthesize ATP, endurance athletes have more dense mitochondrial beds and higher activity of enzymes that are able to break down those fatty acids in the mitochondria. So this improves their capacity to oxidize the fat that actually makes it into the mitochondria itself.

What also happens is those muscle fibers will have an increased capillary density. So if we think about capillary density as being critical for delivering the fatty acids that are broken down from adipose tissue to the muscle cells, higher capillary density will mean there is more surface area for the exchange of those fatty acids between the vasculature and the sarcolemma to occur and ultimately improve the delivery of fatty acids to the muscle cell itself. Once those fatty acids get to the muscle cell itself, endurance-trained individuals also have an improved ability to transport those fatty acids across the sarcolemma. The research has shown that this primarily occurs as a function of increasing the amount of fatty acid binding proteins that are found at the plasma membrane—are bound into the membrane itself. So following a period of training, an individual may have an increase in their FABP p.m. concentrations—more of that protein bound to the membrane—and therefore more facilitation of the take-up of fatty acids across the sarcolemma into the muscle cell itself. Additionally, we will see that if we look at what's going on in the mitochondrial matrix, we'll see that endurance training will increase carnitine palmitoyltransferase 1 and 2 concentrations, which ultimately will facilitate the transport of the fatty acids that are in the sarcolemma across the inner and outer mitochondrial membranes so that they can ultimately make it into the mitochondrial matrix for subsequent beta-oxidation and oxidative phosphorylation.

And then lastly, I previewed and referenced before, is that endurance training increases the overall concentration of intramuscular triglycerides. So there will be more of these triglyceride lipid droplets found within the muscle cell itself but are available to fuel contractions, and those triglycerides that are found within the muscle will move closer to the mitochondria itself, facilitating and providing an easy source of fatty acids that are able to easily make it into the mitochondrial matrix for beta-oxidation and oxidative phosphorylation. So virtually at every single step of the process of getting fatty acids from their triglyceride form in adipose tissue and in IMTG form, there is a benefit to endurance training that favors the increase in fatty acid oxidation observed when we look at papers such as the one that we read a couple of weeks ago.

So what about eating fat as a way to supplement energy either before or during an exercise session, similar to the way in which carbohydrate is used? So I think we have to ask ourselves that question and investigate whether or not there's any reason to advocate for the ingestion of fat before or during an exercise session and whether that would have any impact on performance. And so we should first ask the question: what would be the point of eating fat before or during exercise? And if we think about metabolically why that might be advantageous, a couple of reasons come to mind. The first is just that by eating fat, we ultimately are going to break that fat down theoretically and increase the availability of fatty acids because of the fatty acids that are coming from that exogenous source of ingested fat or triglyceride, and ultimately if we increase fatty acid availability, we would hope that that in some way would increase our ability to oxidize fat and ultimately allow us to rely more on fat and spare some of our stored carbohydrate. So you might expect that this would reduce muscle glycogen breakdown during exercise.

However, if we look at studies that have investigated the role of ingesting fat before or during a bout of exercise, we ultimately see that studies show that this doesn't improve performance at all, especially not when we look at the dramatic effects that can be seen from ingesting or even just mouth rinsing with carbohydrate on a bout of exercise performance. So why do we not see these improvements in performance from ingesting fat? And a lot of it has to do with a lot of the metabolic rush now that we've set up previously in this lecture. And so one of the things that we have to consider is that per unit of mass—okay, so if we have 10 grams of fat versus 10 grams of carbohydrate—we know that fat is going to have a much higher energy content, and as a result, per unit of mass, it's going to empty from the stomach at a much slower rate than carbohydrate. So per unit of mass, it's going to make its way from the stomach into the intestines for absorption at a slower rate, and so that's going to limit its utility as an ergogenic aid or as something that would benefit acute exercise performance before or during. The other thing that we have to account for is that, as we've spent a lot of time discussing today, there are a lot more steps involved in absorbing fats and carbohydrates. We have established that fats need to be transported into the bloodstream, make their way across the muscle cell membrane or the sarcolemma, and then also make their way across to mitochondrial membranes in order to ultimately undergo the longer process of beta-oxidation and oxidative phosphorylation. So in general, in terms of increasing the availability through eating more fats, well, first of all, remember that we're not necessarily limited by our fatty acid availability; we're limited more by our ability to transport it and use it. There are more steps involved in the absorption of fatty acids. The other problem is that most dietary fats, as we have established previously, are found in the form of long-chain triglycerides, and these long-chain triglycerides, because of the size of the molecules when they're eaten, they reach circulation at a much slower rate than carbohydrates do. And the reason for this is because in order to make it into circulation, because of the size of these molecules, they can't just diffuse through the lumen and into the bloodstream as readily as carbohydrates; they have to be packaged into these compounds known as chylomicrons. And we know that the triglycerides from chylomicrons contribute very little to expenditure or energy expenditure during exercise. So because they need to go through the lymph before they go to the bloodstream, it takes a lot longer to get these long-chain triglycerides into the bloodstream itself. And then once that happens, very little of those triglycerides in that form are going to actually contribute to energy expenditure during exercise.

So this also brings them into question: well, if the issue—or a lot of the issue—with ingesting fatty acids and their ability to directly contribute to a subsequent or ongoing exercise session—well, is the same thing true when you look at medium-chain triglycerides or MCTs, given that medium-chain triglycerides actually, because they are smaller, don't need to enter via the lymphatic system? But I don't want to spoil that because we're going to talk about medium-chain triglyceride supplementation and look at whether or not there is any potential benefit to exercise performance from ingesting medium-chain triglycerides as an ergogenic aid versus long-chain triglycerides. So that will be discussed in Chapter 11 later in the quarter in our discussion of supplementation.

