Sleep: An Antioxidant for the Brain? Podcast with Mimi Shirasu-Hiza

Why do we need to sleep? Part of what makes sleep so fascinating, as a field of research, is that it is such an enigma. Sleep is a profoundly vulnerable state, leaving us at the mercy of predators and the environment, and unable to defend ourselves or our possessions. It’s also largely unproductive (which is why so many people want to figure out how to get by with less). Yet we spend about a third of our life in slumber.

Moreover, sleep seems to be nearly universal in the animal kingdom. Indeed, we have yet to identify an animal that clearly does not sleep at all, or one that can forego sleep without experiencing physiological consequences.

All of this, taken together, unambiguously shows that sleep is extremely important. And this makes it all the more remarkable that the actual purpose of sleep remains elusive.

We know quite a bit about the different stages of sleep, the involved neural pathways, and the physiological effects of sleep deprivation. But no singular essential function has been identified yet.

Of course, several plausible theories have been tossed around over the years. One relatively old hypothesis, which had been discarded, has recently begun to resurface.

Oxidative Stress and Sleep

Decades ago, Eric Reimund proposed the free radical flux theory of sleep, basically suggesting that the primary purpose of sleep is to protect the integrity of neural tissue from oxidative damage. He pointed out that the brain demands a great deal of energy, and these metabolic processes inevitably generate a lot of free radicals. Free radicals, if they are not neutralized, can damage lipids, proteins, and DNA. Not ideal!

We know that the processes of sleep facilitate restoration and repair in the body in various ways. Notably, when we sleep, cerebral metabolic rate drops, and it is thought that endogenous antioxidant systems may be more efficient at that time. Thus, Reimund postulated,

sleep functions essentially as an antioxidant for the brain.

Sounds pretty compelling, right? But the hypothesis lost traction, as several studies subsequently failed to find evidence of oxidative stress after inducing sleep deprivation. However, these studies generally examined acute sleep restriction. What about chronic sleep deprivation, and its relationship to oxidative stress?

And that brings me to our guest for this episode.

Guest

In this episode of humanOS Radio, Dan talks to Mimi Shirasu-Hiza. Mimi is an associate professor of Genetics and Development at Columbia University. Her lab uses circadian mutants of fruit flies (Drosophila melanogaster) to unveil the molecular mechanisms that underlie circadian-regulated physiology.

Fruit flies are a surprisingly useful model for understanding sleep. There are a lot of fundamental similarities at the genomic level with respect to sleep and metabolism, and their genes are readily manipulated for experimental purposes. To that end, sleep researchers have identified a wide array of Drosophila mutants that consistently exhibit short sleep, but due to different genetic defects. For instance, some have loss-of-function mutations in ion channels and ion channel regulators. Another type shows a mutation on a dopamine transporter gene. So the specific genes and mechanisms vary, but the outcome is the same – they all just don’t sleep very much.

Mimi and colleagues hypothesized that this collection of short-sleeping mutants might also share a common physiological defect due to their reduced sleep – independent of the specific mechanisms that drive that sleep reduction. And if they could find such a defect, surely that could shed light on the core function of sleep.

They tested this hypothesis in an elegantly designed series of experiments, which we discuss in the podcast.

Study Findings

This is a fairly intricate study, but I’ll recapitulate the main results here.

• The researchers injected several of these mutants with paraquat. Paraquat is an herbicide that catalyzes the production of superoxide anions. It’s pretty nasty stuff and is now restricted in the US due being such a powerful oxidant, but that unpleasant attribute makes it rather useful for research like this. The researchers observed that the short-sleeping mutants died at a higher rate than controls, suggesting they were more sensitive to oxidative stress. They also administered hydrogen peroxide, and found the same result. So, flies that are genetically predisposed to less sleep all share a susceptibility to oxidative stress.
• Then, the researchers increased sleep in flies, by genetic manipulation and by pharmacological intervention. Both methods produced greater resistance to the oxidative challenges, suggesting that increasing sleep protects against oxidative stress.
• Mimi and her team also measured the expression of genes that are known to be activated by high levels of reactive oxygen species in the heads of short sleeping flies and in controls. They found that the short sleeping flies exhibited higher expression of various antioxidant genes and mitochondrial stress response genes, suggesting that short sleep was increasing reactive oxygen species in the brains.
• Finally, the researchers drove neurons to overexpress antioxidant genes, which forcibly reduced reactive oxygen species in the brains of the flies. Altering the expression of these genes, and reducing oxidative stress in the brain, substantially reduced sleep in flies. This suggests that oxidative stress mirrors need for sleep. The findings, overall, support a bidirectional relationship between sleep and oxidative stress.

