Research tools for studying local RNA translation at synapses

研究突触局部 RNA 翻译的研究工具

• 批准号: RTI-2023-00028
• 负责人: Sephton, Chantelle
• 金额: $ 2.86万
• 依托单位: Université Laval
• 依托单位国家: 加拿大
• 项目类别: Research Tools and Instruments
• 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=760667
• 关键词: Research; tools; studying; local; RNA

Neurons are cells within the brain that transmit information to other nerve cells. Synapses are the contact points where one neuron communicates with another. My research program is aimed at understanding the mechanism by which neurons maintain synaptic connections. In particular, we want to understand how the production of proteins (i.e. the conversion of RNA to protein) by RNA-binding proteins (RBPs) occurs at synapses and how RBP activity is regulated. To study the production of proteins at the synapse and the regulation of RBP activity, fresh brain and spinal cord tissues are collected from mice. Harvested tissues are precisely cut to a 250 micron thickness with a vibratome machine, recovered in a special solution to keep them living, and then treated with different drugs or compounds. Using biochemical fractionation techniques, synapses are isolated from the fresh tissue slices and analyzed using a battery of biochemical and molecular approaches. To conduct these highly specialized experiments, we require the use of a vibratome machine that is dedicated for fresh tissues. The current vibratome that we are using is old and in disrepair. Currently, we are borrowing a new vibratome from another laboratory, but as my research team grows, the vibratome that we borrow is frequently not available. Additionally, there are logistical issues regarding where the vibratome is located. We must relocate our mice and supplies from our laboratory on the 3rd floor to the vibratome on the 5th floor to conduct our experiments. This is problematic because we must stop our time-sensitive workflow to brin our samples back to the 3rd floor for the biochemical isolation of synapses. As such, the logistics of coordinating our experiments have put our sample quality in jeopardy and slowed the progress of our projects. Thus, a vibratome is a critical and essential piece of equipment for my research program and to my trainees. This grant application is proposing the acquisition of a new vibratome for our studies. Our research program is taking an innovative and unique approach to understanding how synaptic connections are maintained and how synaptic connectivity impacts brain health and aging. Findings from our studies will act as a stepping-stone towards the long-term goal of our research program, which is to determine how protein synthesis maintains synaptic connections over the life-time of a neuron. The acquisition of a vibratome will allow us to push our research further so that we can have a better understanding of how synapses are maintained. We hope our findings will provide novel insights into the regulatory mechanisms that control local protein production and lead to the development of therapeutic approaches to treated age-related disorders, like amyotrophic lateral sclerosis (ALS), where synaptic loss is an early marker of these diseases.

神经元是大脑中将信息传递给其他神经细胞的细胞。突触是一个神经元与另一个神经元通信的接触点。我的研究计划旨在了解神经元维持突触连接的机制。特别是，我们想了解RNA结合蛋白（RBP）如何在突触上产生蛋白质（即RNA转化为蛋白质）以及RBP活性如何调节。为了研究突触处蛋白质的产生和RBP活性的调节，从小鼠收集新鲜的脑和脊髓组织。收获的组织用振动切片机精确地切割成250微米厚，在特殊的溶液中回收以保持它们的生命，然后用不同的药物或化合物处理。使用生物化学分离技术，从新鲜组织切片中分离突触，并使用一系列生物化学和分子方法进行分析。为了进行这些高度专业化的实验，我们需要使用专用于新鲜组织的振动切片机。我们目前使用的振动器已经老旧失修。目前，我们正在从另一个实验室借用一台新的振动器，但随着我的研究团队的壮大，我们借用的振动器经常无法使用。此外，还有关于振动器位置的后勤问题。我们必须将我们的小鼠和用品从3楼的实验室转移到5楼的振动切片机上进行实验。这是有问题的，因为我们必须停止我们的时间敏感的工作流程，将我们的样本带回三楼进行突触的生化隔离。因此，协调我们实验的后勤工作使我们的样品质量处于危险之中，并减缓了我们项目的进展。因此，对于我的研究计划和我的学员来说，振动器是一个关键和必不可少的设备。这份拨款申请是为了我们的研究而提出的。我们的研究计划正在采取创新和独特的方法来了解突触连接如何维持以及突触连接如何影响大脑健康和衰老。我们的研究结果将作为我们研究计划长期目标的垫脚石，即确定蛋白质合成如何在神经元的生命周期内维持突触连接。获得振动器将使我们能够进一步推动我们的研究，以便我们能够更好地了解突触是如何维持的。我们希望我们的研究结果将为控制局部蛋白质产生的调节机制提供新的见解，并导致治疗年龄相关疾病的治疗方法的发展，如肌萎缩侧索硬化症（ALS），其中突触丢失是这些疾病的早期标志。