I want to shift our focus away from fat as a fuel source during exercise and some of the underlying physiology or the need to intake fat before or during exercise to this idea of adopting a low-carbohydrate, high-fat diet. Low-carbohydrate, high-fat diets are by far one of the most debated and currently researched items in sport nutrition, and there have been quite a number of studies recently that have looked at these diets as a means not only to lose weight, but it has been suggested that long-term adaptation to a low-carbohydrate, high-fat diet may have potentially beneficial performance outcomes for different types of athletes. And I want to set some of that up and look at some of the literature on what happens to performance when these low-carbohydrate, high-fat diets are adopted. And so I think the first thing that we need to do before we look at studies that have investigated the performance effects of low-carbohydrate, high-fat diets is sort of define what a low-carbohydrate, high-fat diet looks like. And there are certainly varying extremes and approaches to low-carbohydrate, high-fat diets, which is one of the reasons why studying them in the context of not just weight-loss interventions but as potential ways to improve performance becomes difficult is because there is some individual variability in how these diets are carried out. But in general, if we look at most low-carbohydrate, high-fat diet area approaches, one of the ways to operationally define these is to look at the relative macronutrient contribution or the percentage of overall energy intake that's coming from carbohydrate, fat, and protein when trying to operationalize what a low-carb, high-fat diet is. And it's been generally accepted that a diet can be considered low-carbohydrate and high-fat if it has less than 20% of its energy coming from carbohydrate. Now, some approaches—some of the extreme ketogenic diet area approaches—may push dietary carbohydrate contribution to as low as 5% of energy or less, but I think we'll—were thinking about athletes and active individuals—a pretty solid number that has been defined is less than 20% of energy coming from carbohydrate; it's a pretty low-carbohydrate diet for these types of individuals. Now subsequently, if it is a low-carbohydrate, high-fat diet, there also needs to be some criteria in terms...

Of the contribution of fat as an energy source to the overall energy intake for a given day, and so most low-carbohydrate, high-fat diets will not only have individuals go below 20 percent of their overall energy intake coming from carbohydrate, but as a result, they will also shift more of that energy to coming from fat. So more than 50% of energy in these diets is coming from dietary fat. And then what's obviously highly variable, depending upon how far below 20% carbohydrate and how far above 50 percent fat the individual's diet goes, the amount of protein is going to be highly variable, albeit an important factor to consider, especially when we're thinking about some of the performance impacts of these diets.

Now, as you may have seen, there are many cases where people adopt low-carbohydrate, high-fat diets successfully. Certainly, people are able to withstand this, the low current of low-carbohydrate diets and don't die; they're able to survive these types of—and this begs the question of how. Because if we think about the fact that the brain, specifically in the central nervous system's favorite energy source is glucose, and that it primarily gets that glucose from the blood, which is tightly regulated, well then how do people survive? How do we metabolically make up the difference if we are supplying carbohydrate at a relatively low rate? How do we keep blood glucose tightly regulated, and how do we still, in potentially some of the gaps where carbohydrate availability might be falling short due to the dietary challenge that we're placing on it when adopting one of these diets? And so this is where the process of ketosis comes into play.

And so I want to walk you through a few steps of what happens when you limit carbohydrate availability via a long, low-carbohydrate, high-fat diet. One of the things that happens over time is, as you go on in limiting this carbohydrate availability and adopting a low-carbohydrate, high-fat diet, one of the first things that's going to happen in order to resupply blood glucose, if we remember, is we're going to have to undergo gluconeogenesis. So in order for blood glucose to not drop to dangerously low levels, we know that the liver is able to resupply some of that glucose, either by making new glucose via gluconeogenesis or from breaking down liver glycogen. But ultimately, what happens as this continues to occur and we limit our other forms of available carbohydrate, eventually hepatic glycogen stores or liver glycogen stores are going to become depleted. And one of the things that ultimately is going to happen is we're going to break down more fatty acids; we're going to beta-oxidize more fatty acids, and as a result, now we're going to have more acetyl-CoA that is formed. So we're calling to ultimately need to take the acetyl-CoAs that are being generated from the breakdown and fueled by fatty acid breakdown and run them through the Krebs cycle so that they can undergo further oxidative phosphorylation. But one of the problems is this is great, right? We have all—we have more acetyl-CoA being generated that's coming from fatty acids, so we're using more fat as a substrate and fuel source and breaking down storage fats in order to make this happen. But because one of the things that needs to happen in order to supply—until—to keep blood glucose stable is we need to recruit some of our Krebs cycle intermediates, specifically oxaloacetate, in order to supply blood glucose. So when we limit blood glucose and we limit our hepatic glycogen stores and deplete them, more oxaloacetate is then drawn into gluconeogenesis, and this is a problem because that oxaloacetate is ultimately necessary for these acetyl-CoAs, which are now being formed at an even higher rate due to an increased need to break down fatty acids to go through the Krebs cycle. Bit so those oxaloacetates are being now drawn into gluconeogenesis, and this is related to the whole “fats burn in the frame carbohydrates” concept because less of that oxaloacetate is now available in the Krebs cycle; well, then ultimately depletion of this intermediary is going to impede the entry of these acetyl-CoAs into the Krebs cycle. So now we have this abundance of acetyl-CoA being formed that can't ultimately go through the Krebs cycle, and we need to have something somewhere for these acetyl-CoAs to go. And ultimately, the way that we deal with this is that these acetyl-CoA molecules—this extra acetyl-CoA—will reach a critical point in which they then need to be condensed, and they're condensed actually in the liver to form what are known as ketone bodies. And there are three primary ketone bodies: acetoacetate, beta-hydroxybutyrate, and acetone. And these ketone bodies are a way to condense the acetyl-choline molecules that are being formed due to the increased need to break down fatty acids in the absence of hepatic glycogen and carbohydrate availability. We do this to condense those acetyl-choline molecules and use them as a fuel source. And so these ketone bodies are convenient and a way to fill in some of the metabolic gaps that might be left from the depletion of carbohydrate because they can cross the blood-brain barrier; they're able to be easily used as a fuel source in a similar way to which glucose is able to be used as a fuel source. So, in turn, ketone bodies are a major fuel source for the central nervous system; they're able to cross the blood-brain barrier, and they're also able to enter skeletal muscle and be oxidized as a fuel source by skeletal muscle. So ultimately, we end up making these ketone bodies as a way to fill in some of the metabolic gaps that are left from the availability of glucose. And when we adopt a low-carbohydrate, high-fat diet, we—we ultimately, over time, it has hepatic glycogen stores become depleted, and more oxaloacetate is drawn into gluconeogenesis to supply blood glucose; we end up having a buildup of acetyl-CoA outside the Krebs cycle, and some of that acetyl-CoA is converted into useful—useful metabolic form known as these ketone bodies. And ultimately, once these acetyl-CoAs make it to either the brain or the mus—or once these ketone bodies make it to either the brain or skeletal muscle, they can—once they enter that mitochondria, they can be converted back into acetyl coenzyme A, where they can undergo subsequent oxidation.

