Testing the role of sleep in homeostatic plasticity

我们一生中有三分之一的时间都在睡觉，没有意识，与世界隔绝。这似乎是浪费时间，然而所有的动物都要睡觉，所以睡眠似乎必须完成一些重要的生物学目的。这个目的可能是什么？睡眠可以提高记忆力，当我们睡眠不足时，我们的大脑工作效率会降低，这表明睡眠具有神经功能。事实上，有一种假设认为，睡眠是大脑保持稳定活动水平所必需的。这一点很重要，因为大脑中的神经元都是相互连接的：当一个神经元发出电脉冲时，它会向其他神经元发送化学信号，这些信号要么激发它们（使它们放电），要么抑制它们（阻止它们放电）。如果兴奋和抑制变得不平衡，神经网络就会失去控制，陷入痉挛（过度兴奋）或沉默（过度抑制）。然而，当我们根据感官经验学习时，我们的大脑会不断变化。为了阻止这些不平衡的兴奋和抑制变化，大脑重新调整神经元及其连接来补偿这些变化并恢复稳定的活动水平，这一过程被称为“稳态可塑性”。研究人员认为，这一过程最好在睡眠期间进行，这是一段不活动的时间，几乎没有感官输入，就像商店在下班后检查库存一样。尽管这一观点有很多支持证据，但目前尚不清楚睡眠是如何参与稳态可塑性的。首先，特定类型的体内平衡可塑性需要睡眠吗？一个有影响力的假设认为，神经元之间的联系主要在我们清醒时加强，在我们睡觉时减弱。这一观点得到了很多（但不是全部）证据的支持。睡眠在体内平衡可塑性中的作用是否会受到另一种逻辑的支配？例如，也许睡眠对调节神经元之间的连接强度很重要，但对神经元自身被其他神经元兴奋的内在能力（“兴奋性”）却没有作用，或者对调节兴奋性而非抑制性神经元的活动很重要。第二，睡眠通过什么分子机制影响稳态可塑性？两个有趣的候选者是“活性氧”（代谢的副产物，可能是危险的，但也起着重要的信号作用）和某些突触蛋白（帮助神经元相互传递信号的分子）的水平。每一种都受睡眠调节，并在体内平衡可塑性中发挥作用。睡眠影响体内平衡可塑性的一个或两个可能是共同的联系吗？我们将利用果蝇的嗅觉系统来解决这些问题。像人类一样，苍蝇也会睡觉，我们已经开发了一个新的模型系统来研究苍蝇完整大脑中的稳态可塑性。在这里，被称为“凯尼恩细胞”的神经元兴奋，并被称为“APL”的神经元抑制。如果我们人为地激活APL 4天（产生过度抑制），该回路会补偿这种扰动，当我们解除人为施加的过度抑制时，凯尼恩细胞的活性会更高。这种效应的产生，一方面是因为APL变得不那么活跃，另一方面是因为凯尼恩细胞得到了更多的兴奋，而这需要睡眠：当我们阻止果蝇睡觉时，它会减少，当我们使用一种基因技巧迫使它们额外睡觉时，它会增强。我们将通过测试睡眠是否需要(1)各种形式的体内平衡可塑性（例如，来自凯尼恩细胞的过度兴奋，过度暴露于自然气味）和(2)不同可能的潜在细胞机制（例如，神经元之间连接强度的变化或内在兴奋性）来测试睡眠需要哪种类型的体内平衡可塑性。我们将通过测量和操纵正常和睡眠剥夺的果蝇的活性氧和突触蛋白水平来测试睡眠如何调节稳态可塑性，并测试这是如何影响稳态可塑性的。

We spend 1/3 of our lives asleep, unconscious and disconnected from the world. It seems like a waste of time, yet all animals sleep, so it seems sleep must fulfill some important biological purpose. What might that purpose be?Sleep improves memory and our minds work less well when we're sleep-deprived, suggesting that sleep has a neurological function. Indeed, one hypothesis posits that sleep is required for the brain to maintain stable levels of activity. This is important because neurons in the brain are all connected: when a neuron fires an electrical impulse, it sends chemical signals to other neurons that either excite them (make them fire) or inhibit them (stop them from firing). If excitation and inhibition become imbalanced, a neural network can spiral out of control into a seizure (too much excitation) or silence (too much inhibition). Yet our brains are constantly changing as we learn based on sensory experience. To stop these changes from unbalancing excitation and inhibition, the brain readjusts neurons and their connections to compensate for the changes and restore stable activity levels, a process called "homeostatic plasticity". It's thought that this process might be best carried out during sleep, a time of inactivity with little sensory input - much as shops do inventory checks after hours.Although this idea has much supporting evidence, it's not clear exactly how sleep is involved in homeostatic plasticity. First, is sleep specifically required for particular *kinds* of homeostatic plasticity? One influential hypothesis posits that connections between neurons are mainly strengthened when we're awake, and weakened when we're asleep. This idea is supported by much, but not all, evidence. Could sleep's role in homeostatic plasticity be governed by a different logic? For example, perhaps sleep is important for adjusting the strength of connections between neurons but not neurons' own intrinsic ability to be excited by other neurons ('excitability'), or for adjusting the activity of excitatory but not inhibitory neurons. Second, by what molecular mechanisms does sleep influence homeostatic plasticity? Two interesting candidates are "reactive oxygen species" (byproducts of metabolism that can be dangerous yet also play important signaling roles) and levels of certain synaptic proteins (molecules that help neurons signal to each other). Each one is regulated by sleep and plays a role in homeostatic plasticity. Could one or both be a common nexus by which sleep influences homeostatic plasticity?We will address these questions using the olfactory system of the fruit fly Drosophila. Like humans, flies sleep, and we have developed a new model system for studying homeostatic plasticity in the intact brain in flies. Here, neurons called "Kenyon cells" excite, and are inhibited by, a neuron called "APL". If we artificially activate APL for 4 days (producing excess inhibition), the circuit compensates for the perturbation, which is revealed as higher activity in Kenyon cells when we lift the artificially imposed excess inhibition. This effect arises both because APL becomes less active and because Kenyon cells get more excitation, and it requires sleep: it's reduced when we stop flies from sleeping, and it's enhanced when we use a genetic trick to force them to sleep extra.We will test what kinds of homeostatic plasticity sleep is required for, by testing whether sleep is required for (1) a variety of forms of homeostatic plasticity (e.g., excess excitation from Kenyon cells, excess exposure to natural odours) and (2) different possible underlying cellular mechanisms (e.g., changing connection strength between neurons or intrinsic excitability). We will test *how* sleep modulates homeostatic plasticity by measuring and manipulating reactive oxygen species and synaptic protein levels in normal and sleep-deprived flies and testing how this affects homeostatic plasticity.