Determining the membrane circadian clock across evolution

生物钟存在于所有动物体内，是生命的基础，这使得研究动物体内的生物钟变得容易保存、衰老和做实验。因此，几乎所有的时钟基因以及它们是如何形成分子钟的，都是在果蝇身上发现的，后来发现在小鼠和人类身上也是保守的。这一点以及昼夜节律深刻影响生理学和健康的各个方面的事实，导致诺贝尔奖被授予了飞行昼夜节律研究人员。分子钟由生物钟基因组成，这些基因在大脑的生物钟神经元中每24小时开关一次。在我们这个“24/7社会”中，越来越多的人由于轮班工作、接触科技产品、抗睡眠药物、不规律的睡眠和饮食模式，他们的生物钟与外部世界不同步。这种所谓的“社交时差”，就像时差一样，会导致健康风险的惊人增加，与心理健康危机、癌症、糖尿病、成瘾、代谢和睡眠障碍有关，30%的人患有失眠症。此外，40-70%的老年人患有阿尔茨海默病和帕金森病的慢性睡眠障碍，导致更明显的昼夜节律和睡眠缺陷，从而导致他们的病理。我们还表明昆虫时钟对传粉者的健康和粮食安全很重要，并且受到靶向时钟神经元离子通道的杀虫剂的负面影响。大脑中的主时钟由一个由时钟神经元组成的神经回路组成，它通过受分子时钟双向影响的电信号接收和传输一天中的时间信息。电信号是由膜上的离子通道产生的，这些离子通道控制着神经元之间的化学信号的释放，这些化学信号在整个时钟中传递时间信息，并传递给大脑和身体的其他部分。我们已经证明，在苍蝇和老鼠身上，时钟神经元的膜在白天比晚上更容易电兴奋。这些离子通道产生的电信号被认为有助于同步时钟神经元，并与分子钟相互作用，从而产生一个整体上强大而持续的昼夜节律。我们已经证明这种能力随着年龄的增长而减弱。然而，关于这种所谓的膜钟的确切组成和机制，以及它是如何受到衰老影响的，目前还存在知识缺口。我们希望确定在果蝇、小鼠和衰老中产生昼夜时钟神经元兴奋性差异的时钟神经元离子通道。因此，我们的目标是通过果蝇、小鼠和计算模型来破译膜时钟的组成和机制，以验证果蝇和小鼠时钟神经元中存在一组保守的离子通道，这些离子通道产生每日的电变化。这将揭示昼夜行为的电学基础，以及是否通过操纵膜时钟变成老鼠，可以让苍蝇变成夜行动物。我们将测试衰老是否会改变这些日常变化，以及它对昼夜节律和健康跨度的影响。因此，目标是：1：在果蝇和小鼠中，确定哪些离子通道产生时钟神经元电活动的昼夜差异；2：在果蝇和小鼠中，生成基于离子通道的时钟神经元计算模型，并使用动态钳（DC）来测试因果关系；3：在果蝇中，确定由我们的模型和DC4预测的时钟神经元离子通道的特定变化的昼夜节律后果；在果蝇中，确定衰老对产生生物钟神经元电活动昼夜差异的离子通道的影响本研究将确定在整个健康周期内产生强大昼夜节律所需的生物钟神经元离子通道，揭示我们将测试的干预措施，以恢复旧生物钟神经元的兴奋性并逆转昼夜节律老化。这将为昼夜节律、睡眠和衰老障碍开发新的药物靶点和治疗方法。

Circadian clocks are found in all animals and are fundamental to life allowing the study of clocks in animals that are easy to keep, age and do experiments with. Hence, nearly all clock genes and how they form a molecular clock were identified in flies and later found to be conserved in mice and humans. This and the fact that circadian rhythms profoundly affect all aspects of physiology and health, led to Nobel prizes being awarded to fly circadian researchers. The molecular clock consists of clock genes that switch themselves on and off every 24hours in the clock neurons of the brain. In our '24/7 society', an increasing proportion of the population experience de-synchronisation of their circadian clock with the external world, due to shift-work, access to technology, anti-sleep drugs, irregular sleep and eating patterns. This so-called 'social jetlag', like jetlag can contribute to an alarming increase in health risks, being associated with the mental health crisis, cancer, diabetes, addiction, metabolic and sleep disorders, with 30% of people experiencing insomnia. Furthermore 40-70% of the elderly population experiences chronic sleep disturbances with Alzheimer's and Parkinson's causing more pronounced circadian and sleep deficits contributing to their pathology. We have also shown that insect clocks are important for pollinator health and food security and are negatively impacted by insecticides that target clock neuron ion channels. The master clock in the brain consists of a neural circuit of clock neurons that receive and transmit time of day information via electrical signals that are bi-directionally influenced by the molecular clock. Electrical signals are generated by ion channels in the membrane which control the release of chemical signals between neurons relaying temporal information throughout the clock and to the rest of the brain and body. We have shown that the membrane of clock neurons is more electrically excitable in the day than at night in flies and mice. These ion channel generated electrical signals are thought to help synchronise clock neurons and interact with the molecular clock to generate an overall strong and sustained circadian rhythm. We have shown this weakens with age. However there is a knowledge gap in the exact components and mechanism of this so-called membrane clock and how it is affected by ageing. We wish to determine the clock neuron ion channels that generate day v night differences in clock neuron excitability in flies, mice and ageing. Therefore our aim is to decipher the components and mechanism of the membrane clock using flies, mouse and computational models testing the hypothesis that there is a conserved set of ion channels that generate daily electrical variations in fly and mouse clock neurons. This will reveal the electrical basis of diurnal v nocturnal behaviour and whether by manipulating the membrane clock to become mouse like you can make a fly become nocturnal. We will test if ageing alters these daily changes and what effect it has on circadian rhythms and health span. Therefore the objectives are:1: In flies and mice determine which ion channels generate the day/night differences in electrical activity of clock neurons2: In flies and mice generate ion channel-based computational models of clock neurons and use dynamic clamp (DC) to test causality3: In flies determine the circadian consequences of making specific changes in clock neuron ion channels predicted by our models and DC4: In flies, determine the effect of ageing on ion channels that generate day/night differences in clock neuron electrical activityThis research will identify the clock neuron ion channels required to generate robust circadian rhythms throughout the health span revealing interventions that we will test to rejuvenate the excitability of old clock neurons and reverse circadian ageing. This will develop new drug targets and treatments for circadian, sleep and ageing disorders.