So this is sort of the physiology underlying how the body ultimately adapts to a low-carbohydrate, high-fat diet without dangerously dropping blood glucose or having inadequate energy for the central nervous system and skeletal muscle to perform the work that needs to be done. Now, given this underlying physiology and the fact that in scenarios like a low-carbohydrate, high-fat diet where carbohydrate intake is restricted over a longer period of time, we should—we see an effect on fat mass, given that in order to supply the energy needed, we're going to have to break down more fatty acids in order to form acetyl-choline and ultimately ketone bodies. Should there be an effect on fat mass when an individual undertakes a low-carbohydrate, high-fat diet? And so this is only recently begun to be investigated in athletes, but if we look at—compared to traditional diets—when we control for calories, we will see, in the short term—this is from a study in mountain bikers where they had them follow a more standard mixed diet over a period of, I think it was four to six weeks, and then in randomized crossover fashion had them keep everything about their training and diet the same except for shifting the macronutrient content to a ketogenic diet or a low-carbohydrate, high-fat diet, and then they looked at their body mass as well as their percentage of body fat in comparison at the end of each one of these periods. One of the things that we'll see across many studies is that there typically is a reduction in body mass observed in the short term when individuals undertake a low-carbohydrate, high-fat diet, but is all of that due to a reduction in body fat? And the answer is that some of it is, but some of that initial loss of weight is also due to changes in water. And the reason why this happens is because when you restrict carbohydrate over time, especially in individuals like the ones we're looking at here who are going to be engaging in relatively large volumes of exercise, one of the things that's going to happen is you're going to deplete your storage of muscle and liver glycogen, and we know that one of the things about a molecule of glycogen—if we think about—if we think about the chemical structure of glycogen is C6H12O6—well, with each molecule of glycogen that is stored, we also are going to then store six molecules of water. So when that glycogen is depleted, all of that water also is gone. So some of the initial loss of weight, and one of the reasons why you'll see individuals lose quite a bit of weight in the first few weeks on a low-carbohydrate, high-fat diet, is due to a loss of water. But in the short term here, as we'll see in this study, there is also a reduction in body fat that can be observed as well. In this particular study, these participants reduced their body fat percentage by around three—three and a half percent, and that occurred pretty rapidly, again like a four-to-six-week period. But if we were to look at weight loss studies of low-carbohydrate, high-fat diets over the long term, we would see that without further energy restriction, i.e., without creating enough of a negative energy balance, these initial losses in weight and fat would start to level out as time goes on. So while this might be an effective short-term strategy at reducing body weight and maybe in reducing body fat, if the goal is long-term fat loss, these diets have not been shown to be any more effective than a well-controlled moderate energy restriction diet. But we'll save a lot of the discussion—this is a weight loss diet—for our unit on weight management, but this is really where this type of diet began to work its way into the performance literature. And if we're thinking about it in the utility for athletes, it might be useful if you're seeking a short-term reduction in weight—let's imagine you're trying to—to make weight for a weight class-specific sport—or if you're in a period of time where you're trying to cut weight, but not just fat, and you're trying to maximize the amount of fat that you lose—I'm thinking of like a bodybuilding competition—there might be some benefit to restricting carbohydrate intake during that period of time. But overall, as a long-term weight loss strategy, if you don't also control for energy deficit, it's not going to be any more effective than a well-controlled moderate energy restriction diet.

