Welcome to the Empirical Cycling Podcast. I'm your host, Kolie Moore, joined as always by my co-host, Kyle Helson, and I want to thank everybody for listening, so please subscribe to the podcast if you have not yet done so already, and please consider telling a friend and sharing the podcast. That really helps us out and gets the word out, especially if you like the kind of stuff that we're doing. If you'd like to give us an iTunes rating, that would be really fantastic, and a review, possibly. And remember that we're an ad-free podcast, so if you'd like to donate and support the show, you can do so at empiricalcycling.com slash donate. We also have the show notes on the website. And if you have any coaching and consultation inquiries, questions, or comments, you can please send an email to empiricalcycling at gmail.com. We also have merch at empiricalcyclingpodcast.threadless.com. So last episode, we started taking apart the Fick equation, and we discussed arterial venous O2 difference. So if you missed that podcast on AVO2 difference, I actually strongly suggest you check that out. If for no other reason than we're going to be talking about diffusion a little bit. And we're going to be seeing some of those concepts come up in this episode again. So if you're okay with diffusion already, that's cool. So my goal for this episode is to get ready to prepare to talk about the heart, which is going to be the second half of the Fick equation. So there's a lot in here that's crucial to understand about the dynamics of blood in order to understand the dynamics of the heart. and so this is all going to tie together at some point and then we're going to see everything as a whole picture and that's going to really let us know how should we best train VO2 max. So this week we're going to take a look at how oxygen is transported to tissues and we're going to talk about the roles played by hemoglobin and the blood and how training can affect these kinds of things. So Kyle, what do we know generally? about breathing and blood and hemoglobin, the three things that we're going to take a close look at today. I think a lot of people know that, well, breathing, they think of your lungs, your nose, mouth. Air goes in, air goes out. You breathe in oxygen, you exhale CO2. People know that your blood is what transports the oxygen, moves the oxygen around to your muscles, other tissues, and then also carries away waste CO2. And then of course, like we talked about last episode, hemoglobin is what binds oxygen and carries around and transports it from your lungs to your tissues, your muscles. Commonly, I think people talk about how, oh, like, oh, yeah, your lungs, they know something, oh, something about your lungs is where your blood is actually able to pick up that new oxygen or dump that waste CO2. Right, yeah. Yeah, yeah, and the lungs are the primary organ for gas exchange. So O2 enters and CO2 leaves. It's really that simple. But the thing is, like, it's also not that simple. because a lot of the structure of the lungs don't actually do anything because it starts with your trachea and becomes more and more branched until it reaches the tiniest little pockets of your lungs called the alveoli. Now, the upper respiratory tract has important functions for filtration, humidification, and usually warming, but sometimes even cooling, especially if you're in really hot temperatures. But the alveoli are the business end of the lungs for gas exchange. So they're really small sacs, and we have them because of, well, kind of like a shoreline or like a fractal problem. So, you know, Kyle, you're smarter than me, so why don't you talk about this and surface area and stuff like that, because we're going to get into the lung specifically in a second. So the idea behind a shoreline or this like fractal problem is that, okay, If you have the length, see, simple example is a beach. So if you're going to try to measure how long a beach is, you can say, okay, if I've got a ruler that's a foot long, and I measure the length of the shoreline, I'm going to get some length. But then if I have a ruler that's only an inch long, I can now contour the measurement to more of the little nooks and crannies along this beach. And I'm going to get a number that can be much, much longer. And then you can keep asking this question down to like, okay, so if I have a ruler that's like the width of a grain of sand long, how long am I going to measure the length of this beach? And that number can get really, really big, like many, many, many times, tens of times, orders of magnitude larger. And so if you actually ask the question, well, what's the length of the beach? It turns out that number is this fractal problem where it's a... number that sort of depends on the size of the ruler that you have. And a fractal is also famous where it's self-similar. So if you have a beach and then you, you know, that's kind of got this random sort of jaggedy pattern, if you zoom in to a smaller scale, it has a similar sort of random jaggedy pattern. And you can theoretically zoom in. infinitely and keep seeing that pattern repeat over and over and over again. Right. And so we can actually see kind of the same thing in structures like tree branches and lungs, right? Broccoli. So broccoli is also a good example. You have the little florets maybe that you eat, but if you've seen a big whole bunch of it, it looks basically the same, just bigger when you buy it in the grocery store as a big whole chunk. And then, you know, if you... Compare that to the structure of trees or whatever, like you said. It's a pretty similar sort of self-repeating pattern. And so when we want to apply this fractal thought pattern to the lungs and surface area, because, spoiler alert, it's the surface area of our lungs that lets us exchange gas very, very, very quickly. Let's say at max exercise, you're inhaling four liters per breath. If your lungs were like one single sphere, they would have a surface area of about 1,200 square centimeters, or like 0.1 square meters. And note that for spheres in this example especially, a sphere is the object that minimizes the surface area for a given volume. Right, and this actually presents us a problem. in the lungs. Because if we want to maximize surface area, how do we do it? Because spheres have little surface area for the volume. We could make it like tubes. We could have really, really thin, long tubes in our lungs instead of alveoli, little spheres. But then we run into the problem of we need surfactant and we need to make sure that it doesn't collapse, especially. And so the lungs have special structures that we're not going to talk about that helps them not collapse, but at the same time maximizes their surface area. So now if we had like two spheres of two liters each instead of one sphere of four liters, now we have a surface area of about 767 square centimeters per sphere, which means that we have 1500 square centimeters as opposed to the one spheres. 