GENETIC DETERMINANTS OF ODORANT RESPONSIVENESS

本研究的总体目标是了解嗅觉信息在嗅上皮水平上的处理逻辑及其向嗅球的投射。我们测试的假设是，嗅觉受体（OR）基因的产物是嗅觉感觉神经元（OSN）的气味反应的关键决定因素。我们的方法涉及构建或使用基因靶向小鼠，其中表达给定OR基因的活神经元可以通过其绿色荧光蛋白（GFP）的表达来鉴定和分离。这种遗传策略提供了一个独特的和前所未有的机会，研究活的神经元，表达一个定义的或从内源性基因。将用钙成像技术测定对气味剂组的反应。这种强大的和新颖的组合，在小鼠和钙成像的靶向诱变将揭示哺乳动物嗅觉编码的一般原则。在具体目标1中，我们将批判性地研究OR对表达它们的OSN的气味反应的贡献。我们将确定一个特定的OR基因的表达是否与一个特定的气味反应谱密切相关。同时，我们将确定一个OR基因的表达是否是必要的和足够的赋予气味反应谱。我们的目标是通过功能手段研究单受体/单神经元假说，并对OR在气味反应中的作用进行关键评估。在具体目标2中，我们将以相反的方式研究嗅觉。从一组定义的气味剂开始，我们将确定OSN中表达的OR的光谱，这些OSN对这些气味剂有反应。面对嗅觉系统令人生畏的数字复杂性，我们设计了两种方法来将这个问题简化为可管理的格式。我们的目标是定义配体/受体的气味和ORS的生理相关的背景下的关系。在特定目标3中，我们将克隆映射到嗅球特定区域中10-20个肾小球的OR基因，并确定表达这些基因的OSN子集的气味反应谱。我们想为球茎的一小部分编制一个综合地图，包括遗传解剖和功能数据。我们的目标是以前所未有的分辨率率先组装嗅觉系统图谱。综合起来，这些研究将成为哺乳动物嗅觉生理学和分子生物学之间期待已久且急需的桥梁。

The overall aim of our studies is to understand the logic of the processing of olfactory information at the level of the olfactory epithelium and its projection to the olfactory bulb. The hypothesis that we test is that the products of the olfactory receptor (OR) genes are critical determinants of the odorant responsiveness of olfactory sensory neurons (OSNs). Our approach involves the construction or use of gene-targeted mice in which live neurons expressing a given OR gene can be identified and isolated by virtue of their expression of the green fluorescent protein (GFP). This genetic strategy provides a unique and unprecedented opportunity to study living neurons that express a defined OR from endogenous genes. Responses to panels of odorants will be assayed with calcium imaging techniques. This powerful and novel combination of targeted mutagenesis in the mouse and calcium imaging will reveal general principles of mammalian olfactory coding. In Specific Aim 1, we will critically examine the contribution of ORs to odorant responses of the OSNs that express them. We will determine if expression of a particular OR gene correlates tightly with a particular odorant response profile. Simultaneously we will determine if expression of an OR gene is both necessary and sufficient to confer an odorant response profile. The goal is to examine the single receptor/single neuron hypothesis by functional means and to provide a critical assessment of the role of ORs in odorant responsiveness. In Specific Aim 2, we will study olfaction in the reverse manner. Starting with a defined set of odorants, we will determine the spectrum of ORs that are expressed in OSNs that respond to these odorants. Faced with the daunting numerical complexity of the olfactory system, we have devised two ways of reducing this question to a manageable format. The goal is to define ligand/receptor relationships of odorants and ORS within a physiologically relevant context. In Specific Aim 3, we will clone OR genes that map to 10-20 glomeruli in a defined area of the olfactory bulb and determine the odorant response profile of the subsets of OSNs that express these genes. We want to compile an integrated map for a small part of the bulb, comprising genetic anatomical and functional data. The goal is to pioneer the assembly of an atlas of the olfactory system at a resolution that has never been achieved before. Taken together, the proposed studies should form the long-awaited and much-needed bridges between the physiology and molecular biology of olfaction in mammals.