Okay, so why does all of this matter?

Because chronic sleep restriction, which is really what Mimi and her team are examining here, is slowly becoming the norm for people in the Western world. Average sleep time has been steadily decreasing (one third of adults sleep less than 7 hours per night), and reduced sleep is associated with a variety of diseases that are linked to oxidative stress.

We know, for instance, that sleep fragmentation and sleep deficiency increases risk of Alzheimer’s and Parkinson’s diseases, and evidence of oxidative damage has been reported in the brains of patients with neurodegenerative diseases.

That’s just a quick highlight of Mimi’s latest work. To learn more, please check out the interview below!

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Contributions

Dan prepared for and conducted the interview, Ginny wrote the blog post, and Mimi continues to do all the hard work in the lab!

Transcript

Shirazu Hiza - 00:07: Oxidative stress, at least on the physiological or cellular level, is really an imbalance between some levels of reactive oxygen species and defenses against them.

Speaker 1 - 00:17: HumanOS. Learn, master, achieve.

Dan Pardi - 00:33: Welcome back everybody to humanOS radio. Today I have with me Mimi Shirazu Hiza. She is an associate professor of genetics and development at Columbia. And the principal investigator in the Shirazu Hiza Lab. Mimi received her PhD in Biology from UCSF and served as postdoctoral fellow at Stanford University in the [David Schneider 00:00:53] lab. Currently, her lab uses circadian mutants of fruit flies or Drosophila[inaudible 00:00:57] molecular mechanisms underlying circadian regulated physiology. With a particular focus on the roles of immunity and in neurological disease. I became aware of Mimi's work when I saw a publication that recently came out in Plos Biology entitled 'A bidirectional relationship between sleep and oxidative stress in Drosophila'. What is the relationship between sleep and oxidative stress? Without further ado Mimi, welcome to the show.

Shirazu Hiza - 01:21: Hi. Thanks for having me. It's really a pleasure.

Dan Pardi - 01:24: I've been thinking about this idea that determining the purpose of sleep is really been a challenge for so long. And we certainly know that many important things happen during the process of sleep, to aid in repair and the functioning of the human body for the next day. And we know there're serious consequences to both getting acute sleep loss and also for not getting enough sleep on a regular basis or a chronic level. And in your work you're exploring this connection between sleep and oxidative stress. And the first thing we wanna do is do a little housekeeping, if you will, and perhaps we can define what we mean when we are talking about oxidative stress for our listeners who don't know that.

Shirazu Hiza - 01:58: Oxidative stress, at least on the physiological or cellular level, is really an imbalance between some levels of reactive oxygen species and defenses against them. That's sort of very simplified, but the cell has various mechanisms to control damage caused by oxidative or reactive oxygen species. And generally, there's a very tight regulation of the levels of the reactive oxygen species that are allowed to run around in the cell. You have a little bit more than usual that will induce oxidative stress responses, that will then dampen the levels of reactive oxygen species. In general that's a pretty good homeostatic system. Oxidative stress, is any time the body is out of whack. You have more stress on the system than it can take at that given moment.

Dan Pardi - 02:48: Recently I interviewed Professor Michael Ristow, and we talked about the story of this oxidative theory of aging. Where the thinking was that if you can eliminate all free radicals, because they do damage, then that would be a good thing. What we've now learned is that reactive oxygen species play an incredibly important role in triggering cellular mechanisms that lead to all sorts of healthy outcomes. Things like improving the oxidative status of the cell, by triggering the production of internal endogenous antioxidants. Two, triggering the differentiation of stem cells and activating anticancer pathways. And you don't want to eliminate those, but you also want to keep them under control.

