Development and Validation of an NPY-sensitive Microelectrode for Measuring NPY Release from Hippocampus

该研究计划旨在开发和验证对神经肽 Y 敏感的微电极 (NPY) 用于测量海马体中 NPY 的释放。这样我们就可以找到相关性 NPY 水平与焦虑症之间的关系。为了做到这一点，有两种电化学策略 旨在实时监测生物分子。大脑中的非电活性分子很难测量 具有高时间和空间分辨率的神经肽一直是一个挑战。基于电化学 技术功能强大，可用于测量表面和化学性质 它们已广泛用于检测检测限非常低的分子。的组合 具有快速扫描循环伏安法和连续电化学阻抗的高选择性适配体 测量将提供两种新的策略来了解 CA1 区域中 NPY 的存在。 可能与 NPY 一起释放的不同分子将使用开发的技术进行测量 微电极证明选择性。转基因小鼠将受到过度监管和过度监管 四环素改变 NPY 水平并使用开发的 NPY 敏感确认测量 微电极。尺寸达 25 微米的铂微电极将提供适当的 用于分子吸附和解吸以及适体修饰过滤NPY的底物 来自其他混杂分子的信号。伴随电化学和电生理学 CA1 中的测量将用于从测量的不同其他信号中过滤 NPY。的确认 将测试 NPY 的效果，记录 SC 中低频电刺激的 fEPSP 和TA途径。为了验证 NPY 水平，将使用 ELISA 来比较与 我们在海马提取物中开发了 NPY 敏感微电极。 电化学阻抗谱和快速扫描循环伏安法已被证明很重要 允许测量法拉第电流以及非法拉第电流的技术，提供了 电活性物质以及与电极表面相互作用的非电活性物质。的 快速扫描循环伏安法与电化学阻抗测量的结合将增强 研究界使用微电极实时测量生物分子，例如 神经递质和神经肽。

This research proposal aims to develop and validate microelectrode that is sensitive to neuropeptide Y (NPY) for measuring the release of NPY from hippocampus. In this way, we will be able to find correlations between NPY levels with anxiety disorders. In order to do this, two electrochemical strategies have been devised to monitor biomolecules in real-time. Non-electroactive molecules in the brain are difficult to measure with high temporal and spatial resolution and neuropeptides have been a challenge. Electrochemical-based techniques are powerful and can be used to measure the physical and chemical properties of the surface and they have been vastly used for the detection of molecules with very low detection limits. The combination of highly selective aptamers with fast scan cyclic voltammetry and continuous electrochemical impedance measurements will provide two novel strategies to understand the presence of NPY in the CA1 region. Different molecules that are potentially released together with NPY will be measured using the developed microelectrodes to prove selectivity. Genetically modified mice will be under and overregulated using tetracyclines to change NPY levels and confirm the measurement using the developed NPY-sensitive microelectrodes. Platinum microelectrodes measuring up to 25 micrometers will provide the appropriate substrate for the adsorption and desorption of molecules as well as the aptamer modification to filter NPY signals from other confounding molecules. Concomitant electrochemical and electrophysiological measurement in CA1 will be done to filter NPY from the different other signals measured. The confirmation of the effects of NPY will be tested recording fEPSPs in response to low-frequency electrical stimulation in the SC and TA pathway. In order to validate NPY levels, ELISA will be used to compare the measurements done with our developed NPY-sensitive microelectrodes in hippocampal extracts. Electrochemical impedance spectroscopy and fast scan cyclic voltammetry have shown to be important techniques that allow the measurement of faradaic as well as non-faradaic currents providing a picture of the electroactive species as well as non-electroactive species that interact with the electrode’s surfaces. The combination of fast scan cyclic voltammetry with electrochemical impedance measurements will empower the research community using microelectrodes for real-time measurement of biomolecules such as neurotransmitters and neuropeptides.