1200 square centimeters. So now if we start dividing it up, we're going to get more and more and more surface area. Yeah. And note that also spheres pack really nicely. So you can famously fill in a three-dimensional or a two-dimensional area or volume with spheres. When they touch, they can actually pack the most in. That's the... the shape that packs the most efficiently. Right. And so now that our lungs are so good at this fractal sphere thing that the approximate surface area of the average human's lungs is 50 square meters. That's crazy, yeah. Yeah, we were just talking about 1,200 square centimeters. like 0.1 square meters for like a single sphere, but we have about 50 square meters surface area in our lungs. And the reason that this is critical is that alveoli are lined with, they're very, very, very thin and they're lined with capillaries the whole way around, meaning that we now have drastically, I don't think I'm, I think I'm underusing that word, now we've drastically increased the amount of blood that can exchange Gas with the air in our lungs. But our lungs are actually so good at the surface area thing and also at being thin and presenting a lot of capillary surface to the inside of the alveoli that for this 50 square meters of surface area, we only have to put in about 200 milliliters or maybe three to four to... Percent of our total blood volume at once in the pulmonary system. That's really impressive. Just as a reference, you have somewhere between five to seven liters-ish of blood. Seven liters for a very large person, yeah. Yeah, depending how big you are. Yeah, typically, in this episode, we're actually going to look up total blood volume, and we're going to see numbers in the four to five and a half liter range. Andre the Giant. Yes, Andre the Giant has seven to eight liters of blood. But now we need to think about the nature of the exchange because reading articles and stuff like that in the popular press and seeing people explaining this on the internet, you know, the quote-unquote on the internet thing, I get the impression that people don't really understand what's happening in the LVLI for gas exchange. Biggest misconception is that when we exhale oxygen, people think that it's from our blood that quote unquote doesn't get used. And we're going to talk about hemoglobin in a little bit and this is going to make a lot more sense. But when we think about this, hemoglobin's purpose, the purpose of the blood is to move O2 oxygen from the lungs to tissue and give it safe harbor so it doesn't wreak free radical havoc on our vasculature and other places. Because if you remember the What's So Radical About Oxygen Wattstock episode, that goes into free radical damage and stuff like that. There's actually, we're going to do a follow-up episode on that at some point in the near future because there's a lot of really cool stuff in redox biology coming out. And I'm excited to get to that. But go listen to that episode if you haven't yet. It's very interesting in terms of potential damage of just oxygen wandering around. So there's two reasons that we breathe out oxygen. The first is that a lot of the air we breathe out is actually in dead space and not the alveoli proper. So that O2 is not going to go anywhere. The second is that not all the capillaries of the alveoli are totally full of blood, especially when we're standing up. and near the top of the lungs since gravity tends to pull blood to the bottom of the lungs which is obviously not the case with like swimming and stuff like that or possibly recumbent cycling. So it depends on your posture. What about leaning over on a bike, right? You're not exactly sitting up. Yeah, I guess if you're in a really aggressive TT position that would help as well. So hemoglobin is actually going to take up what it needs very, very Very Quickly. So when it leaves the lungs, it's basically 100% saturated, like 98, 99%, something like that. Helped in no small part by hemoglobin's cooperativity, where the more O2 binds, the more O2 it wants to bind and the faster it will bind. And we're going to get into that in great detail in a little bit. But if we imagine that we actually breathed out O2 from our blood, this means that We would be losing O2 saturation from our blood whenever we exhale or half the time that we're breathing. And if we just make a rough assumption that the heart pumps about twice as many times per second as we breathe, and yet O2 saturation remains very high consistently, we can pretty well see that no, we're not actually breathing out any oxygen from our blood. Yeah. And we kind of talked about this last episode too, where your blood is actually Not able to deliver all the oxygen that your muscles would be willing to use. So if your blood was still maintaining some of that oxygen, that completely contradicts what we talked about last episode. Yeah, completely. And also, if we think about what would happen if we did breathe out oxygen, we would always be wanting to take very short and forceful exhales and long, slow inhales. Otherwise, if we just breathe normally and our heart's pumped normally, we would actually get a sinusoidal saturation of oxygen in the blood. And that would probably be not good for exercise, I'm going to assume. Hemoglobin is able to become fully saturated very quickly, like I said. So the O2 pressure in the lungs is about 160 torr or 160 milligrams of mercury. But venous O2 pressure is 40 torr or possibly even less. So the diffusion happens very quickly into the lung capillaries in the alveoli. But I think we had mentioned actually that air pressure in one of the last episodes, or possibly several of the last episodes, air pressure is about 760 torr. But we didn't note, and we're going to note now, that this is all of the gases combined. Right, yeah. Atmospheric pressure of nitrogen is like 570 torr. So oxygen pressure is actually quite a bit lower itself, but it's just a general idea of, you know, that it's all partial pressure. What's really happening with the gas in the lungs themselves, like what's actually happening? So the O2 is diffusing into the blood like a normal diffusion, like we talked about in the last episode of O2 diffusing from the blood into tissue. The regular Fick diffusion equation that we talked about last episode applies directly right here, where gases are going to move from a higher to a lower pressure passively thanks to entropy. and the attempt to equilibrate the gas pressures of, well, gas literally everywhere. So you can imagine if you hold your breath long enough, the partial attitude pressure in your lungs is actually going to drop more and more and more because it's going to be like mixing in your lungs. And so everything that's up in your upper trachea, you know, that dead space is going to be mixing like Down, that oxygen is going to be diffusing down into your lungs, into your alveoli when you're holding your breath. So if you hold it until you're just about to pass out, you should exhale significantly less O2 in that breath than you would if you didn't hold it. Interesting note, it's actually the buildup of carbon dioxide that gives you that sort of like Tight feeling of wanting to exhale. So if you do actually exhale a little bit, you can actually hold your breath significantly longer because you prevent the buildup of that excess CO2. And that's one of the reasons why nitrogen is, if you breathe in heavy concentration of nitrogen with low concentrations of oxygen, you can't actually tell that you're not getting enough oxygen and you can pass out really easily. Yeah, and that's actually why carbon monoxide is so deadly, because it'll bind to hemoglobin and displace O2, and it binds more preferentially. And the thing is, we don't actually have O2-sensing apparatus in our regular vasculature. We have CO2-sensing apparatus. We have chemosensors, and that's what detects CO2. But it's not like... because in evolutionary terms we never came across carbon monoxide or huge pockets of nitrogen that much and so it really didn't matter. It mattered if you hold your breath and you've got CO2 there because of course if you breathe there's going to be oxygen. But now we can also understand why altitude would make it harder to breathe and can lower hemoglobin