Shirazu Hiza - 03:22: That's right. Exactly. That makes perfect sense to a lot of people. You want to have a little bit of a good thing, but not too much of a bad thing. There's a reason why the active oxygen species are allowed to exist in the cell evolutionarily. And they do play a very important messenger and signaling roles. It's important to note, when I first started my lab at Columbia, we really focused on circadian regulation of immunity. We were looking at various immune responses and metabolic responses that were circadian regulated, that contributed to pathogenesis with a variety of different bacterial infection. And one of our main tools in studying circadian immunity was to look at circadian mutants. And asked what was their phenotype when stressed by infection with different pathogens. What didn't always sit super well with me was that, these circadian mutants, some of them didn't sleep particularly well.

04:14: And I know for myself, it's very anecdotal, right? But it's very intuitive, when I don't sleep, I get sick. And when I get sick I feel sleepy. And I thought it seemed like there would be a role for sleep in immunity and that's really what we wanted to study, but couldn't. There weren't really good tools to study that for a long time. In the last five or ten years, these sleep mutants have been discovered in Drosophila. That have perfectly good circadian regulation, but don't sleep very much. The timing of their sleep is normal, but they sleep less than wild-type. And we thought that would be a really great opportunity to dissect the two different contributions of circadian regulation and sleep, to immunity against bacterial infection. We thought, perfect. I put a student on this project. She gave a whole bunch of bacterial pathogens and said, find a phenotype.

05:08: And she comes back to me and says, "I can't find a consistent phenotype." What she had done is collected a whole bunch of very diverse short sleeping mutants. There's a mutant that are mutants that are all short sleeping, but they're short sleeping for different reasons. There're mutant for different genes, in different pathways, and different mechanisms. And we thought if immunity against infection is a function of sleep then all of these mutants, no matter what the reason for their short sleep, they're all short sleeping, they're all gonna be sensitive to that particular pathogen.

Dan Pardi - 05:38: It makes [inaudible 00:05:40] sense. And let's talk how the mutation confer some sort of protective effect. Then, otherwise, you've got multiple reasons that are causing these particular flies to sleep less. And now we can really study if sleep itself has an important role here in-

Shirazu Hiza - 05:53: In immunity, but unfortunately there wasn't a consistent response. Some of the mutants were sensitive to infection, some of them were resistant to infection. Some of them were perfectly wild-type [inaudible 00:06:04] ... It just depended on the infection. They had phenotypes strip across the board, it was variable. And we thought, it seems if it was something really core, really fundamental, they would all have the same response or the same defect. At that point we thought, maybe our hypothesis is totally incorrect. And these guys they're short sleeping, but they're short sleeping for different reasons, and maybe they won't have a common phenotype. And I actually told my student, "Maybe we should find her a different project." But she was very persistent, she'd gotten hooked on the idea that we could potentially use these mutants to find a common function for sleep.

06:39: And she asked for a little more time on the project, and I said, "Sure." And she went ahead and tested a variety of different stresses. And eventually found that every single short sleeping mutant that we had collected was sensitive to oxidative stress. And she tested a couple of different ways of delivering oxidative stress and in every single time, there was a pretty strong and consistent phenotype. We decided that we'd been barking up the wrong tree. In fact, it wasn't infection, at least in Drosophila ... Drosophila immune system is quite different from mammals. If this ... Sleep may be involved in immunity in mammals. But at least in the fly, what we could find, was that this very evolutionarily conserved process, sleep is required for a very evolutionarily conserved function. Which is defense against oxidative stress.

Dan Pardi - 07:25: When you think about the utility of Drosophila fruit fly and its applicability to humans. Talk about [inaudible 00:07:31] in this model is actually useful for an evaluation and for such an investigation.

Shirazu Hiza - 07:35: Well fruit fly is really powerful, mostly because of it's genetics. It's been studied for a very long time and it's very manipulable in terms of individual gene function. We know a lot about it and we have a lot of tools that we can use to overexpress, delete or otherwise manipulate in a tissue specific way, the very specific individual gene function. The other thing that's nice about the fruit fly is that, even though it seems really different from us, it's actually very similar on a genomic level. And in terms of conserved physiology, like metabolism or sleep, there are more similarities than differences with humans. The model of Drosophila in terms of sleep, it's been pretty well established in the last five or ten years. People have proven that there are a lot of basic similarities, and a lot of the genes that have been found ... Just for circadian rhythm, which there were those three guys who got the Nobel Prize last year for their work on circadian regulation in Drosophila.

