Why is it that when you’re binge watching your favorite new series on Netflix, you can stay up for hours past your normal bedtime, even if you were tired before you started watching? On the other hand, if you were bored by a game or puzzle, you’d be much more likely to be lulled to sleep at that time. Sleep and goal-directed behaviors are mutually exclusive: you can’t do both at the same time. This is actually true of most animals, and it makes sense from an evolutionary standpoint: While sleep is undoubtedly important, there are times when it’s not a great idea – like when you’re being stalked by a predator. While this relationship is intuitively clear, scientists at Stanford have for the first time clarified the circuitry between the brain’s reward and arousal systems. And this could have implications for formulating the next generation of sleep pills.
Could Targeting the Dopaminergic System Result in Better Sleep Pills?
In the latest episode of humanOS Radio, I speak with Luis de Lecea, Professor of Psychiatry and Behavioral Sciences at Stanford University School of Medicine. Recently, he and his colleagues published a study in the prestigious journal Nature demonstrating that dopamine neuron activity (in the ventral tegmental area of the brain) is necessary in order to be awake. Furthermore, when they inhibited these neurons, there were able to promote what seemed like natural, healthy sleep. And it is that last part that is particularly exciting because this finding could lead to new sleep pills to help us sleep better when we need them (and yes, we need better sleep pills because the ones available now have lots of problems).
The other really interesting aspect of this work is how they used light to turn these neuron groups on and off at will. This research technique is called optogenetics, it’s new, and it’s leading to all sort of exciting discoveries about how brains work. So, listen below to hear how Professor de Lecea led this research to make this exciting discovery!
Luis de Lecea - 00:07: The more we know about the structure of sleep and all the behaviors associated with sleep, we'll have both developed new ways to intervene and increase the quality of sleep.
Kendall Kendrick - 00:18: HumanOS. Learn. Master. Achieve.
Dan Pardi - 00:28: Okay, Professor Luis de Lecea, thank you for joining HumanOS Radio.
Luis de Lecea - 00:32: My pleasure.
Dan Pardi - 00:33: Let's start off and, tell us a little bit about how you got into sciences, particularly how you started to study sleep and sleep circuitry.
Luis de Lecea - 00:40: It's actually an interesting story, I think. I got into science through microbiology at heart. I started this course doing science in the late eighties, and that the time, the hype was biochemistry, microbiology, the first genome had not been sequenced yet. My main interest was to determine which genes were making up the brain. It was a very naïve approach, of course, back then. I set up to investigate how many cell types, how many types of neurons there were in the brain. At the time, the technology was rather limited, but I was very attracted by this technology called subtractive hybridization, which allowed to compare different populations of nuerons. [inaudible 00:01:19] different populations of neurons.
Dan Pardi - 01:20: Okay.
Luis de Lecea - 01:21: And, that was really what my thesis was all about. To try to identify if different markers for different cell types in different brain areas.
Dan Pardi - 01:29: And where did you do that work?
Luis de Lecea - 01:30: I started in Belgium. I continued in Barcelona, and I wrapped it up in San Diego.
Dan Pardi - 01:35: Great.
Luis de Lecea - 01:36: So, I continued post-doctorate work with the same philosophy of trying to identify markers of different cell types in the brain and then I came up with this very interesting molecule, which was the [inaudible 00:01:46] neuropeptide. It was expressed exclusively in the neurocortex, so we call this molecule Cortistatin. It was expressed in the cortex and it was similar to another raw peptide called somatostatin. Then, I have no idea about how to study the function of this molecule. So, I asked my blog mates, "How could I study the function of a molecule that is expressed only in the cortex?" Someone suggested, "Well, you should learn how to EEG," and then I approached someone in the lab next door who had done an EEG. And that's really how I got into the sleep world.
Dan Pardi - 02:16: It's always interesting to ask because sleep has really grown as a field that has attracted a lot of great researchers over the last twenty to thirty years, but it's not an obvious track for a lot of people. I think, maybe nowadays, it's moreso, but back then, it was still emerging in a lot of ways.
Luis de Lecea - 02:31: Yeah, absolutely. I was extremely lucky to find Steve Erickson and his associates. They were really incredibly open to teach me the basics of sleep, where they did a lot of the experiments for me at the beginning, and we were taught by students then. So, it was really a coincidence to have these experts next to me.