saturation because the partial pressure of O2 at altitude Drops the higher you go. And this means that the pressure gradient in the alveoli versus the blood gets smaller and diffusion slows because of course the space is the same. But of course, if we want to look at pathologic conditions where there's like scar tissue in the lungs or there's liquid in the lungs, like you've got like schmutz in your lungs from like a... like bronchitis or something like that. Yeah, like that liquid is actually going to give the O2 a longer path to travel or the scar tissue is going to give the O2 a longer path to travel and that's what slows diffusion and affects lung capacity with those kinds of conditions. Don't smoke, kids. Don't smoke. Please don't smoke. So now let's take a look at what makes hemoglobin So awesome at not only taking up oxygen very quickly in the alveoli, but let's also let it go so when it gets to working muscles or tissues in need. That makes sense because if it was too good at binding, it would just hold on to the oxygen and not actually serve its function of delivering the oxygen anywhere. It would just bind and then sit there indefinitely. It's my oxygen. You can't have it. Hemoglobin doesn't do that. And so before we get really started on hemoglobin, I want to congratulate everybody listening to this because you all have the most interesting protein in the world and a lot of it. So we talked about myoglobin in the last episode and we mentioned the heme ring. So just a refresher, the heme has an iron in the center of it bonded to five nitrogens, which makes it stable enough to reversibly bind oxygen. Reversibly is the big key here because as we talked about last episode again, as opposed to when oxygen normally binds iron and creates rust. So we're not rusting. Internal rust, yeah. So myoglobin is a protein that has one heme group on it, and this protein is in the muscles. Hemoglobin has four heme groups, which is pretty cool in itself, but these four O2 binding sites have a distinct interactive ability on the protein that we're going to talk about in a second. So hemoglobin itself, has four what are called subunits. And it's two of each subunit. And we just call them alpha and beta subunits. Alpha 1, alpha 2, beta 1, beta 2. How creative. Yes. So it means it has like rotational symmetry. So it like alternates alpha, beta, alpha, beta in a circle. And each subunit has one heme group and each one combined one O2 molecule. So all the four subunits together make a hemoglobin. Alright, so hemoglobin displays what are called cooperative kinetics. Kyle, have you heard of cooperative enzyme kinetics before? I have not. Alright, so this is going to be fun. I'm really excited now. So the O2 binding curve of myoglobin is what's called a rectangular hyperbola. So imagine if you took the CP, the critical power curve, and you flipped it upside down. So it looks kind of like that, except it approaches the ceiling. And it's a very basic enzyme curve, and a lot of enzymes have this same curve. Hemoglobin, it has a sigmoidal oxygen binding curve. That means it's S-shaped. So the graph of hemoglobin binding to O2, this curve, we have it in the show notes. So go check that out. But to shortly describe it, what happens is starting at zero, if we're going to take the inverted CP curve, basically we would shoot straight up for a little while. Then we're going to like level off a little bit and then we hit our ceiling. Hemoglobin oxygen binding curve starts at zero and then it's going to go to the right and then it's going to slowly come up and then it's going to shoot up at one point and then it's going to slowly level off again and then it's going to gradually approach a maximum. So that is a sigmoidal or S-shaped curve and this curve means it displays what are called cooperative kinetics. So hemoglobin binding to oxygen changes the shape of the hemoglobin molecule itself. And this is where cooperative enzyme kinetics come from. So what does the cooperative kinetics mean in terms of hemoglobin and oxygen binding? It means as O2 binds hemoglobin, the shape changes a little bit in order to allow hemoglobin to bind more oxygen or vice versa. So as less oxygen is bound to it, it actually makes it easier to unbind O2 near tissue that needs it. This is awesome, right? Yeah, that's really cool. I should not know that. Yeah. Now, all right. Four of the five nitrogens that bind hemoglobin iron are on a big ring that's actually called porphyrin. It's porphyrin without the iron in the middle of it. It's heme with the iron in the middle. We mentioned this last episode. So the iron in this ring is kind of like a beach ball getting bisected by a trampoline with the fifth nitrogen. Binding from underneath. It's got four around it otherwise from the porphyrin ring. So the shape of the heme group, it actually changes shape, does physically change when O2 binds. So what's happening is without O2, the trampoline, this is where the trampoline analogy actually runs into a problem because it's, it... You know, it looks a little droopy. Like imagine pushing the trampoline down in the middle a little bit. This isn't actually what's happening. So what's happening is the beach ball in the middle actually has to be a certain length from the corners of the trampoline, if we imagine it as a square. Which means in order to make the lengths right, we actually have to move it out of the plane a little bit. The ring cannot expand because of all the other bonds that have to happen. So what's actually happening is it almost looks like the beach ball is kind of getting held up by roof rafters. So this quote-unquote trampoline takes on what's usually called a domed shape. But binding O2 changes the electronic state enough that it turns it into what's called the relaxed formation. And now the bond length actually changes a little bit. So now it becomes a flat trampoline. And the iron-porphyrin-nitrogen bonds actually get shorter by 0.1 angstroms and the doming subsides. So the shape change of the heme group means that the protein attached to the heme group literally moves in space and that moves the fifth protein that's actually attached to the iron underneath. And it's this movement. It's like 0.1 angstroms or something like that. It's tiny. An angstrom is 1 to the minus 10 meters, by the way. It's roughly the size of a hydrogen atom. Yes. Now, we're moving one-tenth the amount of the size of a hydrogen atom. And this... Can entirely change the structure of the whole hemoglobin molecule. The hemoglobin structure is so tightly coupled between the heme groups and the entire structure based on the grabbing of the heme group around it and that one nitrogen group from underneath that the whole structure changes based on O2 binding the heme and this takes microseconds. It is Almost instantaneous. And for all purposes, it practically is instantaneous. And roughly speaking, binding two O2 molecules to a hemoglobin is enough binding energy to change the entire shape of the whole hemoglobin molecule from what's called T to R. So there's taut and relaxed. Original again. Yes. Very descriptive. Yeah. So the T shape. is deoxyhemoglobin and it has low O2 affinity. Meaning if there's somewhere better for O2 to be, it's already got its bags packed up and it's ready to go. But the binding of 2O2 changes it to the R shape. So now if we have two oxygens on hemoglobin, it's going to... Now we talked about the T and the R, the taut and the relaxed for heme. It is very much the same for the entire molecule. So they just took the same name and renamed the two forms of the entire hemoglobin molecule T and R to