Deciphering the role of microRNAs in neuronal maturation using live imaging

使用实时成像解读 microRNA 在神经元成熟中的作用

• 批准号: RGPIN-2022-04630
• 负责人: StAmour, Isabelle
• 金额: $ 2.04万
• 依托单位: Université Laval
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2022
• 资助国家: 加拿大
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=762043
• 关键词: Deciphering; role; microRNAs; neuronal; maturation

The optimal functioning of the brain depends, among other things, on a delicate balance between the excitatory and inhibitory activity of neurons. GABA, a neurotransmitter or chemical messenger involved in cellular communication, is the major player of inhibitory activity in the brain. Indeed, normally, when GABA binds to a neuron, the activity of this neuron is inhibited. However, it is interesting to note that for immature neurons, this neurotransmitter rather plays an opposite role and has an excitatory function. During the neuronal maturation, the action of GABA will gradually change from excitatory to inhibitory. This critical period is believed to play an important role in development and its timing can modify the neurodevelopmental trajectory of an individual and indeed have lasting effects on the brain functions. Whereas the proteins responsible for this change are well known, the mechanisms that regulate them are much less understood. RNAs are known to be essential cellular building blocks that carry crucial information for all functions, including protein expression. In the early 1990's, researchers discovered a new class of small RNA molecules, called microRNA. Research on this subject has since shown that these molecules are essential for neuron survival and function. MicroRNAs operate by finely regulating the protein expression in cells. In humans, over two thousand different microRNAs have been identified, each controlling a few dozen proteins with diverse functions. Our hypothesis is that some of these microRNAs are important in the fine-tuning of the excitatory to inhibitory switch of GABA action in neurons. In order to study the role of microRNAs in this process, we need to be able to monitor the effects of GABA on the activity of living neurons while varying the amount of individual microRNAs in the cells. For this research program, we will use mouse neuronal primary culture to test the developmental maturation of single cells as a function of the number of individual microRNAs. Immature neurons are usually excited by the action of GABA while mature neurons are not. We have separated this project into three distinct objectives. Firstly, we will identify which microRNAs play a significant role in neuronal maturation. Secondly, we will evaluate the effect of modulating the number of these microRNAs on neuronal activity. Finally, we will develop a new methodology, combining state-of-the art RNA sequencing approach and multiple optical microscopy techniques, to relate the molecular content of single neurons to their activity. This program will allow a better understanding of the role of microRNAs in the fine-tuning of neuronal maturation. This multidisciplinary effort will require the development of innovative strategies to study the action of microRNAs at the single cell resolution and will ultimately shed light on neurodevelopmental disorders in a multitude of contexts.

大脑的最佳功能尤其取决于神经元兴奋性和抑制性活动之间的微妙平衡。 GABA 是一种参与细胞通讯的神经递质或化学信使，是大脑抑制活动的主要参与者。事实上，正常情况下，当 GABA 与神经元结合时，该神经元的活动就会受到抑制。然而，有趣的是，对于未成熟的神经元，这种神经递质却起着相反的作用，具有兴奋功能。在神经元成熟过程中，GABA的作用会逐渐从兴奋性转变为抑制性。这个关键时期被认为在发育中发挥着重要作用，其时机可以改变个体的神经发育轨迹，并且确实对大脑功能产生持久影响。尽管造成这种变化的蛋白质众所周知，但调节它们的机制却知之甚少。众所周知，RNA 是重要的细胞构建模块，携带着包括蛋白质表达在内的所有功能的关键信息。 20 世纪 90 年代初，研究人员发现了一类新的小 RNA 分子，称为 microRNA。此后对该主题的研究表明，这些分子对于神经元的生存和功能至关重要。 MicroRNA 通过精细调节细胞中的蛋白质表达来发挥作用。在人类中，已鉴定出超过 2000 个不同的 microRNA，每种 microRNA 控制着几十种具有不同功能的蛋白质。 我们的假设是，其中一些 microRNA 对于神经元中 GABA 作用的兴奋性到抑制性转换的微调非常重要。为了研究 microRNA 在此过程中的作用，我们需要能够监测 GABA 对活神经元活动的影响，同时改变细胞中单个 microRNA 的数量。 在本研究项目中，我们将使用小鼠神经元原代培养物来测试单细胞的发育成熟度与单个 microRNA 数量的函数关系。未成熟的神经元通常会因 GABA 的作用而兴奋，而成熟的神经元则不会。我们将该项目分为三个不同的目标。首先，我们将确定哪些 microRNA 在神经元成熟中发挥重要作用。其次，我们将评估调节这些 microRNA 数量对神经元活动的影响。最后，我们将开发一种新的方法，结合最先进的 RNA 测序方法和多种光学显微镜技术，将单个神经元的分子内容与其活动联系起来。 该计划将使人们更好地了解 microRNA 在神经元成熟微调中的作用。这项多学科的努力需要开发创新策略来研究单细胞分辨率下 microRNA 的作用，并最终揭示多种情况下的神经发育障碍。