Dan Pardi - 02:48: So, you've recently published some very exciting work connecting the reward system with arousal systems, but before we get into that, there's a few other things I'd like to talk about, about your background. One of them is the research that you've done revolving around narcolepsy. Very important work identifying critical aspects that helped to advance the field for narcolepsy. And then, secondly I'd like to talk a little bit about some of the newer techniques that you're using to do some of this work, because they're very cutting-edge. A lot more scientists know about them. General population does not, but it's absolutely science fiction-like.
Luis de Lecea - 03:17: That was the hypocretin orexin work. It immediately followed the cortistatin work. So, that was again, with the finding markers of specific brain regions, in that particular study, we focused on the hypothalamus. And, yes, indeed, one of the first molecules that came up from that screening was very attractive. Gene expression pattern just restricted to hypothalamus. But, again, we had no idea what this molecule was doing, and we suggested that it could be involving the thermal regulation, and maybe sleep and arousal. It was just speculation at that time, but only a couple of years after that study came out then [inaudible 00:03:50] published this cell paper in which he had knocked out this gene, and he found that the mice were narcoleptic. Then, [inaudible 00:04:02], following up this discovery, it showed that narcoleptic dogs also have this gene and a whole bunch of discoveries followed up after that. It was really an amazing privilege to be part of this set of discoveries.
Dan Pardi - 04:14: Yeah, that's a big one. Not every scientist gets to make a discovery that leads to understanding a disease that much better than we did previously. So, kudos to your work on that. Let's talk a little bit about this very interesting, science fiction-like technique: optogenetics. You've done a lot of work in that area. You've done a lot of talks about it, but can you tell us a little bit about what it is?
Luis de Lecea - 04:36: Sure. Optogenetics is a technique that is based on the discovery of this molecule that responds to blue lights. This molecule was originally found in blue algae, and the groups in Germany, and here in Stanford, hypothesize that this molecule in the algae could be introduced into neurons to make them sensitive to light. That was a great idea. So, when I joined Stanford, [inaudible 00:04:59], that was our Assistant Professor then, had just published a paper showing that indeed he could render neurons sensitive to blue light in culture. And that was a fantastic tool that needed to be implemented in people. The reason why we thought it was interesting was because before optogenetics, the only way to manipulate brain activity was either through an electrode, which would give you the precision of the location of the electrode, but it would not know which cells you are stimulating, or recalling from. The other, alternative method, would be pharmacology, which provides specificity, but it doesn't provide temporal resolution. When you study sleep, you need actually, both. You need to know where you're recalling from, you know, cell type, and a good temporal resolution because the sleep cycles in rodents are relatively short. So, that's really what drove us to try optogenetics in the first place. Indeed, that was the first paper [inaudible 00:05:47], in my lap and [inaudible 00:05:48] was a grad student back then. [inaudible 00:05:54] worked together to manipulate the hypocretin orexin cells in the hypothalamus and demonstrate that these cells were very important to facilitate the transition between sleep and wakefulness.
Dan Pardi - 06:01: Yeah. Just for the audience, if they don't understand what temporal resolution means, can you describe that?
Luis de Lecea - 06:07: Sure. Temporal resolution means that we can manipulate the activity of those neurons within milliseconds. That is the physiologically relevant time scale for neurons.
Dan Pardi - 06:16: Got it. The original discovery was that there are these blue light sensitive cells. You could then actually put these into neurons and then use light to then activate the neurons, which was better than the techniques that were used previously. You had better specificity, better time resolution. You could start to then discover new aspects about neurons and neuron function that was here to for, impossible.
Luis de Lecea - 06:37: That's correct. I forgot to mention that the beauty of this step, you can express this molecule in genetically identified neurons. In this case, we chose to target these hypocretin orexin neurons. What this does, is really to define functionally, a neurocircuit. And that has really revolutionized neurobiology in the last ten years.
Dan Pardi - 06:55: Yeah, so you could see, for a particular behavior, all the different neurons that are firing together to make that behavior or brain activity the functional unit that happens.
Luis de Lecea - 07:04: That's exactly right. You can identify the functional units underlying a specific behavior.
Dan Pardi - 07:08: Incredible. On that note, recently, this month in fact, you guys published a paper in [inaudible 00:07:13] neuroscience, right?
Luis de Lecea - 07:14: Yes.
Dan Pardi - 07:14: Yeah, and this is looking at the connection between the reward system and arousal circuits. So, tell us a little bit about the thinking that went into the design of the study, originally. What were the connections that you were interested in that made you think that there was something here?