Now, fat mass is obviously just one side of the equation when it comes to body composition, and so if we now begin to think about the underlying physiology and its potential effects on lean mass—which obviously matters for a lot of athletes—or most athletes as well—what might be effects of restricting carbohydrates on lean mass? And so this is a—this is a question that's a lot more difficult to answer because it's multifaceted in nature. And if we take a look, for example, at a study that was done in 2010, this was done in overweight subjects, so not well-trained athletes, and these overweight subjects were fed a hypocaloric diet, so they created an energy deficit, and they were fed a low-carbohydrate, high-fat diet. And so that's the group here that is going to be depicted in the blue bars. This group of overweight subjects had a low-carbohydrate, high-fat diet, fed a hypocaloric amount of energy in order to induce weight loss, and they were also combining this diet with resistance training. And this was compared to a group of individuals who were participating in the same—in the same resistance training program but were not undergoing the low-carbohydrate, high-fat diet. So they were—they were given a standard kind of mixed diet; it would be a more normal breakdown of macronutrients. And what we can see here, if we look at the change in body weight—so this is the overall change in weight—this is the overall change in fat mass—so how much of that weight change was due to changes in fat—and then how much of that weight change was due to changes in lean mass—if we look at what happened in these groups of individuals, we'll see that the individuals who were given the low-carbohydrate, high-fat diet, when compared to the individuals who just did the exercise program, we'll see that the individuals on the low-carbohydrate, high-fat diet had significant reductions in body mass and—and fat mass, as we might expect to happen based off of the fact that these are overweight subjects and based off of the underlying physiology. But one of the things that we'll also notice is that these individuals, when they combine this diet with resistance training, they didn't lose much lean mass, but they also were not able to gain lean mass. So they maintained their lean body mass; they didn't lose any, but they were not able to gain lean mass, which could be seen as a negative here because one of the things that we're trying to do when resistance training many times is to build lean mass. So in some way, shape, or form, these individuals were limited when they were adopting the low-carbohydrate, high-fat diet from gaining lean mass. So what might have limited the individuals on the low-carbohydrate, high-fat diet from gaining this lean back? Well, we then need to also think about if we are decreasing the availability of carbohydrate, we're not only probably going to increase the reliance on fat, but although it represents usually a relatively small proportion of overall energy expenditure during exercise, we're also now going to increase our reliance on amino acid metabolism. And so we'll talk about in our unit on protein and amino acid the different ways in which amino acids can enter the oxidative pathway and how reducing carbohydrate availability might increase our reliance on amino acids to provide either an oxidative fuel source or to serve as a way to supplement gluconeogenesis and preserve blood glucose. But on these diets with low carbohydrate availability, you're going to increase the need to break down amino acids, and sometimes some of that comes from the breakdown of lean tissue or amino acids being removed from the free amino acid pool for energy that can't be used to then serve as building blocks for muscle tissue. So one of the things that was key and why this study didn't observe an increase in lean mass is a—it may be because those individuals were increasing their reliance on amino acid for metabolism, but they also were fed a hypocaloric diet. Remember that this is an overweight group of subjects where weight loss is the goal, so in addition to manipulating the energy content of their diet, they also needed to put them at a negative energy deficit. And so this doesn't leave enough energy surplus for a lean mass accrual, but also what typically happens when you have a hypercord diet is you then decrease the absolute amount of protein in the diet that is being in taken. So if we went back and looked at the diet of the individuals in this study, they were consuming less than one gram per kilogram of body weight per day when adopting the low-carbohydrate, high-fat diet. And so usually protein intake scales pretty well with energy intake, so in these situations, we might need to increase the dietary content of protein because of the increased reliance on amino acid metabolism and the potential need to break down amino acids and use them to stave off some of the breakdown of lean tissue. So does that happen? So this was a subsequent—this was a study where they looked at carbohydrate restriction—a low-carbohydrate, high-fat diet—in the context of exercise. In this case, they were performing both aerobic and resistance training, and in this particular study, the goal was to make sure that the individuals were being fed enough calories to maintain their body weight, but also that they were being fed enough protein. So if we look at the results from this particular study, but we can see that compared to the control group, we saw a significant reduction in fat mass when these individuals were being fed enough calories and protein, as just as we did before. But you also now see that in response to this diet and exercise program that the individuals who were undergoing the low-carbohydrate, high-fat diet were also able to now gain lean mass, as you would expect—as an anticipated outcome from the resistance training component of the exercise training program. And these increases in fat-free mass were occurring as long as an adequate amount of protein is being consumed, and we'll look at the specific recommendations for protein intake for individuals who are trying to gain lean mass on a normal diet. But one of the things that changes about that is when you increase the reliance on amino acids as a substrate for metabolism, those amino acids are going to be coming from the breakdown of lean tissue, and less of the amino acids that are in the free amino acid pool can be used to build new tissue. So more protein intake is recommended for individuals who are adopting these types of diets but are seeking to also increase their lean mass as a function of a low-carbohydrate, high-fat diet as well. And so the article that you read gives a pretty large range, but a range on the less recommended 1.3 to 2.5 grams of protein per kilogram of body weight a day if we mass accrual during a period of low-carbohydrate, high-fat diet adoption is necessary.