reflect what's actually happening. But just because the entire hemoglobin is in the taut or the relaxed state does not mean all of the hemes are. So the hemes will still change shape individually based on O2 binding or not. So what's happening is the individual subunits actually do change shape a little bit, but two O2s binding is enough to change the entire shape. So the shape of the curve, this S shape, what it means is that the more O2s bound, the more hemoglobin, quote unquote, wants to bind O2. The less O2s bound, the less hemoglobin, quote unquote, wants to bind O2. So we can see this would be a huge advantage in tissue that requires a lot of oxygen. But we can also see that in the presence of a ton of oxygen, it would be easy to stick 202 onto the T-state deoxyhemoglobin and snap it into the R oxyhemoglobin shape and it's going to be even more ready to accept more oxygen. Oh, yeah. And that makes it useful because in those... Two different locations that we talked about last time, your lungs or your muscles, you have those, you have the exact opposite states that you would want. So you have the states that you would want because the oxygen concentrations are the opposite from your muscles to your lungs. Right, yeah. Yeah, and let's look at the O2 binding curve in the show notes. So somewhere between like 20 and 50 TOR or millimeters of mercury. The curve drops off, or if you want to go the other direction, it goes up really fast and kind of levels off a little bit after these little curves. So at about 60 tor, hemoglobin is about 85% saturated. At 20 tor, it's about 20% saturated. Now at... 50% Now, we usually look at 50% saturation for enzymes because it's a nice middle point, kind of gives us a good idea once we know the shape of the curve and, you know, a couple other things. So 50% saturation happens around 30 torr. And so this is a good middle point because above this, you know, we're going to see 100 and something in the lungs and we're going to see like you know in cells that are exercising at max capacity at like they're like one to three or four tor or something like that yeah um so we can so we now we have a protein that's going to do its job perfectly at either end of the spectrum um as opposed to myoglobin which stays uh with really high saturation until very low pressure so at 60 tor Myoglobin is 98% saturated. At 20 tor, myoglobin is about 90% saturated. And the 50% saturation for myoglobin is about 3 tor. Oh, so yeah. So it is better designed to work in places that don't see very much oxygen compared to hemoglobin, which is designed to... Be exposed to environments with lots more oxygen, i.e. your lungs. Yeah, lots more oxygen and a little less oxygen. Because remember that in terms of diffusion, hemoglobin's concentration and distance delta is, you know, really only on the cellular surface. Even though cellular surface is, of course, driven by mitochondrial disappearance and the creation of zero-pressure zone. Listen to the last episode for that. So hemoglobin may never effectively quote unquote see a zero pressure zone the way that myoglobin will which can explain why they've each evolved for the task that they've evolved for and they each do their job extremely well but we're not done with why hemoglobin is so cool not by a long shot. So the shift of hemoglobin shape due to O2 binding actually changes its responsiveness to acidity. Acidic environments shift the entire O2 binding curve to the right. Literally, this is known as a right shift, meaning it's more likely to release O2 at higher O2 partial pressures. So this means it's going to give away oxygen more quickly at higher O2 pressure environments. So these environments are very much like you would probably find around rapidly respiring muscles churning through ATP and spitting out protons and acid. And so what this shape change that happens between the T and the R conformations when O2 binds to hemoglobin, it means that protons are likely to bind in certain spots on hemoglobin to help with the shape change even more. Alright, so we don't have to go far to see this phenomenon. So if our normal pH is about 7.4, if we make it a little more acidic to 7.2, it changes the saturation point, the 50% saturation point of the hemoglobin binding curve to the right from 28 tor to almost 40 tor, depending on how much acidity there is. Pretty big change. This pH from 7.4 to 7.6, the shift will actually go to the left, so it's harder for O2 to be released, but it's easier for it to bind. So that's... So think about exercising muscle versus the lungs. Oh, yeah, where you have no acid, basically. Yeah, so it moves the 50% saturation point down to almost 20 tor. So we actually have a pretty big ability to shift how hemoglobin binds. Now this is pretty great, but it gets crazier. In red blood cells, there's an enzyme called carbonic anhydrase. It is one of the perfect enzymes in the world. Because there's not really any training benefit from knowing about this, but biologically it's amazing. So carbonic anhydrase takes... The CO2 exhaled from muscle and it combines it with water to form bicarbonate because if CO2 were left as a gas it would actually make bubbles in your blood. Most CO2 actually is transported in our blood as bicarbonate. That's cool. Also, bicarbonate is a convenient buffer. It is a buffer. However, making Bicarbonate from water and CO2 also does something else like it releases a proton. It increases the acidity inside cells which further helps hemoglobin unload O2 to respiring tissues. Cool. In the lungs on the other hand, O2 is so abundant that it sticks itself to hemoglobin whether it likes it or not and it disrupts this. It snaps this whole shift backwards so it changes the state of the hemoglobin and it unbinds these hydrogen ions that had bound to the hemoglobin and carbonic anhydrase is going to extremely quickly going to take that proton and the bicarbonate and make CO2 again and water. Almost Instantaneously. Carbonic anhydrase is one of the fastest enzymes known on the planet. It's what's called catalytically perfect. In other words, it can do its reaction faster than anything can diffuse into it. Oh, okay. So if we imagine in the brownie and soup of stuff bumping around in cells, Carbonic Anhydrase finds a bicarbonate and it can rip it apart and grab a proton and make water and CO2 faster than it'll take for another bicarbonate to bump into it. There's no backlog of bicarbonate or something like building up. It's always ready to... Right, well, actually, there is a point at which, well, we're going to talk about this, obviously, another time, because this is really getting into the weeds, but actually, over 1.0, this is why we cannot trust RER values to tell us how much carbohydrate and how much fat that we're burning because of this kind of thing. But regardless, so anyway, it's so fast because when blood goes through our lungs, it doesn't spend a lot of time there. Carbonic anhydrase needs to be really fast in the tissues to make sure that hemoglobin unloads more oxygen and in the lungs is able to unload CO2 as quickly as possible. So that's why it's one of the interesting reactions and most efficient things in all of biology. I think note here... Some people may be under the misconception that the hemoglobin is what is transporting the CO2 back out. Actually, I'm glad you brought this up because this is yet another misconception that's out there. So it's actually heme. Well, you're right in that heme itself does not bind CO2, but hemoglobin, the protein, does bind CO2. One hemoglobin is actually capable of binding four CO2. What happens is in the four