08:31: One of the reasons they did get the Nobel, is because, A, they were the first to discover circadian regulatory genes. But B, it turns out the machinery is really similar in flies as it is in humans. Everything that they discovered in the fly turns out to be applicable to humans. Sleep is a similarly very ancient, evolutionarily conserved behavior. Meaning almost all animals we know, sleep in some way or another. And that sleep is characterized by very similar types of hallmarks, two of which are, for example, regulated basically by two different mechanisms. One of which is circadian biology that regulates the timing of sleep. And in mysterious homeostatic mechanisms that regulates the amount that you sleep. For both flies and people, if you don't get to sleep as much as you need to, then at the next opportunity that you have to sleep, you'll sleep more. Suggesting that there's something keeping track of how much you need to sleep and trying to getting you that amount of sleep when it can.

Dan Pardi - 09:32: This concept of the homeostatic measuring some factors that are building up when you're awake, and then that is then partially determining the type of sleep that you get. If you're awake longer then you sleep longer.

Shirazu Hiza - 09:45: That's exactly right. When people started studying sleep in Drosophila, one of the main goals of that work was to try and understand the sleep homeostat. Because we understand that circadian biology fairly well, again using Drosophila. But the sleep homeostat is really mysterious. And we don't understand the mechanisms by which sleep amount and quality get regulated. And that's an important problem for people, especially in modern day society. I don't know about you, I'd never sleep enough.

Dan Pardi - 10:15: I travel the world talking about it, I talk about it on my show, I speak to organizations about it, and it's still a challenge.

Shirazu Hiza - 10:21: It is. That's right-

Dan Pardi - 10:22: It's still a challenge to really optimize it in our world. We're sort of fighting an uphill battle. And yet having good information about its determinants and technology that can support us and all this ... Luckily it's one of these things that we can work against. It's not this force that's high in the sky, that we really just can't do anything about. With the right knowledge, we can act in the right way, and yet we're increasingly learning of all the different things that do have an impact in this area. Great explanation of why we use fruit flies, why it's relevant, and how they have this sleep homeostat. And also circadian processes that are partially determining this situation as well. What did you exactly do with these fruit flies in your experimenting?

Shirazu Hiza - 11:00: We expose them to two different types of oxidative stress, one of which is Paraquat. Which was actually a pesticide, that's no longer used now because it generates oxidative stress and has been associated with Parkinson's. Knowing that it messes up mitochondrial function and generates a lot of oxidative stress, if you inject it into an animal in giving them a very specific high dose of oxidative stress. And over powering their natural cellular antioxidant defenses. And basically just count the number of days until they die or hours, however the case may be. And we tested a panel of short sleeping mutants and their wild-type control. And just [inaudible 00:11:40] or ask whether the short sleepers were sensitive to Paraquat injection, relative to the wild-type. And in fact they were, in everything every single case, and we can do this ... It's very consistent, we could do this experiment over and over again and get the same result.

11:53: [inaudible 00:11:53] To test whether or not that was specific to the types of reactive oxygen species that are generated by Paraquat. We tried a different source of oxidative stress, hydrogen peroxide, hydrogen peroxide actually is a reactive oxygen species. We injected that directly, and also fed that to our short sleeping mutants and compared their survival to their wild-type controls. And found, again, that they were across the board, sensitive to that source of oxidative stress. We are [inaudible 00:12:22] correlation, that these short sleeping mutants are sensitive to oxidative stress. But to show causality, we had to turn the experiment around. We used two different methods to increase sleep and asked what was the effect on their response to oxidative stress, as far as survival. The two methods that we used were, one was genetic. By stimulating sleep regulatory neurons in the brain by overexpressing a particular ion channel, we could get the flies to sleep more. And then we tested them for their response to oxidative stress in both forms again. And consistent with our hypothesis, increasing sleep caused the flies to become now resistant to oxidative stress.

