NEURONAL NICOTINIC RECEPTORS FUNCTION, PHARMACOLOGY AND MODULATION BY RNA APTAME

点击翻译按钮获取中文摘要

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Neuronal nicotinic acetylcholine receptors (nAChR) are recognized as important elements in neurotransmission in the central (CNS) and peripheral (PNS) nervous systems. The major nAChRs found in the CNS and PNS are alpha-7, alpha-4/beta-2, and alpha-3/beta-4 subtypes. Activators (agonists), inhibitors (antagonists), and modulators of nAChR have been used as drug leads for treatment in Alzheimer's disease Parkinson's disease, and other neuropsychiatric illnesses such as schizophrenia, attention deficit, mood, and anxiety disorders, and chronic pain. We are interested in studying the effects of 4S, 6S- cembratrienediol (4S), an abundant cembranoid compound found in tobbaco on the alpha3beta4 nAChRs. Previous work indicate that tobbaco cembranoids accumulate in the brain in significant amounts to modify the action of nicotine. The purpose of this project is to develop receptor-binding RNA aptamers that can inhibit and/or label alpha3beta4 nicotinic receptors at the 4S cembranoid sites. RNA aptamers generated by the SELEX technique (Systematic Evolution of Ligands by Exponential enrichment) have the properties of high- affinity ligands binding to complex protein molecules including membrane-bound receptors. For this project, we will employ the SELEX selection of RNA aptamers using membrane proteins prepared from cell lines stably expressing alpha3beta4 receptors in the presence of 4S cembranoids to displace the receptor-bound aptamers. The final pool of aptamers will be sequenced and characterized according to their nAChR subtype specificity and effects on receptor function. Subtype-selective aptamers will be fluorescein-labeled and used to characterize nAChR interplay in neuronal networks.