Now, as you might have imagined, this picture is certainly more complex than just the effects of this type of diet on body weight. And so while these might be good short-term solutions for reductions in body weight and maybe even a little bit of loss of body fat, these diets are really starting to work their way into the endurance performance world. And if that's the case, the question then becomes—other than just losing fat—assuming that most endurance athletes have generally pretty low levels of body fat to begin with—or let's imagine that they use a diet like this to decrease their fat mass in the short term, early in the season—are there other adaptations, other than fat loss, that may either benefit or hinder performance in these types of athletes? So for right now, we're thinking about endurance athletes, and we're thinking about exercise that is performed, you know, around or below 75% of VO2 max, where the majority of that energy could be supplied by fatty acids. And so if we start to think about what are some of the ways—other than just losing body fat—that a low-carbohydrate, high-fat diet might improve endurance exercise performance, we might arrive at something such as—well, the increased ability to use fat as an energy source. It's possible, by restricting the availability of carbohydrate, that we train our bodies to rely more on fat as an energy source and maybe get a similar training adaptation onto the one that we see when an individual undergoes a period of endurance training. And so if that occurs, then potentially we can rely less on carbohydrates, spare our storage of muscle glycogen during exercise, or use less muscle glycogen during exercise, and potentially, because of the increased availability of fatty acids in the diet, increase our storage of intramuscular triglycerides. So these are some ways that potentially metabolically a low-carbohydrate, high-fat diet might benefit endurance performance. But we also have to consider that there are several related ways in which a low-carbohydrate, high-fat diet might hinder endurance exercise performance, and these are things such as increasing the risk of hypoglycemia. And given that we know that we're gonna have to limit the intake of carbohydrate and potentially challenge blood glucose to a greater degree during exercise, especially exercise at higher intensities, there may be an increased chance of incidents of hypoglycemia or low blood glucose due to the low availability of carbohydrate or content of carbohydrate in the diet. The problem may be that although we used less muscle glycogen, we also may have less muscle glycogen available. So by limiting the intake of carbohydrate, you may decrease your ability to replenish and store muscle glycogen. Some other things that could potentially occur because of the negative effects of limiting carbohydrate availability potentially on the central nervous system are increased feeling of fatigue. So this could be related to both the prevalence or risk of hypoglycemia as well as the decreased ability to store muscle glycogen, which ultimately may manifest itself into greater perceived exertion—having less left in the tank at the end of an endurance bout. And then, of course, if we reduce the availability of carbohydrate, while we can recruit fatty acids to help—or try to fill in some of that gap—or potentially even more ketones to help fill in some of that gap, we know that when the intensity of exercise shifts to higher gears that ultimately we're going to need to recruit our anaerobic energy pathways in order to supply energy at a rate that is commensurate with the demand of the exercise, and we know that after we've gone through our very limited phosphocreatine and stored ATP stores that we accomplish that largely by anaerobically breaking down carbohydrate.

So, if you have fewer or less glycogen available, it could potentially impair our ability to break down carbohydrate and aerobically, especially given that anaerobic glycolysis typically relies very heavily on glycogen, specifically muscle glycogen. So there are some ways in which this could be metabolically advantageous, and there are also some ways in which it could become a metabolic hindrance, each of which potentially exerting different effects on performance. So let's take a look at studies that have been done on the short-term adaptation to a low-carbohydrate, high-fat diet and how it impacts performance.

And so the initial studies that came out looking at this were done in some models where low-carbohydrate, high-fat diets were adopted over a one- to three-day period. In this particular study, what they had individuals do was they had individuals go on a three-day low-carbohydrate, high-fat diet or a high-carbohydrate diet, something that would be more indicative of what's normal, the normal diet of an endurance athlete. And so they had these individuals, in these cases, do different 120-minute cycling bouts. So basically, this was a two-hour trial in which they were asked to cover as much distance in 120 minutes as they possibly could, maintain as high of an intensity over this two-hour period as they could. They did so in a randomized crossover fashion with some washout in both dietary conditions: a high-carbohydrate and a high-fat condition.

What they looked at in these models was how the individuals' intramuscular substrates changed as a result of both these diets before the ride, after the ride, and 24 hours after the ride. And then, of course, they looked at their metabolic responses and performance during the 120-minute exercise bout when adopting these dietary strategies. And so this has been corroborated by multiple studies in terms of what these individuals found. But if we take a look at the primary findings from this study, we'll see that when practiced for a short period of time, low-carbohydrate, high-fat diets have, first of all, been shown—as you may expect them to—increase the storage of intramuscular triglycerides.

So if we take a look at the intramuscular triglyceride content of these individuals before the ride, immediately after the ride, and 24 hours after the ride when they were still undergoing these dietary approaches following the ride itself, the individuals had significantly more intramuscular triglycerides stored when they ate the high-fat diet. So by increasing the availability of fat in the diet, they were able to restore and even top off their storage of intramuscular triglycerides to a significantly greater degree than those who were adopting the high-carbohydrate diet. If we look at what happened in response to muscle glycogen, we can see, first of all, that the individuals came in with pretty similar levels of glycogen before the actual ride itself, and that both groups significantly used a lot of that muscle glycogen, and really to the same degree. So they came in with similar levels of muscle glycogen before the diet and before the ride; they used the same amount of muscle glycogen during the ride. But one of the things that we saw following the ride is, if we look at their ability to replenish their muscle glycogen, the individuals with the high-carbohydrate diet were able to, within 24 hours, replenish their muscle glycogen to levels that were much, much closer to their pre-exercise levels, whereas the individuals on the high-fat diet, because of limited availability of carbohydrate in the diet, were not able to restore their muscle glycogen to the same degree. So they had increased ability to store fat, closer to baseline, but they had an impaired ability to store carbohydrate.