Luis de Lecea - 07:31: Yeah, that's again ... and another surprising story, because it was surprising to us that nobody had looked at the activity of these reward neurons across the sleep-wake cycle. Intuitively, these neurons had to be involved in sleep and wakefulness, because amphetamines and psychostimulants are well-known to induce wakefulness, and maintain wakefulness for long periods of time. We were surprised that there was no direct proof that there was the case. That's why we used optogenetics. To stimulate these dopaminergic neurons, neurons that experience dopamine, that they're mainly regarded as the main driver behind the rewards and brain function.
08:08: So, she found that, indeed, when she stimulated those cells, the animals would not only wake up, but they would maintain wakefulness for very long periods of time.
Dan Pardi - 08:17: So the optogenetics could almost act like amphetamine was given?
Luis de Lecea - 08:20: Amphetamine, cocaine, it was exactly the same. The beauty of optogenetics, as you mentioned earlier, and the millisecond temporal resolution allowed us to define the minimal amount of activity that was sufficient to maintain wakefulness for very long periods of time, and amazingly we say, "Oh, 500 milliseconds ... one second per minute, was sufficient to maintain wakefulness in mice for six hours or longer.
Dan Pardi - 08:41: Interesting. So, listener, instead of giving amphetamines or drugs to activate these circuits, light pulses are given to these special neurons in these mice that are genetically engineered, and it's having an equivalent effect.
Luis de Lecea - 08:52: That's exactly right.
Dan Pardi - 08:53: Keeping them up. Okay. So, that was the connection, and through a group of researchers in your lab, set up these mice. What was the study actually like to evaluate the circuit.
Luis de Lecea - 09:02: Yeah, so, the actual surprise came immediately after because these days, it's essential not only to provide proof of the sufficiency, but also journals want to see necessity. That means that what happens when you inhibit those neurons ... so does inhibiting those neurons actually inhibit the behavior? That was the follow up experiment. To our surprise, when inhibited, these dopaminergic cells, the animals went to sleep. That had not been described before. So, then a whole new set of questions were raised, like, well, if we inhibit brain rewards, well maybe the animals get bored, and that's why they fall asleep.
09:36: So then, we challenged the animals with a whole set of stimuli that normally make them more alert or make them more aroused, and those include an opposite sex animal, [inaudible 00:09:49] specific, then also palatable food, or the other, a predator, like [inaudible 00:09:57]. Under these circumstances, the animals would explore the environment, but would still fall asleep.
Dan Pardi - 10:00: So even in the presence of things that usually keep them awake, like the delicious tasting food, the smell of opposite sex mice or the smell of a predator, they usually stay awake when they're in an environment that has those things, but instead they slept even when those things were present?
Luis de Lecea - 10:14: That's correct. There was another surprise, which was that when we inhibited dopamine, that we transferred the animals to a new cage, the animals would not fall asleep. That was somewhat puzzling because that meant that changing the cage was more arousing then other ... a predator? That really didn't make any sense to us.
Dan Pardi - 10:31: Yeah, right. When you're inactivating these neurons, can you inhibit them only a certain amount, or is it really more like an on/off switch, or can you affect the strength of how much you're turning them on and off?
Luis de Lecea - 10:41: Yeah, that's a great question. Yes, indeed, you can tune the inhibition in several ways. It doesn't have to be on and off, but the more conditions you try, the more complex the experiments become. So, for these particular conditions we just tried on and off.
Dan Pardi - 10:55: Sure.
Luis de Lecea - 10:55: So, mentioning before, what we did was then to change the [inaudible 00:11:01] new cage, and the animals that had dopamine inhibited would not go to sleep. They were very active, actually. So, we were puzzled with this finding until [inaudible 00:11:07], the leading author on the paper, realized that what the animals were doing was actually building a nest, compared to the control mice. The control mice would not touch nestlets, which is the building material for a nest. When the control animals were transferred to a new cage, they would just explore the new cage, running around, running around, and would not touch these nestlets. However, the animals that were treated with this, for to inhibit neurons, they were exclusively devoted to building a nest and then they would fall asleep.
Dan Pardi - 11:34: Interesting. So, the animals that had their dopamine neurons turned off in the VTA, in one environment where there is usually arousing stimuli, they would just sleep. But when those mice were transferred to a different cage, they didn't sleep right away, but all of the activity was devoted to then creating a sleeping area for themselves.
Luis de Lecea - 11:52: That's exactly what happened. And even to our surprise, when we transferred the mice with the old nest, in the old home cage, then the animals would fall asleep immediately. So, if there was a nest already built, the animals would fall asleep immediately.
