Shocked

by David Casarett

  In 2010, Oivind Toien and his colleagues rescued a group of five “nuisance bears.” (These are wild bears that have proven themselves to be so incorrigible and so prone to menacing human populations that they are euthanized.) Instead, the scientists recruited them into a research study. Spared from execution, the bears happily went to sleep in man-made dens, watched by curious researchers.

  What those researchers found at first didn’t bode well for bears’ prospects in the hibernation club. Although the bears’ temperature did drop, it dropped by only 4 or 5 degrees Celsius. Alas, the official scientific report makes no mention of steam issuing from their nostrils.

  However, the researchers also found that the bears’ metabolism dropped by about 75 percent. That is, their temperatures didn’t decrease nearly as much as expected, but their metabolic rate did. Meaning that if you judge by the (old) temperature criteria, bears don’t hibernate; if you look at metabolism, they do.

  So bears are back in the club, and another important lesson is learned. At least in some species, metabolism can drop way out of proportion to temperature. That may seem like a trivial discovery, but it’s not.

  Everyone thought that you’re not hibernating unless you’re cold and hibernating. But the discovery that bears can reduce their metabolic rate without a large reduction in temperature means that there might be ways to reduce metabolism that don’t involve temperature regulation. At least, not directly.

  Remember the French Cocktail and the ensuing debate between Laborit and Dundee? Remember all of those bags of ice placed on all those groins? What if all of that drama isn’t necessary at all? What if we could slow metabolism directly? That’s an exciting possibility.

  There’s another reason to be excited that the search for the secret of hibernation is shifting to metabolism. If we’re looking for something that alters metabolism specifically, it may be easier to find a trigger that works in both animals and people. Despite all of the differences between mice and bears and lemurs and people, at the level of our cells, we’re more similar than different. So if we hit upon something that could reduce metabolism at the cellular level, this could be a game-changer.

  Think back to the fly-fishing example, and the guy across the stream who is catching all those fish. I said earlier that if you don’t have an exact match for whatever flies he’s using, then you’re out of luck. But that’s not quite true.

  What if you don’t have to find an exact match? What if you happen to have a fly that is universally attractive to all trout, everywhere? Then you don’t need a #10 caddis or a #12 coachman or whatever that guy who thinks he’s God’s gift to trout is using. You can use that all-purpose fly.

  Back when I was a resident, whenever I had a day off I’d sneak away and go fly-fishing in some of the small streams in northern Iowa and southern Minnesota. Those fish were a tough audience. They were very picky. And because I didn’t know the local streams, most of the time I had no idea what those trout liked.

  But I had a trick that would often salvage a fishless day. Most of those streams ran through open meadows, and I knew that those meadows were full of grasshoppers. I also knew that trout like grasshoppers. A lot. Grasshoppers are to trout on a hot summer day what buffalo wings are to a football fan on a Monday night. So even without knowing any of the local patterns or hatches, I’d usually do pretty well with a #12 hopper.

  That’s what researchers like Cheng are looking for. They aren’t hoping for an exact match of a particular animal’s physiology with ours. They don’t want a cocktail that will mimic all of the complex changes that a hibernating body undergoes. They just want the equivalent of that #12 hopper. They want something that’s close enough to trick a body into thinking that it wants to hibernate.

  SAM, SMÉAGOL, AND FRODO

  As Cheng and I talk about bears and mice, it’s starting to sound like AMP might be the biological equivalent of that all-purpose #12 hopper. I’m starting to be convinced. But I still haven’t heard a good explanation of how it works. I’ve seen it induce a state of hypometabolism in a mouse. Or at least that’s what it looked like to me. But how?

  By way of an answer, Cheng shows me two graphs that are similar to the one we started with. Both have the same shape of the VO2 curve that drops precipitously and then flattens out. And both show a temperature curve that drops more slowly.

  I tell him I still don’t get it.