Dan Pardi - 13:07: You short sleeping mutants and then you did some genetic manipulation to make them sleep more by modifying a ion channel in a sleep active area-

Shirazu Hiza - 13:16: We actually did that with just wild-type flies, otherwise wild-type flies. Right. Just to come at it from a clean slate, so to speak, in case if there was something messed up, as you were saying, about some other extra effect that their genetics we're having. But starting with wild- type flies had them sleep more either genetically, by stimulating the sleep regulatory centers in their brains or pharmacologically. We fed them a GABA agonist that caused them to fall asleep or sleep more. And then again tested them for oxidative stress, and sure enough, again, they were resistant to oxidative stress relative to untreated flies.

Dan Pardi - 13:51: Very interesting. Do we know anything about sleep staging in flies?

Shirazu Hiza - 13:55: We don't do electrophysiology ourselves, but as far as I know, there isn't really the same kind of staging of sleep in the same way as with mammals. Since we don't really know what the function of sleep is, we don't really know what the functions of each of those stages are either. It's interesting to think about whether or not different types of sleep confer different kinds of benefits.

Dan Pardi - 14:17: Yeah, a very good point. All we do know here is that, given either genetic manipulation or pharmacologically, with GABA agonists, you were able to increase the time span sleeping in these fruit flies? And then that conferred protection against these two forms of oxidative stress. One this herbicide Paraquat that catalyzes the production of superoxide ions, and then secondly, a direct oxidant which is hydrogen peroxide. Was there a reason why you chose these two distinct oxidative stressors?

Shirazu Hiza - 14:44: Not really, for technical reasons, but also they've been used in the field quite widely with fruit flies in particular. But also to test the generality of the susceptibility to oxidative stress, we've also done it with [hyporoxia 00:14:58]. That wasn't included in the paper, that's more recent work. But just to figure out the method of delivery of oxidative stress or the source of what type of oxidative stress, whether or not that was important. Whether or not it was something specific that we could learn But it really does seem it's a very general function.

Dan Pardi - 15:13: They sleep shorter, they're more susceptible to this form of stress. If they sleep longer, they become more resistant. What was the next step then?

Shirazu Hiza - 15:20: This is where we get to the oxidative [Locke's 00:15:23] theory. This fellow Raymond in the '90s who proposed a hypothesis, without any data, but it's an interesting idea. That basically the brain is a very metabolically active tissue, and metabolic activity generates reactive oxygen species and oxidative stress. Which he speculated would build up in the brain and trigger sleep. And that sleep, by shutting down the metabolic activity of the brain, would then allow antioxidant processes to clear the reactive oxygen species and restore homeostasis. And then at a certain point you'd have a low threshold of oxidative stress that would be hit and the brain would wake up. This was sort of his hypothesis for how the homeostat works. It makes sense, in an intuitive way, right? If this is the function of sleep then you would want some part of the machinery that's controlling sleep, to sense some part of the product of sleep or the function of sleep.

16:22: Sort of like the thermostat in your house. A thermometer is heat sensor to tell what the temperature is, so it knows when to turn on and off the heat. And keep your house within a window temperature. [crosstalk 00:16:34] speculated that oxidative stress would somehow trigger sleep and that's if sleep was taking care of oxidative stress. Because of this, we set out to see if changes in oxidative stress in the brain, since we know that the brain is sleep regulatory, at least part of the sleep regulation machinery. We thought, if we change oxidative stress in the brain, maybe that will change sleep behavior. My student overexpressed antioxidant enzymes in all neurons in the brain, sort of a hammer approach. But sure enough, what it did is decrease the amount of sleep of the individual fly. Which suggests that by not having as much oxidative stress, they don't need to sleep as much.