We can see that kind of playing itself out here. The third thing that we can see from looking at the respiratory exchange data—so this was from data collected from a metabolic cart during the two-hour exercise session—we can see that the high-carbohydrate group had a significantly higher respiratory exchange ratio during this ride compared to the high-fat group. And this is probably due to an increased reliance on fatty acids, but also could be due to an impaired ability to break down carbohydrates. So this could be good for glycogen storage over the long term, but one of the things that unfortunately is noted—likely due to impairments in the anaerobic breakdown of glycogen and also potentially due to impaired glycogen replenishment—is that, as the exercise session went on, the high-carbohydrate…if we look at this, this top panel here, this is the percentage of their VO2 max, or the intensity that they were able to maintain across this two-hour bout of cycling, and what we see is that over the course of this two-hour bout, the high-carbohydrate group was able to maintain their higher intensity of exercise. So they're somewhere, as we can see, around 75 or 80 percent of their VO2 max, and they're able to sustain that over the entire two-hour bout without much decrease, whereas in the high-fat group, they're able to maintain their intensity to a degree, but somewhere around 60 percent of the total time. As we go into the later stages of this session, that high-fat group was not able to maintain as high of a percentage of their VO2 max or maintain as high of an intensity later in the session. So this impaired performance is likely due to potentially an impaired ability to break down carbohydrate and reducing the availability of carbohydrate storage in the diet and then the muscle itself.

So while this may potentially spare some…this may improve our ability to store fat and use fat, it may also impair our ability to store and use carbohydrate, which become extremely important, especially later in an exercise session, especially when intensity needs to be maintained or increased. So what happens when you restore carbohydrates periodically? One of the challenges to the findings from this paper is that the individuals were not given a chance to replenish their muscle glycogen. And so subsequent studies looking at this short-term adaptation in this particular fashion—again, this would be a randomized crossover trial—so this took place over the course of a week. In these particular individuals, they were again, in this case for five days, they were fed either a high-fat or a high-carbohydrate diet; they were given a performance test, the same 120 minutes at a fixed percentage of their VO2 max, and they were given these diets. Then, on the last day, on day six, the individuals given the high-fat diet were then given an opportunity to eat a high-carbohydrate diet the day before their performance trial. So you can see here everything was the same; their training was exactly the same in the six days leading up; the only difference here were the diets, the high-fat versus the high-carbohydrate diet. And then, regardless of the diet in the previous five days, they were given a high-carbohydrate diet and given a chance to replenish their muscle glycogen before undergoing this two-hour performance trial.

And what we can see when this, the results of this study were published, is that this five days with one day of carbohydrate restoration, we saw that again, in a similar fashion, if we look at this particular graph here, this is the amount of energy during that two-hour exercise trial that came from fat versus muscle glycogen and plasma glucose. We can see that significantly more fat was used by the individuals who were fat-adapted or had undergone this five days of a low-carbohydrate, high-fat diet. And in a related concept, we can also see that they were able to use significantly less muscle glycogen and carbohydrate overall, mainly because they were able to use more fat. And this happened even after the carbohydrate availability was restored. So by giving them a day to replenish their muscle glycogen, they didn't negate the effects of the previous five days of fat adaptation. So we can see that we're able to rely more on fat during exercise even when carbohydrate availability is restored. That shows up in the substrate utilization overall in terms of their absolute contributions to energy expenditure during the trial. It also shows up in, again, the same effect being observed where the individuals who were fat-adapted still had a lower respiratory exchange ratio over the course of the trial compared to the individuals who were on the high-carbohydrate diet. And again, we see a reduced reliance on muscle glycogen and an increased reliance on fat. But what we don't see is a clear performance benefit. So this was that time trial that they completed; so they were given, they were given a fixed distance to cover in a set amount of time, so a lower amount of time; that would…this would be the performance test. To cover that given distance would be a performance benefit. And so we can see the mean data here in the high-carbohydrate diet versus the low-carbohydrate, fat-adapted individuals, and what we can see is that in each condition—remember, this is a randomized crossover trial, so all of these individuals are serving as their own controls—so we see the means plotted here, but we also see the individual effects being illustrated by each one of these bars. And we have a pretty small sample size here, and we have one individual who there was a clear benefit of the fat adaptation in terms of his or her performance in that they completed the trial almost twenty minutes faster, but that's an outlier, and we have to kind of discount that and look at the overall average. And if we look at the overall individual effects, we see that there really wasn't much of an effect in performance.

So while these individuals were able to rely less on glycogen and more on fat during the exercise, there wasn't a clear performance benefit, and there also wasn't a clear performance hindrance. So physiologically, the tables are kind of set for some metabolic and metabolic advantages here, but they don't seem to really show up in the current literature as a performance…as having any performance-enhancing effects. And many of the individuals who are proponents of this diet will combat this research by suggesting that the reason that you don't see any adaptation to performance is because it takes longer than a few days for individuals to adapt fully from a metabolic perspective, from a muscular perspective, and from a performance perspective to these low-carbohydrate, high-fat diets. And so this is starting to be investigated more as time goes on. But one of the things that you can imagine is difficult in getting long-term data from individuals who are participating in low-carbohydrate, high-fat diets, one of the reasons being that many people have a lot of difficulty in sticking to low-carbohydrate, high-fat diets, especially in the Western world where carbohydrate is a pretty abundant source of food in a given society.