subunits of hemoglobin, The two alphas and the two betas, there's what's called an N-terminal domain. And this is just one end of the long protein chain that gets folded up. And this end, the nitrogen actually binds CO2, which turns it into what's called carbaminohemoglobin. And this is just another way of further shifting the curve to the right for more O2 unbinding ability. So it decreases the affinity, so it right-shifts everything. And about 10% of our total CO2 gets transported this way. But for the most part, yes, it is moved in as bicarbonate. And of course, this process is very easily reversible. So when the carbaminohemoglobin gets to the lungs, this process quickly reverses. And then CO2 just heads off into the atmosphere. We're not actually done yet. We're almost done, but we're not actually done yet. Other things that can cause this shift as well. So we've seen CO2 and acidity, but a product of glycolysis called 2,3-bisphosphoglycerate and shifting temperature can also shift the curve. So if we see more 2,3-bisphosphoglycerate or diphosphoglycerate, we are actually going to shift the curve to the right. And if we see an increase in temperature, You know, exercising muscles create a lot of heat. We're going to shift the curve to the right so it's easier to unload O2. Now, we'll circle back for a second because can you possibly imagine a protein like this, like hemoglobin, getting to the lungs and releasing oxygen? Right, yeah, that doesn't make any sense. There's like, firstly, it's going to be very difficult just based off of diffusion and and Gradients. Like, if there's already a bunch of oxygen there, it's not going to want to be like, oh yeah, let me do work and push more oxygen. Like, that's actually like, that would require energy in all likelihood. Yes. And possibly most importantly, like we mentioned before, hemoglobin and amyglobin give oxygen a hiding spot from the rest of the cells in your body. Because without it, you know, free radicals are going to be a big problem. So if hemoglobin is going to unload oxygen on its way back to the lungs at some point, because if it's going to unload it in the lungs, it'll unload it before the lungs, this is going to create problems also. Oh, yeah. Yeah, yeah, yeah, because you just keep your brain only spitting out oxygen. Like in a non, maybe not predictable way. Yeah. Or kind of like stochastic, like random way. Yeah. And only like one to two percent of all the oxygen dissolved in your blood is actually just like in plasma. Most of it's like in your hemoglobin. So I hope I've made my case for why hemoglobin is the most interesting protein in the world. Nerd. All right. All right. So now we've laid bare the mechanism of O2 transport. One hemoglobin, 4O2. That's, that's, it's, it's very simple equation. One hemoglobin transports 4O2. Hemoglobin is in red blood cells. So, if we have more hemoglobin, we can transport more oxygen. So, is it really that easy? Well, it depends. Ask the Festina team. Ask the Discovery team. Yeah. Let's have David Miller on. Yeah. So we mentioned last episode that the heart is probably the biggest long-term limiter of VO2 max, but the blood is part of that. Thinking about the big components of blood with respect to VO2 max, we're going to take that first before we talk about its relationship to the heart. So well-trained endurance athletes usually have more red blood cell volume than untrained people, like a lot more. The number I see in textbooks and papers quoted everywhere is 40%. I mean, I have no reason to doubt this. I didn't find a reference for it literally anywhere. And I'm sure that there are some high-level endurance athletes with more. I'm sure there are some high-level endurance athletes with less. I'm sure it is a very nicely shaped Gaussian distribution. But why would an athlete need so much blood? Like we just talked about, its primary function... is to transport, well, not its primary function. One of its main functions is to transport oxygen and CO2. But it also transports other molecules to meet metabolic needs. So we transport fatty acids, we transport vitamins, we transport glucose, we remove waste products, you know, trash removal, that's a metabolic need too. Well, the only thing I was thinking is that also well-trained athletes have higher Capillary Density, so like more places for the blood to go. Whereas like if you're like sedentary and maybe a little bit fat, like your fat cells don't require the high density of capillaries that muscles do. Well, you're actually a little ahead of us. That's a really good thought. And probably some of our listeners from last episode are thinking that too. Let's think about hemoglobin mass for a little bit. So total hemoglobin mass and red blood cell volume, absolute volume, plasma absolute volume, and absolute blood volume, total blood volume correlate all with VO2 max, R is about 0.75. So it's not a perfect predictor, but it's a very good predictor. Pretty good, yeah. So we can actually see how important it is in any number of experiments. where removing red cells removes VO2 max improvements from a training intervention. It may have been made over a couple weeks, like four weeks, six weeks, eight weeks, 12 weeks, something like that. Who volunteered for that study? Come on. Right? Yeah. Yeah. I mean, these are phlebotomy studies. You manipulate blood values and you see what happens. And it's a pretty classic thing. And one of the things that we usually think about with red cell volume is hematocrit. That's looking at a concentration. That's looking at what percentage of our total blood is our red cell volume. You know, let's think about having more red cells for a second. Because on paper, having more hemoglobin per unit of blood is good, right? So having higher hematocrit should increase O2 carrying capacity. But it turns out, you know, there's not a good correlation between concentration measurements of red cells and hemoglobin. So if we graph concentration in like grams of hemoglobin per liter or something like that with VO2 max, R is like 0.1 or something like that. We've got a reference in the show notes. It's got a bunch of graphs on it that we're pulling from here. But they don't even actually report the R value. It's so low. It literally looks like a random cloud. Yeah. 0.1 is not. Yeah. No correlation at all. Yeah. Well, they didn't even report it. It's not on the graph because it just, you look at it and you go, oh, there's no way. Yeah. So even if you like took a linear regression of it and you could try to find something from it, I don't know why you would try. You shouldn't. Despite some of the studies that I've seen recently. Oh my God. It would be no better than a random number tenor here. Yes. But concentration, is an important factor because if we look at red blood cells as a percentage of total volume, of course we're looking at hematocrit, but there is such thing as too little or too much hematocrit. So over about 50% hematocrit oxygen delivery ability, I know, sorry to use delivery, but we know it's a bad analogy, but I'm going to disclaim that every time I use this term. The capability of oxygen delivery drops drastically over 50% because the blood actually becomes too viscous to move easily. Oh, that makes sense. Yeah. Yeah. But if we have someone, well, but of course, if we have too little, then we can transport a lot of plasma and there's not going to be much blood in it, not many red cells in it. That's going to be a problem as well. because our blood vessels and our capillaries can only fit so much blood at a time. And as we mentioned last episode, capillaries are