Drug-conjugated nanocrystals for imaging and assays in neuroscience

细胞表面受体、离子通道和转运体是神经系统信号传导和兴奋性的重要组成部分。因此，它们代表了目前制药行业正在探索的大多数药物靶点。基础研究表明，这些蛋白在神经元和靶标上的分布是不均匀的，这种分布可能受到多种信号转导途径和内源性调控程序的影响。此外，许多药物似乎在慢性占用后改变其蛋白靶点的反应性、分布和/或表面丰度。目前，膜蛋白的检测、定量和定位主要是利用放射性标记配体或间接利用抗体技术实现的。由于放射性示踪剂研究的空间分辨率较差，膜蛋白表面结构域选择性抗体探针的可用性有限，可用荧光团的发射光谱广泛及其光化学降解，这些方法受到限制。在本提案中，我们将继续发展我们的新颖，非同位素，标记策略，主要关注药物偶联荧光纳米晶体（纳米偶联物），它可以允许细胞表面受体和转运蛋白的成像和定量分析。具体而言，我们将：1 .合成改进的纳米晶体探针。2。建立血清素转运蛋白的动态成像。3。开发利用药物共轭纳米晶体独特特性的药理学分析。为了实现这些目标，我们组建了一个由化学家、显微镜学家、药理学家和神经科学家组成的跨学科团队。本文提出的实验利用了荧光纳米晶体独特的光学特性，这是传统的有机荧光团或荧光蛋白无法完成的。在这个提议中，纳米技术与神经科学的结合在某种程度上推动了这两个领域的发展。

Cell surface receptors, ion channels, and transporters are critical components of signaling and excitability in the nervous system. As such, they represent the majority of drug targets currently being explored in the pharmaceutical industry. Basic studies have revealed that these proteins are nonuniformly distributed on neurons and targets and this distribution can be impacted by multiple signal transduction pathways and endogenous regulatory programs. Moreover, many drugs appear to alter the responsiveness, distribution and/or surface abundance of their protein targets following chronic occupancy. Currently, the detection, quantitation and localintion of membrane proteins is achieved largely using radiolabeled ligands or indirectly with antibody techniques. These approaches are limited due to the poor spatial resolution of radiotracer studies, the limited availability of surface domain-selective antibody probes for membrane proteins, the broad emission spectra of available fluorophores and their photochemical degradation. In this proposal we will continue to develop our novel, non-isotopic, labeling strategies with a principal focus on drug-conjugated fluorescent nanocrystals (nanoconjugates) that can permit the imaging and quantitative analysis of cell surface receptor and transporter proteins. Specifically we will: I. Synthesize improved nanocrystal probes. II. Establish dynamic imaging of the serotonin transporter protein. III. Develop pharmacological assays that exploit the unique properties of drug-conjugated nanocrystals. To accomplish these aims we have assembled an interdisciplinary team of chemists, microscopists, pharmacologists and neuroscientists. The experiments proposed here exploit the unique optical properties of fluorescent nanocrystals and cannot be performed with traditional organic fluorophores or fluorescent proteins. In this proposal nanotechnology interfaces with neuroscience in a way that advances both fields.