  Cheng grins and points at the legends under each. The graph on the right looks almost identical to the one on the left, except that it describes the result of doses of AMP that were ten times greater. The dose of AMP went up tenfold, but the effects were essentially the same.

  I think about all of the drugs that doctors prescribe every day, and what Cheng is showing me just doesn’t make sense. Take lisinopril, a drug that’s used to control blood pressure. Ten milligrams is a typical starting dose. That works for many people. They don’t make a 1 mg pill, but if they did, and if you gave it to the average person, it wouldn’t have any effect. They don’t make a 100 mg pill either, but if they did, and you were dumb enough to give that to another average patient, that patient’s blood pressure would probably drop precipitously, causing dizziness, lightheadedness, and maybe a loss of consciousness. That’s a typical dose-response relationship. More drug leads to a greater effect, up to the point of toxicity and even death.

  Cheng understands the oddness of his results too. “The first time I saw this, I thought there was a mistake. It couldn’t behave like this. It’s just wrong.” He sounds like he was seriously offended that his AMP isn’t behaving in the way that it should.

  It wasn’t wrong, though. In fact, the lack of a dose-response curve offers a couple of clues as to how AMP works. It’s what is called an allosteric reaction.

  An allosteric reaction, Cheng explains, has three “actors.” There’s a protein. Then there’s whatever molecule the protein usually binds to. Then there’s a third actor—call it X—that binds to the protein. That third actor, X, changes the affinity of the protein for the molecule to which it normally binds. Depending on whether X increases or decreases that affinity, it’s described as either a positive or negative allosteric reaction.

  Think about two people who don’t really get along. Call them . . . Sam and Sméagol. Left to their own devices, they’d never spend any time together. But then add a third person. The X factor. Let’s call him Frodo. With the addition of Frodo to the mix, Sam and Sméagol manage to tolerate each other, at least for the duration of their journey to Mount Doom. They’re never friendly, and they never really get along. But they coexist. That’s the way a positive allosteric reaction works.

  Allosteric reactions tend to produce results at predetermined levels. A protein by itself will have a certain attraction for another molecule. If you add X, the protein’s attraction for that molecule is kicked up a notch, or down a notch, but only within a predetermined range. Just as there’s no way Frodo will ever convince Sam and Sméagol to become BFFs, an allosteric reaction can’t convince a protein to bind more tightly to a molecule than it’s programmed to.

  So—and here’s the most important point about allosteric reactions—you get the same change in attraction and binding whether you add X at a 1 percent concentration or a 10 percent concentration. Sound familiar? That was exactly the result that Cheng was seeing.

  Even better, once Cheng figured out AMP was part of an allosteric trio, he knew there aren’t that many allosteric reactions in the body. So he could make an educated guess about how AMP works. That’s because he knew about an interesting and very well-known negative allosteric reaction that happens in red blood cells. A molecule called 2,3-diphosphoglycerate (known either as 2,3-DPG or BPG, for biphosphoglycerate) reduces the attraction of hemoglobin for oxygen. In the presence of BPG, hemoglobin is slower to bind to oxygen, and quicker to release it to tissues that need it. So at a given pressure of oxygen in the blood, adding BPG means
that less of that oxygen will get bound to hemoglobin.

  The key point here is that the interaction between BPG and hemoglobin is a negative allosteric reaction. If you expose red blood cells to a little BPG, you reduce their capacity to carry oxygen. Expose them to a lot, and you get the same effect.

  The most important implication of this result is that it pinpoints the location of AMP’s effects. “Now,” Cheng announces, “we think we know where this happens.”

  We do?

  “Where is BPG?” He looks at me steadily.

  I’m stumped.

  “Where is it . . . in the body?” he clarifies, helpfully. Thanks for that.

  Well, red blood cells, obviously, I tell him.

  “And?”

  I stretch my memory back to the first year of medical school, without much success, but then it hits me.

  Nowhere else?

  “Right!” Cheng says. (Actually, it also appears in the placenta, he admits. But I’m pretty sure my male mouse didn’t have one of those.)