really small. Like a lot of blood cells, red blood cells, erythrocytes actually need to fold to go through capillaries. So it's pretty tight in there. They go one by one. It's single file. Now, if we have someone who's sedentary start an endurance exercise training program, Plasma volume increases much more rapidly than red cell volume, especially early on, and this is what's called exercise-induced anemia. Interesting. Yeah, so I'm sure a lot of athletes out there have gone to get their, gone to get a blood test. I remember a friend of mine once, the first time I heard of this, he goes, he was like, he's literally the fittest dude I knew at the time. And he comes back and he's like, yeah, my hematocrit's 38. And I'm like, wow. Yeah, I was like, what? and it's I think it was like early summer or something like that um and so he's he was actually building plasma volume he might not have been that fit or at the time or like something like that um but yeah he like he had exercise induced anemia but it's actually temporary because after a few weeks red cell volume is going to come up and plasma volume is going to come back down a little bit so they're gonna like kind of meet in the middle like plasma volume like overshoots and then like is like, okay, I gotta chill out for a minute. And red cell volume's like trying to catch up slowly because we build red cells pretty slowly. Yeah. And red cells do get constantly renewed by our body, but they have a long-ish but not infinite lifetime. So, why can we still deliver, again, disclaimer on that analogy, I'm sorry, the same amount of oxygen to tissue When the concentration of the red cells drop, the red cell volume remains the same. It sounds like you're actually like quote unquote delivering more blood. Oh, because I forgot to mention earlier that when this plasma volume really increases a lot and red cell volume is coming up slowly, it doesn't affect the VO2 max. So why does this happen? My bet is that it's an evolutionary adaptation. to be able to account for not only different hydration states because you don't see your V2 max drop as you lose a couple pounds of body water through a workout because you're sweating and you're breathing hard and you lose a lot of water through your breath and it's to account for blood volume expansion with endurance training and heat adaptation. Yeah, and that would be interesting too because figure prehistoric people wouldn't necessarily have consistent sources of good Water, that they would be, they're not going to the faucet every hour or whatever and drinking a glass of water. They're not able to be as consistently hydrated as a modern humanist. Right. And you can imagine how terrible it would be if you were like, you know, you can't find any water, and you've got to like, go run. Hunt and gather. Yeah, you've got to gather, you've got to do all, you've got to work, you've got to do stuff. And if your V2 Max is coming down, because you've lost a couple pounds of blood volume and body water, like this is going to be horrible. You have a bad, it's like a, it's a negative feedback loop because like, oh, you wouldn't be able to go get those resources that you need and you'd be digging even deeper hole. Yeah, so how does this happen? Like, I personally have a couple theories on all of this. I haven't seen any myself. So this is just me spitballing. So I think blood regulation, might be a big part of this because exercising muscles send more signals back into the incoming blood vessels to open more capillary beds based on need. But this is obviously not infinite since without some regulation, you could probably easily pull enough blood away to suffocate some of your organs. That would not be good if you exercised too hard and just black out. I mean, some of us do, but that's not the point. And there might be a couple reasons that we pass out anyway. And we talked about this last episode too. Because when we saw the two legs versus the one leg experiment, we saw that the blood volume that was going to a single exercising quad was like three liters maximally, but at VO2 max, with both legs exercising 100%, it only got up to like four liters a minute. Obviously, having more blood than that would probably be detrimental elsewhere. So your body has to regulate this extremely carefully. Now, my other theories on why this happens. So plasma lubes the blood, right? So it's less viscous. That may help it move faster through capillary beds. So that way, you know, you're getting, you know, the increased rate is making up for the fact that it's a little more dilute. So you would get the same number, you would still get the same number of red cells passing through a surface per unit time, yeah. For time, yeah. My last thought is that red cells go through capillaries. Capillaries are really small. And so with red cells going through one at a time, blood pressure might actually be such that the plasma gets squeezed out of the capillaries. I don't know. This is probably the weakest of my ideas. I don't know. I don't study fluid dynamics. All right. So anyway, so I don't think plasma actually gets the credit it usually deserves in terms of exercise ability and exercise improvements. Because in sedentary individuals, plasma volume expansion can already be seen a couple of hours after finishing their first bout of endurance training. Interesting. Yeah. That's really fast. Like, I know that plasma volume goes up from endurance training. I've never looked into seeing how quickly it is, though. Yeah. And it's not, like, huge, but it's noticeable. It's measurable and it's significant. Increases first is a big step in terms of increasing fitness. But when you detrain, plasma volume can be one of the first things to go. And of course, there's plenty of evidence on both sides of this, actually. So in the published literature, there's a lot of evidence that blood volumes do drop, plasma volume especially, with detraining, and there's a lot of literature that says it does not. So it's probably a lot of individual variation here. But now we're going to get to a study. And we're a long way into this episode, but don't worry, this is not going to take that long to get through. And we're only going to cover half of it because we're going to cover the other half in the next episode since this study so nicely straddles both episodes. And this is one of my favorite studies. referenced this a lot. So if you've like done a consult call with me, if you're one of my athletes, I have almost definitely referenced this paper to you. This is once again by Ed Coyle. I told you a long time ago we'd see a lot of him. Mary Hemmert and Andy Coggin are the co-authors. It's called Effects of Detraining on Cardiovascular Responses to Exercise Role of Blood Volume. So what they did was they took eight endurance trained cyclists, average six years training history. I think it was plus or minus one. The average training that they were doing seemed to be about four threshold workouts a week, 70 to 80% VO2 max and one VO2 max workout, five by five minutes at 90 to 100% VO2 max. Testing was done over two days while training and then again after two to four weeks of total cessation of activity. So just done. Stopped on the couch two to four weeks. So this is a detraining study. And the authors based this study on observations that plasma volume quote-unquote detrains quickly after stopping activity. So the experimental section of this study, so they tested the athletes twice while training and twice while detraining. The first day they did it normally, and then they tested the next day with their experimental procedure. On