"Feeling the Distance: Investigating the molecular mechanisms of intrinsic cell size sensing in neurons using stem cells, bioengineering and imaging"

神经元是一种特殊的细胞，它们相互连接，并与我们身体中的其他细胞类型连接，并依赖于我们神经系统向其他器官发出的信息和命令。像任何其他细胞一样，它们的形状与它们的功能密切相关，并且由于一些靶器官有时可能与大脑相距相当远，神经元本身必须跨越很远的距离。例如，脊髓运动神经元需要将脊髓与我们身体中的每一根肌肉纤维连接起来，有时可能有轴突-神经元使用“电缆”连接到其他细胞的长突起“超过1米长。这种极端的形状给神经元带来了挑战：资源（蛋白质和RNA）的主要生产中心在细胞体中，但高需求和高消耗的主要部位在1米长的轴突末端，这两个部位由一条转运分子链连接。此外，轴突末端突触的需求需要与索马中组分的生产相匹配，但对于非常长的轴突，需求可能会迅速变化，并且细胞末端之间的巨大距离使其不可能运行“即时”生产线。出于这些原因，神经元采用了一些机制来创建本地生产和调节远离细胞体并进入轴突的资源，所有这一切的一个关键问题是，神经元如何感知自己的长度并决定实施这些依赖于长度的适应，以及是否有一类特定的信号负责感知轴突长度，或者这个过程是否更多地受到多个方面的供需之间复杂相互作用的指导。这是基础神经生物学的一个基本问题，但它也与理解几种人类疾病的基础非常相关，因为在几种神经退行性疾病中，轴突远端的能量供需失衡似乎是以某些神经元死亡结束的问题链中的一些早期可观察到的事件，到目前为止，系统地研究这种机制是复杂的，因为在动物模型中，不可能以简单的方式系统地改变轴突的长度，并且细胞培养系统通常具有非常短的神经元。因此，大多数提出的神经元这种感知能力的机制都集中在相对较短的轴突上，通常远低于1 mm。我们开发了一种新的平台，将生物工程，人类干细胞和先进的成像技术相结合，以创建具有可控长度高达并超过1 cm的人类运动神经元有序阵列，我们用它成功地证明了当轴突达到一定长度时，（即“阈值长度”），因此我们完全准备系统地研究神经元大小感知背后的机制，以了解是什么决定了这个“阈值长度”。要做到这一点，我们建议使用我们的平台，并系统地改变我们观察到的所有不同的通路随轴突长度的变化，以确定它们中是否有任何一个直接负责确定“阈值”长度，如果是这样，导致神经元制定这些适应的事件序列是什么。我们的假设是，ATP（细胞能量的货币单位和所有其他功能的基础）的动态将是早期（如果不是第一个）反馈系统之一，它决定了能量水平不再可持续的长度，需要实施更多的本地生产和维护过程。

Neurons are specialised cells that connect to each other and to other cells types across our bodies and rely information and commands from our nervous system to the other organs. Like any other cell, their shape is intimately linked to their function, and as some of the target organs can be sometimes at considerable distance from the brain, neurons themselves have to span vast distances. For example, spinal Motor Neurons need to connect the spinal cord with every single muscle fibre in our body and can sometime have axons -the long protrusions that neurons use a "cable" to connect to other cells" exceeding 1m in length. This extreme shape poses a challenge for the neurons: the main centre of production for resources (proteins and RNA) is in the cell bodies, but the main site of high demand and consumption is at the end of a 1m long axon, with the two sites connected by a chain of transporter molecules. Moreover, demands in the synapses at the end of the axon need to be matched by production of component in the soma, but for very long axons demands might change swiftly and the vast distance separating the end of the cell make it impossible to run a "just-in-time" production line.For these reasons, neurons have adopted mechanisms to create local production and regulation of resources away from the cell body and into the axon, rendering the latter more independent and capable of buffering fast changes in demand.One key question in all this, is how do neurons sense their own length and decide to enact these length-dependent adaptation, and whether there is one specific class of signals responsible for sensing axonal length or if the process is more guided by a complex interplay between supply and demand across multiple fronts. This is a fundamental question for basic neurobiology, but it is also very relevant for understanding the basis of several human diseases, as in several neurodegenerative disorders imbalances in supply and demand of energy at the far end of the axon seems to be some of the earlier observable events in the chain of problems that ends with the death of certain neurons, like in Amyotrophic Lateral Sclerosis.Up until now it has been complex to study this mechanism systematically, as in animal models it is not possible to systematically change the length of axons in a simple way, and cell culture systems generally have very short neurons. As a result, most of the proposed mechanisms for this sensing capacity of neurons is centred around relatively short axons, usually well below 1mm.We have developed a novel platform that combines bioengineering, human stem cells and advanced imaging to create ordered arrays of human motor neurons with controllable length up to and exceeding 1cm, which we have used to successfully demonstrate that several important mechanisms are fundamentally altered in the axons when a certain length is reached (i.e. "threshold length") and we therefore perfectly poised to systematically study the mechanisms behind neuronal size sensing, to understand what determines this "threshold length". To do so, we propose to use our platform and systematically alter all the different pathways we observed changing with the axonal length, to determine if any of them is directly responsible for determining the "threshold" length, and if so what is the sequence of events that leads the neurons to enact these adaptations. Our hypothesis is that it will be the dynamics of ATP (the cell's unit of currency for energy and basis of all other function) that will be one of the early -if not the first- feedback system, which determines at which length energy levels are no longer sustainable and more local processes for production and upkeep need to be implemented.