Drug-conjugated nanocrystals for imaging and assays in neuroscience

细胞表面受体、离子通道和转运蛋白是神经系统中信号传导和兴奋性的关键组成部分。因此，它们代表了制药行业目前正在探索的大多数药物靶标。基础研究表明，这些蛋白质在神经元和靶点上的分布是不均匀的，并且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，许多药物似乎会改变长期占用后其蛋白质靶标的反应性、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的配体或间接利用抗体技术来实现的。这些方法是有限的，由于放射性示踪剂研究的空间分辨率差，有限的表面结构域选择性抗体探针的膜蛋白，可用的荧光团和它们的光化学降解的广泛的发射光谱。在这项提案中，我们将继续开发我们的新的，非同位素，标记策略，主要集中在药物共轭荧光纳米晶体（nanoconjugates），可以允许成像和定量分析细胞表面受体和转运蛋白。具体而言，我们将：一。合成改进的探针。二.建立血清素转运蛋白的动态成像。三.开发利用药物结合纳米晶体独特性质的药理学测定。为了实现这些目标，我们组建了一支由化学家、显微镜学家、药理学家和神经科学家组成的跨学科团队。这里提出的实验利用了荧光纳米晶体的独特光学性质，并且不能用传统的有机荧光团或荧光蛋白进行。在该提案中，纳米技术与神经科学的结合方式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.