day two, There was a plasma volume expansion done, both in the trained and untrained states. And what they did was they aimed to increase blood volume by about 8%, or 700 to 800 milliliters. And this would bring, after detraining, plasma volume back up to normal, but it was also done in the trained subjects as well to see if extra plasma volume might have additional effect over the already expanded with training plasma volume. Does that kind of make sense? Yeah, that's interesting. So they did it in the trained subjects as a control, basically. Yeah, and so that was their training schedule on average for a bunch of weeks before they got to the study. They tested them during their training period, then they tested them after two to four weeks off again. So, what happened? Detraining resulted in an average 9% decrease in blood volume, average 5.2%. to 4.7 liters. And of the loss, the average was 350 milliliters was plasma volume and 130 milliliters was red cell volume. Interesting. Yeah. So it is mostly plasma. Yes. So 2.27 liters of red cells detrained to 2.14 liters, a nearly 6% loss. This meant detrained the VO2max went down on average 4.42 to 4.16 liters a minute or about the same 6% loss. Which makes sense. We said before the kind of R of 0.75 correlation. Great. Yeah. Cool. Yes. And then when they did the plasma reinfusion in the detrain subjects, this brought their VO2 max back up to 4.28 liters a minute on average. Not quite back to normal. And halfway. Yeah. And I remind you, there was no red cell reinfusion. That stayed 6% low. Only plasma volume was replaced. Interesting. Yeah. So now we mentioned hematocrit and, you know, lubing the blood with plasma. So their normal hematocrit when trained was 43.8% on average, of course. But when it detrained, It went up to 45.7%. Re-infusion in the detrained subjects of plasma volume to bring their normal blood values up dropped hematocrit to 39.6%. Which, remember, when we mentioned earlier that concentration is not that correlated with VO2max, this shows us exactly that. And, also, VO2max went up when the concentration went down. Okay, last thing, of course, I'm sure some of you are like, what happened with the train people? So, the train people, did the plasma expansion help bring up VO2max? No. VO2max stayed the same. It actually was within measurement error. So, it actually dropped 0.05 liters a minute, 4.42 to 4.37 liters a minute. So, it's within the margin of measurement error. It's basically the same. My theory is that had they replaced the red cells in the detrained subjects, they would have seen a total reconstitution of VO2max. But they didn't do that. I'm sure those people were competing and they didn't want to get them doping even though it was the late 80s or early 90s or whenever the study was out. 86. Thank you. So what we're leaving out in this study, we're actually going to deal with in the next episode. on the Fick equation when we talk about heart and heart stroke volume. But we're not done yet. So I hope this so far illustrates all of the constituent components of VO2 max, especially those that can come and go in the short term, especially with training studies, four to five weeks or less, relative contribution of each, you know, in terms of plasma volume and hematocrit and heart stroke volume and stuff like that. People start training and their VA2 max starts to go up. Did their heart stroke volume actually increase? You know, it turns out that there's an interplay between this and blood volume. And if you take away the blood volume, you're going to go back to normal is what most studies will show. And the blood volume expansion, though, it's not exactly temporary either in the long term, but it's not also permanent. As we've seen, plasma volume responds to training quickly, and red cell volume responds slowly. So the other component to this is with respect to blood pressure and what might be actually too much blood volume. Because if we have too much volume and we don't have enough places to put it, we become hypertensive. Six liters of blood in a sedentary person is a recipe for a heart attack. They're not fit. Cut back on the salt. Yeah, and so you're lacking, so a sedentary person would be lacking what's called vascular capacitance, somewhere to put the blood, which is another benefit of endurance training when we build capillaries and blood vessels and they presumably become a little more supple. There are things called elastance or compliance in arteries. And having the ability for the artery to expand and contract a little bit is a good thing. If otherwise we have atherosclerosis and that's a bad thing. Too much plaque built up on the inside of your... Yeah, too much plaque so you're stiffened arteries. And so arterial stiffness... is related to hypertension, of course, but it's also related to aging. So it's just something that happens as we get older. And does this have an effect on our aerobic ability? Probably. How much? I am not sure. I'm not a doctor. I haven't studied this specifically. But the kidneys are incredibly important. They regulate plasma volume and red cell volume. and we're going to talk about them specifically in a future episode because a study actually came out about eight people have sent it to me and I've already read it and I've read it like six of those eight times again now on the crit meter potentially in the kidneys so it's still theoretical at the moment there's not ironclad experimental proof and we'll get to that in the future but before you send that to me again I'm aware of that study thank you So when we consider things like blood volume and vascular capacitance, blood flow regulation of muscles, mitochondrial density, anything that has to do with long-term improvements, just remember that adaptations are not symmetrical. Some are faster, some are slower. Plasma volume, like we said several times, is a fast responder. Muscle capillaries, hematocrit, these are slow responders. But something else to note, Increasing plasma volume like this can make concentration measurements look like they're getting worse or better, like hematocrit. You know, if you exercise-induced anemia right there, it looks like something bad is happening because your hematocrit is going down, but you actually haven't lost any red cell volume. All right, so if plasma volume expands, this can also have another effect on things like lactate. So if we make the same amount of lactate and we have more blood volume to put it in, the concentration goes lower. And how do we always report lactate values in the blood? Millimoles per liter. Which, you know, is just millimolar, but, you know, it's easier to see it in millimoles per liter because, of course, in the medical field, everybody always wants to see that, whatever it is, per liter or per deciliter, or however they measure it these days. So if this study had only looked at hematocrit, we might actually have a different interpretation. You know? Because there's a paradoxical nature of how hematocrit goes up with detraining and then drops with early training plasma expansion. And so, you know, your hematocrit drops, your hemoglobin's actually fine, your red cell volume's fine, VH max is going up. What is going on? Yeah. So we're working towards the bottom line here. which is blood volume in general. More blood volume has a lot of benefits. Generally speaking, if we have more plasma volume, then we can have more things like albumin, which is the most abundant protein in plasma and is involved in transporting stuff like hormones and fatty acids, something you might want to use more of at some times rather than