Dan Pardi - 17:17: That's interesting. Is the ability of sleep reduce oxidative stress entirely about reduced energetic load due to the relatively low energy expenditure that we have during sleep? Or are there oxidative processes that are permitted during distinct sleep states, for example, that makes the handling of oxidative stress optimized to occur during the sleep period? Is it really just energetics or is it also-

Shirazu Hiza - 17:39: Induction of specific responses or both? We don't know. That's the thing about this study, it's very intriguing, but it's all ... The tip of the iceberg is how we feel. Because there's so much that needs to be done and we expressed antioxidant enzymes in the entire brain. We know that there are sleep regulatory circuits. Are some of those sleep regulatory circuits also the places that sense reactive oxygen species or oxidative stress, or are those completely different set of neurons? We don't know. Are there defenses that get triggered during sleep? Or is it really about just shutting down metabolic load and just allowing the cell to catch up in a way, to clean up off oxidative stress by just not generating as much during sleep.

18:20: And we think this is partly why, this very clear relationship wasn't discovered previously. It's actually very hard to deprive animals of sleep without inducing a lot of stress. And the stress itself could potentially produce confounding consequences. When Raymond proposed this hypothesis in the '90s, there was a raft of studies after that, that try to come up with evidence either for or against the hypothesis. And there was compelling evidence on both sides, but they get really ... All of their data were at the time based on sleep deprivation studies of rodents that were pretty stressful for the rodents. I think it really took having a genetic model where we could deal with [inaudible 00:19:03] chronic sleep deprivation, as opposed to very stressful acute sleep deprivation to be able to parse this out.

Dan Pardi - 19:09: I think you'll find this interesting, I've experienced within and noticed that taking antioxidants prior to bedtime can have a really negative impact on my sleep.

Shirazu Hiza - 19:19: Is that right?

Dan Pardi - 19:20: Yes. And a study recently came out, I think it was [NSAIDs 00:19:23] or aspirin, suggested that they actually disrupt sleep. It wasn't that it was reducing sleep need, but it makes me feel ... Like I wake up in the morning, like I didn't get any sleep at all. It make it harder to go to sleep. [inaudible 00:19:36], if I took something like [glutathione 00:19:38] and [superoxide 00:19:38] just in case, which combined in the form of a supplement. In the morning, I would notice a quality of arousal that was different than, let's say, caffeine. It was more of a clarity in thinking. Yes, I know [inaudible 00:19:50] quite interesting.

Shirazu Hiza - 19:50: Do you often experiment on yourself Dan?

Dan Pardi - 19:52: In a way this is sort of the field of biohacking. Interesting, because there's evidence-based medicine, which is really about the quality of evidence that you have. And then there's all this new research that comes up. It doesn't quite have the level of confidence behind it through many different trials, suggesting that, yes there's a consistent effect here. But for things that seemingly have low risk but the same reward, I like the idea of investigating to see if they can add meaningfully to my wake time experience or the depth of sleep that I get. It's very challenging to come up with something that's definitively good.

Shirazu Hiza - 20:19: Is a very challenging problem in the clinic as well. And so many people come in, according to my friends who are clinicians, and have a hard time sleeping. And I went to a talk at a meeting recently and somebody had a quote that was sort of modification of [Dustyesky 00:20:36]. Basically said, something along the lines of 'Every happy sleeper is the same, but every bad sleeper is unique.' Do you know the quote that I'm talking about. It's like 'Every happy family is pretty much the same, but every bad family is pretty much unique'.

20:51: There are so many different ways to not sleep well. And I think the problem is we don't know enough about how different aspects of sleep may affect physiology. We don't know how sleep itself is regulated or what its functions are in different aspects of our physiology. It's such a fundamental mystery of science [inaudible 00:21:13]. It's amazing to me how little we know about such a whole behavior. One that affects our daily lives so fundamentally, right? I'm hoping that this will open the door for a lot of people to start, at least thinking about or considering oxidative stress on the cellular level, in terms of sleep function.

Dan Pardi - 21:30: What I have arrived now is, we understand that the currency of energy usage .. Adenosine plays a role from work [inaudible 00:21:36] and others. We bet how that relates to the triggering of immune molecules like TNF alpha and Interleukin 1, that are sort of fundamental to the initiation of sleep, and things like that [inaudible 00:21:47] things that takes place during sleep. I think the homeostat in my mind is probably influenced by a lot of different secondary signals. Oxidative stress is perfectly possible to be one of those.

Shirazu Hiza - 21:58: Exactly. And certainly it seems like, at least in the fly, is a function of sleep. The more that we can figure out about this very complicated physiological process, I think the better.

