Receptor driven cellular behavior, ranging from signaling and excitability in neural activity to immune system response to disease invasion, represents an important class of functional nanobiostructures in biological systems. Receptors are uniquely suited to direct such processes due to their ability to sense the environment, through ligand binding, and their ability to transmit this signal to the cell interior via signal cascades. The goal of this proposal is the development of novel imaging tools and methods to establish a quantitative molecular level understanding of the function of biological receptors. Unfortunately, the study of these fundamental biological components is limited by currently available imaging tools such as radiolabeled ligands or indirect detection via antibodies. These approaches suffer due to the poor spatial resolution of radiotracer studies, the limited availability of surface-domain 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 strategy involving ligand-conjugated fluorescent nanocrystals (nanoconjugates). Specifically, we will: 1. Design, synthesize, and characterize novel nanoconjugate probes for imaging neural receptors. 2. Develop a molecular level understanding of nanoconjugate-receptor interactions. 3. Demonstrate dynamic imaging of neurotransmitter receptors in order to map their regulation and function. To accomplish these specific aims, we have assembled a multidisciplinary team including chemists, physicists, pharmacologists and neuroscientists. Completion of this grant will result in the development of a novel class of nanoconjugates based on highly fluorescent small-molecule and peptide conjugated CdSe/ZnS core shell nanocrystals. These probes will enable investigators in neuroscience and membrane biology to answer questions previously unanswerable due to the limitations of current methods. The nanoconjugates described herein present the opportunity for dynamic imaging of fundamental processes involving neural G protein-coupled receptors and transporter proteins. Such """"""""real time"""""""" experiments will provide new insight into questions concerning the molecular details of these neuroreceptors, their trafficking and localization in response to external stimuli. Such results will provide new molecular level insights into neural processes such as depression, addiction and learning. Additionally, the nanoconjugates may serve as the basis for new drug discovery methods to identify unique drugs that target specific receptors previously implicated in neurological disorder.
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