others. And it's found in eggs. It's found in eggs. And more plasma might mean more red blood cells for more oxygen transport. Theoretical, but looking promising, possibly. But especially with the heart, more blood volume means we're actually going to get more heart stroke volume. Based on one of the two main contraction strategies, quote-unquote contraction strategies, of the heart. And you've probably heard of the Frank Starling Law, which is that when the heart gets filled more, it's going to contract harder to get that blood out, because otherwise you've got extra blood sitting in there, and then you can't fill as much, and if you can't pump it out, then your heart's just going to explode eventually. Sounds like a bad day. It sounds like a very bad day. Not recommended for athletes especially, or anybody who's Alive. Alive. Yeah, likes living. And so that's where we're going to start for the next episode because heart stroke volume is really, as we've mentioned before, the biggest long-term limiter of VO2 max, but of course it's hard to pump more blood if you don't have enough blood, so these things are interlinked. So we're going to get into the heart stuff next episode. and we're going to see a lot more about this interlinking between blood volume and heart stroke volume and we're going to look at some of the very few papers out there that are brave enough to actually try to tackle the mechanism of increased heart stroke volume. Do you have any final thoughts Kyle? Yeah I think this is kind of cool because I think a lot of people especially in cycling have heard of hematocrit and think you know generally if you You hear hematocrit in the doping context, but you hear like, oh, higher hematocrit must be better. And this must be the case, especially because like the UCI had that rule that you weren't allowed to race if your hematocrit got tested over 50% or whatever. So people would, you know, ride that line really close. And that kind of implies that just always more hematocrit is better. It does imply that. And actually, we remember hearing stories about people having to get up in the middle of the night. and Spin or their blood was going to coagulate and congeal because they had too many red cells. Right, right. So it's a health and safety thing, you know, despite the doping thing. Right, exactly. And this kind of goes to show that it's not a universal truth that higher hematocrit means that you're fitter or faster or whatever. And then again, looking at this concentration is like an oversimplification of the things that go into making you actually fit. Right. And that paper that came out recently that's been sent to me eight times, one of the things that happened is they actually managed to increase the athlete's red cell volume. And actually, we didn't mention the blood volume thing. How do we measure blood volume? Let's get that out of the way real quick. Because one of the reasons, one of the things you might think is like, I want to measure my blood volume. It's hard. Because what we have to do is we have to take a known quantity of something. In the COIL study, they used a blue dye. And it's not weird. It's not like you're actually like tie-dying your blood or anything like that. The blue dyes are actually very common in the sciences. There's a lot of different molecules that can Reflect Blue Light. So what you do is you put a known quantity into the blood and you let it dilute. It takes a while and then you draw out a known quantity and you measure how much of your tracer is in it and then you can just reverse calculate how much blood there is. That makes sense. Yeah. So that's why we don't see this kind of thing more often because it's time consuming and, you know, you've got to like... I don't know. It's a measurement that a lot of people don't actually take that into consideration that much. An autopsy is also another very accurate way to measure blood volume, but typically people aren't itching to volunteer for that either. Yeah, just drain it out. So what were we talking about right before that? Oh, just that, you know, more hematocrit is not necessarily better. Oh, yeah. Okay, so I'm sorry. So in that study, they actually managed to increase the athlete's hematocrit. with a certain method of training. And most of us coaches who have looked into this kind of thing, we've known that this is probably maybe a thing for a while. However, what happened is, despite the increase in hematocrit, they did not increase their VA2 max. Oh, interesting. Yeah. Yeah, that's maybe a surprise to... Everyone involved. Yeah, but see, the thing is, like, this is, we're gonna get a little into the weeds now, but I remember reading Secret Race, and that was Tyler Hamilton, right? Yeah. Yeah, so in Secret Race, Tyler Hamilton says that, you know, you cannot just dope and make more red cells with artificial erythropoietin, and then be faster suddenly. He says, you still have to do the training. So the question is, in the longer term, in this study where they added more red cells, very legally, they were not doping, if they trained with this over the next couple weeks after this red cell volume increase, would they have gotten fitter than the group that didn't actually add this extra hemoglobin? I think it's interesting. I think it's a good, it's still a good, I mean, blood doping is like the elephant in the room with all of cycling and a lot of other sports too, but like it, it, it's hard not to talk about it if you're going to talk about human body performance and physiology because it's something that is a, it's a common touchstone point for almost everyone in the sport who's been in, heard of cycling, even people who have not participated. You know, increase sort of understanding if people have better familiarity with words like hematocrit. No, that's true. That's very true. Yeah, and I think the other thing is that looking at doping in some lights is instructive. You know, like you just said. But, because normally when we want to look at how things function, we look at The absence of them. We look at accident victims. We look at people who have unfortunate genetic mutations when they're born. We look at diseases and how they affect certain molecules, certain systems in the body, certain proteins. We look at mice. We look at pigs. We look at all kinds of model animals that would be typically lacking a gene. Lacking this function. And so it's interesting to see what happens when we add function. If we improve something, what happens? Usually we're looking at people who are, and animals that are not having a good life. Let's put it that way. Alright, so as always, I want to thank everybody for listening. You know, I want to, of course, thank everybody who's written to me. Sorry I couldn't get back to everybody. written to me saying, you know, thank you for the content, thank you for the episodes, and thank you for helping me get a lot faster and fitter. That's why we're doing this, and I love hearing that, so even if I don't respond, I'm really sorry about that. Please, please keep writing. If you have any questions or comments, you've got any coaching and consultation inquiries, you can just shoot me an email, empiricalcycling at gmail.com. If you have any merch needs, if you want to get an Empirical Cycling Podcast bottle or a shower curtain, I don't know, I gotta get a shower curtain. That's at empiricalcyclingpodcast.threadless.com And an iTunes rating would not go amiss as well if you are enjoying the content and especially sharing the podcast with your friends and loved ones. And I hope everybody's staying safe out there. Thanks, everyone.