  This means that ordinary red blood cells might have a role in hibernation. As we’ve seen, people have looked to the brain and the liver and even the eyes, if you count Aristotle’s strange fascination with swallows. However, no one was really thinking about red blood cells.

  But how does it work?

  The answer to that question isn’t complicated, but getting to it requires a basic understanding of how our bodies break down glucose (sugar) to produce energy. This process, known as glycolysis, is the way a cell uses the energy that is stored in glucose to create adenosine triphosphate, or ATP—the energy molecule we met in chapter 3. Glycolysis has two interesting features that are relevant to Cheng’s work.

  First, remember that adenosine comes in three forms. There’s adenosine monophosphate (with one phosphate molecule). And there’s adenosine diphosphate (with two) and adenosine triphosphate (with three). When you add AMP to red blood cells, it becomes ADP by pulling a phosphate molecule off ATP.

  That, in turn, leads to a cellular shortage of ATP, which slows metabolism. Our cells need an initial investment of energy to make energy, and in the same way that a tough economy restricts the availability of startup capital and makes it more difficult to get a loan, a shortage of ATP makes it that much more difficult for cells to produce new ATP. Indeed, Cheng and his colleagues have found that AMP-treated blood cells have more glucose than they should, which suggests that they’re not breaking it down at the usual rate. That is, AMP seems to slow down metabolism.

  It’s the second feature of this process, though, that gets us back to BPG. When there’s a shortage of ATP, then BPG—normally a by-product—is suddenly produced in large amounts. And that happens—if Cheng is right—in every red blood cell in the entire body. That means that a very focused change would induce a whole-body hypometabolic state. By simply shifting the oxygen-hemoglobin binding curve a bit, AMP—if indeed it’s AMP that’s responsible—could force a systemic downregulation of metabolism. That’s one change, to one molecule, that then affects every organ in the body. Amazing.

  It’s a little like using gas prices to change people’s behavior. You can do all sorts of things to encourage people to walk more and drive less. But as a single, global intervention, there isn’t anything that works quite as well as tripling gas prices at the pump. So maybe that’s AMP’s role—to create a tax on oxygen that means that cells everywhere need to conserve.

  But is that safe? It seems a little risky. Surely there’s a potential downside to decreasing oxygen supply. Like brain damage. Or death. But Cheng smiles.

  “It’s perfectly safe, as long as you keep ambient temperature low so that the body temperature can drop. It’s the cooling of the body that allows for the decreased need for oxygen. With a low enough temperature . . . no problem.”

  THE BADASS BEAGLE

  In order to reassure me that what he’s doing is safe, Cheng fires up a video to prove his point. He points at the screen.

  “This is a dog,” he announces.

  Indeed it is. He has floppy ears and wiry fur. He looks like a beagle having a bad hair day, and he’s very cute. I find myself thinking back to experiments with thirteen-lined ground squirrels and “mongrel dogs of medium size,” and I’m fearing the worst.

  First the dog is given a haircut, to reduce his ability to keep warm. That’s not too bad. There are a few tufts of fur around his ears that are sticking up at 45-degree angles, but this just makes him look a little badass. At least, as badass as a beagle can look. Then he’s injected with AMP.

  The video was taken in a lab in China by one of Cheng’s collaborators, and he admits they weren’t sure of the optimal dose of AMP for a dog. So they made a guess based on their work in mice.

  He shrugs. I wince. This, I’m thinking, is not going to end well.

  But the dog seems unperturbed about his immediate future health prospects. He sits there panting and flopping his ears back and forth on Cheng’s laptop screen. As we’re watching, the dog stumbles around a little. Like an undergraduate at a frat party. Then the dog passes out, as gradually and naturally as if he were falling asleep.

  I’m worried. But it seems that Cheng isn’t. And he reminds me why there’s really nothing to worry about.