Dan Pardi - 22:08: If one of the functions of sleep is to act as an antioxidant for both the brain and the body, increasing overall resistance to acute oxidative challenges. Then leading to change and sensitivity of cells in various ways, so that they're prepared for the next day of oxidative stress. Are things that are thought to be good inducers of health, like plant phytochemicals, exercise, et cetera, that induce reactive oxygen species. Do we really need to proportion our intake of phytochemicals and the length of exercise if we are chronically under sleeping? You're getting really good sleep night after night, you can tolerate more. And if you're not, maybe what's right for you is none or a little bit. And that's a question that's probably really hard to answer, but a very interesting one in my mind.

Shirazu Hiza - 22:50: Absolutely. And in terms of a balance of oxidative stress, and regulation of [inaudible 00:22:55] function and sleep. One thing we're really interested in, is thinking about this in the context of disease. A lot of diseases, particularly neurological diseases that are associated with oxidative stress. If we're not sleeping enough or were exposing ourselves to more oxidative stress. Then we need to ... How does that impact in an individual who's genetically sensitized to neurological diseases that exacerbate their symptoms.

Dan Pardi - 23:21: [inaudible 00:23:21] gene correlates with diseases that are also associated with oxidative stress. Things like Alzheimer's disease, Parkinson's, Huntington's. There's a pretty clear connection between sleep and those conditions. That could be the mediating influences here.

Shirazu Hiza - 23:33: Right. You know for a long time it was thought that they were symptoms of pathology but they may in fact be contributing to pathogenesis as well. It's food for thought, right? Makes you think about sleeping just a little bit more, at least putting some more effort into taking care of ourselves and sleeping more.

Dan Pardi - 23:50: Does your lab plan to do more work now that you've found such an interesting connection here?

Shirazu Hiza - 23:54: Yeah. We absolutely hope to keep working on this problem. I have a student who's looking at the sleep regulation and also function. We're interested in understanding, for example, which parts of the brain are required for the reactive oxygen induced sleep. And also thinking about which organs of the body might be particularly sensitive to oxidative stress, the brain, the gut, muscles, et cetera. Maybe all of them, maybe the entire body needs sleep equally. And then also thinking about tweaking metabolism or looking at metabolism and whether and how a metabolic processes may shift between wake and sleep. Is associated with a shift in whole body metabolism, cellular metabolism. Trying to figure out which aspects of that are fundamental to sleep and might be helping to clear oxidative stress. We're moving towards a metabolic definition of sleep. Sleep as a process that is regulated by the brain, for the body.

Dan Pardi - 24:53: Every different tissue has its sensitivity to oxidative stress, calibrated to its own energy usage. And I would imagine that areas of high energy usage would be more prone, under conditions of deprivation and extra high energy users because you're up longer, to vulnerability. And that's where we see things like chronic disease take place.

Shirazu Hiza - 25:13: [inaudible 00:25:13] imagine. Exactly. But the thing is to and see if we can test that. If we can come up with experiments to test the hypothesis or that prediction. And we're still thinking about clever experiments we can do with Drosophila, to test these very specific aspects of the model. But I think that's where all the fun comes in. Just trying to come up with a very specific experiment to answer the question.

Dan Pardi - 25:33: It feels like a rich area to explore.

Shirazu Hiza - 25:35: Yeah. Oxidative species. That is a tricky beast. With a very tightly regulated, really difficult to measure and quantify. And both good and bad, in terms of their function and consequences, into extremely complicated problem.

Dan Pardi - 25:50: It's like a moving target that changes [crosstalk 00:25:54].

Shirazu Hiza - 25:54: That's right. Constantly moving, and probably different in different parts of the body and all kinds of really incredibly sophisticated mechanisms to deal with reactive oxygen species. I imagine that this will be something that we could work on for a really long time.

Dan Pardi - 26:10: I definitely want to stay in touch to hear about what you find next. Thank you for the work you do and the time that you've taken to spend with us. And I'm sure our audience appreciates it as well.

Shirazu Hiza - 26:19: Terrific. Thank you so much.

Speaker 1 - 26:23: Thanks for listening and come visit us soon at humanOS.me.