Dan Pardi - 12:06: Interesting. So, you would think that nest building might leverage their reward system, but this would indicate that it doesn't. So, that seems to be a goal directed behavior, which is what dopamine is typically used for, these ... fueling goal directed behaviors. So, does that indicate that there's a separate circuit for nest building that's not related to brain reward?
Luis de Lecea - 12:25: Well, I think it is connected to brain reward because it only appears when dopaminergic cells are inhibited.
Dan Pardi - 12:31: Ah, yeah. Okay.
Luis de Lecea - 12:32: So, it's like an on/off switch ... like turning off dopamine cells unleashes this behavior that is conducive to sleep.
Dan Pardi - 12:39: I see, yeah. So it's not that it's separate. It's actually facilitated by its inactivity.
Luis de Lecea - 12:43: Yes. Exactly.
Dan Pardi - 12:44: Okay. That's so interesting. I'm trying to think about how that would make sense if your using your dopamine system for a little while. It becomes taxed, perhaps? And then, its inactivity will usher in a new set of behaviors that helps you go to sleep, and then it restores, and then you can do more goal directed behavior once you're awake.
Luis de Lecea - 12:59: Yeah, that opens up, again, a whole set of new questions.
Dan Pardi - 13:02: Totally.
Luis de Lecea - 13:02: Plus, it's very interesting, the fact that these sort of proprietary phases for sleep have not been described before, at least related to dopamine. That prompts us to investigate whether interfering with this proprietary phase would actually affect quality of subsequal sleep. And whether, really to define, exactly what brain patterns and behavioral patterns are conducive to sleep. That, I think, is a very new and intriguing question.
Dan Pardi - 13:26: Yeah, that does seem like it could potentially lead to novel therapeutics that would facilitate sleep behaviors other then benzodiazopines.
Luis de Lecea - 13:35: That's correct. And I think that any alternative to benzodiazepines would be great to investigate. Our hypothesis now is that if we deliver short-lived dopaminergic antagonists that facilitate these proprietary phases, then you would make a much better hypnotic than benzodiazapines.
Dan Pardi - 13:49: Yeah, interesting. Aside from just changing the behaviors around sleep, did you notice that sleep was deeper, or anything related to sleep quality?
Luis de Lecea - 13:56: We analyzed sleep quality and it was indistinguishable from a good night's sleep. So, it was just a balanced, full-blown sleep. There were no pharmacological signatures of that sleep, so that is a good sign.
Dan Pardi - 14:06: That's good news, yeah. Did you happen to, also then notice, what their behavior was like after sleep occurred?
Luis de Lecea - 14:11: Yeah, that was perfectly normal. There was nothing, no inertia, nothing that makes us worry about intervening in that sense. So, I think there's, again, potential. There's also the biological question of what does this mean, in terms of ecology, and ethology, where [inaudible 00:14:26] behaviors are related to this proprietary phase and how important, how relevant, these are for the physiology and the well-being of the animal.
Dan Pardi - 14:31: What's the next step to follow up on this research?
Luis de Lecea - 14:34: Well, there's sort of, interesting questions. I think one is actually to define exactly which neurons are responsible for this nest building behavior. We know that they are turned on by dopaminergic inhibition, but we don't know exactly which cells these are, and of course, we need to identify them. We have some ideas about what kind of cells these might be, but this is a work in progress.
14:52: And then, like I said, I'm intervening with this proprietary phase, micrologically, and see how sleep is affected, and then, I said the third line of research would be to integrate this into ... which other behaviors are related to this proprietary case? Which other transmitters may be activated or inhibited during this proprietary phase?
Dan Pardi - 15:09: Wow. That's fascinating. I can't wait to see the follow up studies on that. I agree with you that benzodiazapines and non-benzodiazapine like drugs that facilitate sleep now that are the most common remedies for people that can't get the sleep that they want, have a lot of issues with them, so novel therapeutics that can address sleep in a different way, perhaps in a more targeted way, and also, in a way with less consequences, is really, really desirable. There's a thousand applications that I can think of where people could really benefit from this. So, I hope it pans out.
Luis de Lecea - 15:36: Yeah, thank you. I hope so, too. I think that the more we know about the structure of sleep and all of the behaviors associated with sleep will help us develop new ways to intervene and increase the quality of sleep, which is one of our goals.
Dan Pardi - 15:47: Yeah, another great discovery. Dr. De Lecea, thank you for your time today and discussing this with us, it's really appreciated.
Luis de Lecea - 15:52: Thank you so much. I appreciate it.
Kendall Kendrick - 15:56: Thanks for listening and come visit us soon as HumanOS.me.