Imaging exocytosis via a combined fluorescence and amperometry approach

在化学系生命过程化学（CLP）项目的支持下，密苏里州大学的蒂莫西·格拉斯和凯文·吉利斯教授正在研究神经递质是如何释放到突触中的。神经递质对神经系统的调节至关重要，并控制许多功能，如学习，记忆，睡眠和运动。了解突触释放的机制和神经递质的化学活性对于了解正常和非典型神经元过程至关重要。此外，了解突触囊泡融合和通过胞吐释放递质的基本机制具有广泛的意义，因为它不仅有助于开发神经递质释放受损的疾病的治疗方法，而且还将促进我们对FDA批准的调节递质释放的治疗方法的理解，例如肉毒杆菌毒素A和B。作为这项工作更广泛影响的一部分，将为服务不足的高中生举办一个新的机器人夏令营。实践证明，这些夏令营等活动提高了参与者在中学期间学习STEM（科学、技术、工程和数学）课程的可能性。本项目涉及制备和评估用于荧光检测神经递质的儿茶酚胺荧光化学传感器。这些传感器与PI Glass实验室开发的NeuroSensor类探针有关，该探针已用于检测分离的嗜铬细胞中的去甲肾上腺素。传感器将被开发，产生荧光增强后胞吐作用。这些传感器将与PI Gillis实验室开发的新型透明电极结合使用，通过荧光成像结合电流测量来研究胞吐作用的机制。这些组合实验将测量从单个囊泡释放和保留的儿茶酚胺，以测试从囊泡释放的儿茶酚胺的量受刺激强度调节的假设。该项目有望为神经化学研究开发和应用一套新的重要化学生物学工具，具有潜在的广泛科学影响。该奖项反映了NSF的法定使命，并被认为值得通过使用基金会的知识价值和更广泛的影响审查标准进行评估来支持。

With the support of the Chemistry of Life Processes (CLP) Program in the Division of Chemistry, Professors Timothy Glass and Kevin Gillis of the University of Missouri are studying how neurotransmitters are released into synapses. Neurotransmitters are critical to the regulation of the nervous system and control a number of functions such as learning, memory, sleep, and movement. Understanding the machinery of synaptic release, and the chemical activity of neurotransmitters is vital to understanding both normal and atypical neuronal processes. Furthermore, understanding the basic mechanisms of synaptic vesicle fusion and transmitter release via exocytosis is of broad significance because it will not only aid in the development of therapies for diseases where release of neurotransmitters is compromised, but it will also advance our understanding of FDA-approved treatments that modulate transmitter release, such as botulinum toxin A and B. As part of the broader impacts of this work, a new robotics camp for underserved high school students will be held. It has been demonstrated that activities such as these camps enhances the likelihood that participants pursue STEM (science, technology, engineering and mathematics) coursework during their secondary education.This project involves the preparation and evaluation of fluorescent chemical sensors for catecholamines with a view toward the fluorescent detection of neurotransmitters. These sensors are related to the NeuroSensor class of probes developed in PI Glass’s lab, which have already been used to detect norepinephrine in isolated chromaffin cells. Sensors will be developed that produce fluorescence enhancements upon exocytosis. The sensors will be used in combination with novel transparent electrodes developed in PI Gillis’ lab to study mechanisms of exocytosis via fluorescence imaging in combination with amperometric measurements. These combination experiments will measure both release and retention of catecholamine from individual vesicles to test the hypothesis that the amount of catecholamine released from a vesicle is modulated by the stimulus intensity. This project is expected to result in the development and application of a new set of important chemical biology tools for the study of neurochemistry with potentially broad scientific impact.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.