And so this study looked at individuals reporting to have been on a low-carbohydrate, high-fat diet for a long period of time. This was published three years ago in the journal *Metabolism*, and these individuals were, on average, following a low-carbohydrate, high-fat diet for about 20 months. So they took a bunch of endurance athletes, they pulled them on their normal diets, how long they may have been on their normal diets, and they broke them down into high-carbohydrate and low-carbohydrate intake. And what they found is that, not surprisingly, if we look at…if we look at the peak rate of fat oxidation in the individuals who adopted a low-carbohydrate diet versus a high-carbohydrate diet, their peak rate of fat oxidation was significantly higher in individuals who had spent an average of at least 20 months on the low-carbohydrate, high-fat diet. So they were able to, as we've seen before, drastically increase their peak rate of fatty acid oxidation. And additionally, what they were also able to do is they were also able to shift where that peak rate of fatty acid oxidation occurs. So in this particular study, where the peak rate of fatty acid oxidation in individuals who were adopting a high-carbohydrate diet, or what's more normally practiced, is somewhere between 55 or 60 percent of their VO2 max, the individuals in this study who were on a low-carbohydrate diet had shifted their peak rate of fatty acid oxidation to closer to 70% of their VO2 max. So they were able to rely more heavily on fats at higher intensities when adapting to these diets in the long term. And the other thing that was interesting to show is that when we look at their muscle glycogen concentrations, their muscle glycogen concentrations and their ability to restore muscle glycogen following depletion over the course of longer periods of time had been restored. So they were able to avoid the effect of…of impaired glycogen restoration like we saw in the shorter-term study by periodically reintroducing carbohydrate, probably in the time after their exercise periods. So they didn't necessarily have this impaired ability to store muscle glycogen after long-term adaptation to the diet. But what this doesn't necessarily show still is a clear performance benefit. So while this might be metabolically advantageous, whether or not these changes in fat oxidation rate and shifts in fat oxidation rate make enough of a difference in performance after long-term adaptation to the diet has yet to be shown. But there is some evidence to suggest that this may be a way to rely more on fat and less on carbohydrates, but it does take a significant amount of time to adapt to.

Now the question becomes, well, what about high-intensity exercise performance or anaerobic performance? So one of the reasons why we may not see a performance hindrance, especially after an individual adapts to a diet like this long-term in endurance performance, is because, well, we don't necessarily need to rely a whole lot on anaerobic energy contribution, given that endurance exercise is typically performed at a low enough intensity where fatty acids can provide most of the energy needed. So in terms of looking at this for something like an anaerobic performance or short-term, high-intensity exercise performance, we may expect to find a completely different effect of a low-carbohydrate, high-fat diet. And so this is a recent investigation in which they looked at the effects on anaerobic performance. So these were athletes who…anaerobic performance is a big component of their sports. So they looked at this in trained male basketball players. And what they have these basketball players do is they gave them a lead-in period where they ate their normal diet. So you can see this would be something a little bit more similar to like 50 to 60 percent of the energy coming from carbohydrate, maybe 20 percent coming from protein, with the remaining 30 or 40 coming from fat. And they had them follow this for a week, and then they had them do a baseline Wingate test, so a test of anaerobic performance, looking at overall all-out effort on the bike, how much power they were able to generate over the course of a 30-second Wingate test. And then what they had the individuals do was follow a low-carbohydrate, high-fat diet for a period of seven days. And what they did in this particular diet—again, you can see the macronutrient contributions of each one of our macronutrients to the overall energy expenditure—we see now that fat is making up about 60% of the diet, carbohydrate is making up less than 20, and the rest is being made up by protein. So they had them follow this low-carb diet, and then they had them again perform the same anaerobic test. So they had them perform another Wingate test, and then they reintroduced carbohydrates; they carbohydrate-loaded them, gave them the ability to restore their muscle glycogen, and have them perform another Wingate test to see if carbohydrate restoration following a period of carbohydrate restriction had any subsequent changes in their performance.

So if we look at the results from this study, this table depicts their performance on the Wingate test in terms of their time to peak power, their peak power, and then, of course, what really matters: the total amount of work that they were able to do over the course of the 30-second Wingate test following their normal dietary practice, so their normal carbohydrate diet, or the baseline here, versus their performance on the Wingate test after a week on the low-carbohydrate diet versus their performance on the Wingate test after a week of reintegrating carbohydrate and loading carbohydrate, restoring muscle glycogen. And what we can see here very clearly is that, compared to following their normal diet, they were able to do significantly less work during the Wingate test when performing the Wingate test following seven days of carbohydrate restriction. But after just seven days of reintroducing carbohydrates and loading carbohydrates back into their diet, they were able to completely negate those negative performance outcomes. So they were now able to get their work rates back up to levels that were commensurate with where they were before the low-carbohydrate diet after a week of carbohydrate loading. So what we can see here is that there are certainly some short-term impairments in high-intensity exercise performance that occur when undergoing a low-carbohydrate diet, but that those performance decrements can be restored simply by adding carbohydrates back into the diet. So if these athletes are looking to periodize their training in a fashion that favors maybe adopting a low-carb diet for body fat loss or weight loss during a certain time of the season, but they're concerned that they're not going to be able to restore their high-intensity exercise performance, we can simply do that by restoring carbohydrate back into the diet or loading carbohydrate back into the diet, and those performance outcomes, those negative performance outcomes, may potentially be saved off by doing so.

