DUAL ENZYME-BASED MICROBIOSENSOR TO MEASURE ATP RELEASE FROM THE CAROTID BODY

描述（由申请人提供）：神经化学物质改变细胞或神经元的电活动，诱导基因和蛋白质表达，导致神经元或细胞表型的短期和长期改变。 因此，定义特定细胞和神经元的神经化学特征对于理解健康和疾病中正常发育和衰老的生理和病理功能至关重要。 微生物传感技术和纳米生物传感技术的最新进展为测量从神经元和细胞释放的神经化学物质的实时释放提供了途径。 ATP从中枢和外周神经系统中的神经元释放，并通过与P2 X离子门控通道结合来介导快速突触传递。 颈动脉体中的外周动脉化学感受器在调节对氧张力变化的通气和循环反应中至关重要，并且它们在早期发育期间提供基本的感觉输入以稳定和维持整个生命中的呼吸。 低氧化学敏感性的成熟发生在哺乳动物出生后发育的最初几周内。 我们推测外周动脉化学感受器中氧敏感细胞释放ATP的增加可能在介导低氧化学敏感性的发展中起作用。 为了回应节目预告（PA，03-058）的探索性/发展性生物工程研究赠款（EBRG），我们的目标是1）进一步开发一种新的基于双酶的安培微生物传感器（尖端直径5-10微米），其可以测量溶液中广泛生理条件下的ATP释放，和2）测量灌流大鼠颈动脉体的ATP释放，从而建立出生后成熟期间颈动脉体ATP释放的刺激-反应曲线。 这些研究将与格鲁吉亚理工学院的同事合作完成，他们已经开发出了用于本提案中概述的实验的微生物传感器原型。 由于突触传递是外周和中枢神经系统功能的核心基本过程，ATP是参与这一过程的重要分子，我们相信这种微生物传感器的成功制造将允许普遍应用于多种实验范式，这些实验范式询问在发育和衰老期间冥想突触传递的细胞和分子机制。 三磷酸腺苷（ATP）是一种重要的分子，使神经相互沟通。 有了正确的工具（微生物传感器），我们可以测量这种小分子，因为它是从组织和神经元释放。 这项资助计划开发一种微生物传感器，以测量从颈动脉体释放的ATP，以了解新生儿如何对氧气浓度的变化产生足够的呼吸反应。

DESCRIPTION (provided by applicant): Neurochemicals modify the electrical activity of cells or neurons, induce gene and protein expression, which lead to short and long-term modifications in the phenotype of the neuron or cell. Thus, defining the neurochemical profile of specific cells and neurons is essential to understanding physiological and pathological functions that underlie normal development and aging in health and disease. Recent advances in micro and nanobiosensing technologies has provided an avenue to measure real-time release of neurochemical released from neurons and cells. ATP is released from neurons in the central and peripheral nervous system, and mediates fast-synaptic transmission via binding to P2X ion-gated channels. The peripheral arterial chemoreceptors in the carotid body are critical in regulating ventilation and circulatory responses to changes in oxygen tension, and they provide essential sensory input during early development to stabilize and maintain breathing throughout life. Maturation of hypoxic chemosensitivity occurs within in the first several weeks of mammalian postnatal development. We hypothesize that increase release of ATP from oxygen sensing cells in the peripheral arterial chemoreceptors may operative in mediating the development of hypoxic chemosensitivity. In response to the program announcement (PA, 03-058) for exploratory/developmental bioengineering research grants (EBRG), we aim to 1) further develop a novel dual enzyme-based amperometric microbiosensor (tip diameter 5-10 mcirons) that can measure ATP release across a wide range of physiological conditions in solution, and 2) measure ATP release from the superfused rat carotid body thereby establishing the stimulus-response profile of ATP release from the carotid body during postnatal maturation. These studies will be done in collaboration with colleagues at Georgia Institutute of Technology, who have already developed the prototype of the microbiosensor to be used in experiments outlined in this proposal. Since synaptic transmission is the core elementary process in the function of the peripheral and central nervous system, and ATP is an important molecule involved in this process, we believe that successful fabrication of this microbiosenor will allow for general application to multiple experimental paradigms that interrogates cellular and molecular mechanisms that meditate synaptic transmission during development and aging. Adenosine triphosphate (ATP) is an essential molecule that allows nerves to communicate with each other. With the correct tools, (microbiosensors) we can measure this small molecule as it is released from tissues and neurons. This grant proposes to develop a microbiosensor to measure the release of ATP from the carotid body to understand how the newborn develops an adequate breathing response to changes in oxygen concentration.