  Because the relationship between BPG and hemoglobin in the presence of AMP is an allosteric reaction, it imposes its own limits. Remember how a tenfold change in dose produced the same graph? That’s because if you throw a bunch of BPG at a red blood cell and that cell’s carrying capacity for oxygen decreases, it only decreases so much. As a natural molecule, it’s designed (if you want to use that term) to work within a range.

  After some measurements of the dog’s oxygen utilization and heart rate to confirm that it truly is in a hypometabolic state, as Cheng promised, the dog wakes up, just as the mice did. No harm done. In the last scene, we see him frolicking on linoleum without a care in the world. Cheng tells me that he was later adopted.

  One made-in-China video hardly proves that AMP is safe and effective. And we’re a long way from clinical trials in people. But still, I have to wonder what might be possible someday.

  Indeed, it’s difficult to watch that video clip of the badass beagle and not think about how some version of AMP might be used, someday, on a cardiac arrest victim. Paramedics arrive at the scene, apply cooling blankets, and give an injection of AMP. Then, with a dramatically lowered metabolic rate, and concomitantly lowered oxygen requirements, the patient is taken to a hospital where he can be evaluated and resuscitated in a controlled way. We’d treat a cardiac arrest in much the same way that we’d treat a broken leg—as an urgent problem certainly, but one that can be managed carefully and methodically, rather than with a pell-mell ambulance ride through city streets.

  STUART LITTLE’S NEXT ADVENTURE

  “Ah, there he is.”

  I ask Cheng if he’s sure. That’s my mouse? He looks like he’s starting to wake up from a very deep sleep. Cheng rolls him onto his back with a gloved finger, and #0011 rights himself. He paws at the wood shavings in front of him. I swear he yawns.

  Cheng is grinning. “See?”

  He points out that the mouse would not have attempted to right itself an hour ago. Back then, he was a small furry pillow. But now he’s starting to wake up.

  Together we watch as little #0011 wakes up and warms up. His temperature gradually swings back to normal. By the time Cheng and I leave the lab, the little guy is scampering around his cage. Not even a hangover.

  There are limits to these AMP injections. According to Cheng, the mice wake up after five to seven hours. And if you keep dosing them with AMP, they die. So Stuart Little’s next adventure will probably not be to Mars in a suspended animation pod.

  Cheng explains that the challenge is glucose. If AMP slows the breakdown of glucose into energy, then there’s a limit to how long an animal can su
rvive. If you don’t give those cells another form of energy, they’re going to die.

  I think for a moment about what I know about metabolism. What about fat? I ask.

  Cheng nods enthusiastically. In a hypometabolic state that lasts for a few hours, he explains, the main source of energy is glucose. But in hibernation, it’s fat. The trick is going to be to convince cells to switch over. That’s a challenge, but it’s an opportunity, too. If you can induce that switch, that opens up a whole new door.

  We walk down the tiled hallway toward the elevator, and although our visit is now officially over, there’s still a whole list of questions I’d like to ask. But none of them has to do with BPG or about what goes on inside cells. Instead, I find myself thinking about the implications of this line of research.

  In the elevator, I push Cheng to think about what might be possible for people. If you can put a mouse into a state of hypometabolism with a simple injection, what might that mean for accident victims? Or soldiers with battlefield injuries?

  Cheng is reluctant to speculate. When we’d first met, he warned me that his work focused on mechanisms of hypometabolism, not on clinical applications. And throughout our conversation today, he’s been reluctant to take more than a few steps beyond the comfortable zone of mice and into the domain of men.

  But Cheng may not have a choice. This is the world, remember, that turned Mitsutaka Uchikoshi into an international media sensation. And it’s the world that has become convinced that, no matter what its clinical applications, suspended animation is a great tool for interstellar travel.

  The general public is likely to find results like Cheng’s irresistible because the potential here is very hard to ignore. This entire line of research, and Cheng’s in particular, has an elegance and clarity that seems to demand that we think of implications and applications. What started as a harmless—except to squirrels—fascination with hibernation is gradually turning into a clinical science that could save lives.