So what about things like weightlifting? Something like a Wingate test is a good test of anaerobic endurance, the ability to generate power in the short term over periods of, say, 30 seconds to a couple of minutes before our oxidative pathways begin to take over. But what happens in super high-intensity, short-duration exercise, such as powerlifting or Olympic weightlifting? Are there any compromises that occur to performance for individuals who are adopting these low-carbohydrate, high-fat diets? And so in this particular study—this was a study of high-level Olympic weightlifters or powerlifters—and what they had these individuals do was they took 14 of them, and they randomized them either into a low-carbohydrate, ketogenic diet for 12 weeks—which is a really long time to get athletes to commit to such a dietary approach in the context of a research study, so that's even kind of an impressive element of this study as well—or to maintain their usual diet over the course of that 12 weeks. Once that happened, these two…individuals were given a 14-day washout period in which they went back to their regular practices, and then they were asked to adopt the opposite approach. So all of these individuals, over the course of a 24-week study with a 14-day washout period in between conditions, completed both the low-carbohydrate diet condition as well as the usual diet condition. And what they wanted to look at was whether or not there were any effects on their performance of key tests that would be related to powerlifting.

So if we look at the main outcome here, which was their change in lifting performance—so this would have been an aggregate of their one-rep max for key lifts, such as like the bench, the deadlift, the squat, the power clean—and we look at their performance of these key lifts as a function of change following the usual diet condition versus the low-carbohydrate condition, we see that these would be the overall averages here; we see all the individual data plotted; we see that as a function of the average of these populations, which is represented by these flat bars, there is really no change in their lifting performance. So they were able to maintain their maximal strength and power production whether or not they were following their usual diet or a low-carbohydrate, high-fat diet. So no change in maximal strength and power performance and no decrement seen here. And that makes sense physiologically because if we go back to our fuel sources for metabolism, we'll see that, as you would expect for something like a maximal-effort weightlifting exercise or a power lift where the overall duration of contraction is very short, but that the rate of ATP resupply is high, the majority of that energy can be supplied by the anaerobic breakdown, specifically of phosphocreatine and stored ATP. These aren't carried out for long enough periods of time to really even considerably need to recruit the anaerobic glycolysis of carbohydrates. So metabolically, because the metabolic outcomes of a low-carbohydrate, high-fat diet don't really play themselves into causes of fatigue during maximal power and maximal strength efforts, we don't really see much of an effect on performance when looking at the effects of low-carbohydrate diets on this type of exercise.

So let's summarize what we know about low-carbohydrate, high-fat diets and their effects on performance. There's still a lot of work to be done in this area. This week's research article that I'm asking you to read is a review article that will summarize some of the things that we talked about in the second half of the presentation, but maybe highlight some of the findings from some of the individual studies and set the stage a little bit more for what isn't known and what needs to be investigated in the context of the effects of these diets on performance. But in…

Terms of summarizing what we know so far: first, we know that these can be effective strategies in causing rapid weight loss. Some of that weight loss is due to water loss due to glycogen depletion, but there is an acute effect on fat that can't be ignored, and that you might be able to also maintain lean mass while losing fat mass if adequate protein and energy are consumed when practicing a low-carbohydrate, high-fat diet.

One thing that we definitely showed through the studies that we examined was that low-carbohydrate, high-fat diets, in the short term, tend to increase the availability of lipids as a substrate, but also can confidently reduce the storage of glycogen and carbohydrate as a substrate. And as a function of that, fat oxidation increases during exercise. But fatigue resistance and exercise performance have been shown to be compromised in the short term when adopting a low-carbohydrate, high-fat diet.

In terms of long-term adaptation and carb reintroduction, these may both be effective strategies that may allow someone to better adapt to a low-carbohydrate, high-fat diet and stave off some of the decreases in performance that have been noted in previous studies. It takes a while; wary to take on, as in the study we showed, those athletes that were on long-term low-carbohydrate, high-fat diets have been on them for an average of 20 months—so a pretty long period of time. So a long-term adaptation may improve success with this diet, and then reintroducing carbohydrates periodically seems to erase some of the performance decreases that we've seen when low-carbohydrate, high-fat diets are adopted without periodically reintroducing carbohydrates and giving the opportunity to replenish muscle glycogen.

But one of the things that we don't have clear right now is our data regarding whether these changes in fat metabolism actually translate into performance benefits after long-term adaptation for endurance athletes. We also know that, because of the effects on the impairment of anaerobic breakdown of carbohydrate and potentially limiting source of carbohydrate, that aerobic endurance performance, such as one like we saw with the basketball players and the Wingate test, is impaired when low-carbohydrate, high-fat diets are adopted, but that those impairments can be staved off once carbohydrates are reintroduced into the diet.

Now, one of the things that is clearly not well known, but I'll give you a chance to investigate in the context of the research article as well as the sport nutrition blog prompt that is associated with that research article, is data on repeated high-intensity bouts of effort. So think about this: you can think about this like an ice hockey player, which is the example that I'm going to give you to work with this week, where there are going to be not just one high-intensity bout of an aerobic endurance performance, but potentially multiple occurring over the course of a given competition. Data on these high-intensity, repeated bouts of effort are less clear in terms of the effect of low-carbohydrate, high-fat diets and are definitely something that warrants further examination, which again will be discussed further in the research article that I'm having you read.

And then, in terms of maximal strength and power, given that carbohydrate availability is not necessarily a limiting factor for these types of exercise, adopting a low-carbohydrate, high-fat diet doesn't seem to really have much of an impact on maximal strength and power production.

So this is a little bit of a summary of what we know so far. I want you to now go ahead and read the review article on the low-carbohydrate, high-fat diets and exercise performance that will go over some of these things in more detail, as well as post some new questions, and then complete your sport nutrition blog posts for the week.