Transduction Mechanism in Olfactory Receptor Neurons

嗅觉受体神经元的转导机制

• 批准号: 7873021
• 负责人: KING-WAI YAU
• 金额: $ 34.5万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-01 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7873021
• 关键词: Action Potentials; Adenylate Cyclase; Affect; Amphibia; Arrestins; Attention; Binding; Brain; Cilia; Complex; Cyclic Nucleotides; Detection; Disabled Persons; Disease; Dissociation; Event; Feedback; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; Genetic; Genetically Engineered Mouse; Golf; Grant; Health; Human; Individual; Insecta; Membrane; Mus; Nose; Odorant Receptors; Olfactory Receptor Neurons; Pathway interactions; Phosphorylation; Phototransduction; Physiological; Probability; Rana; Relative (related person); Reporting; Retinal Pigments; Shapes; Signal Transduction; Smell Perception; Solutions; Stimulus; Suction; System; Time; Vertebrate Photoreceptors; genetic manipulation; insight; public health relevance; receptor; research study; response; retinal rods

DESCRIPTION (provided by applicant): PROJECT SUMMARY The long-term objective of this proposal is to understand in quantitative detail the transduction mechanism in vertebrate olfactory receptor neurons (ORNs). During the current grant period, we have been able to resolve the response triggered by a single odorant-binding event on an ORN (the so- called "unitary response"). Surprisingly, we found that randomly encountered ORNs (and therefore randomly encountered odorant receptors, or ORs) all gave unitary responses of similar amplitudes, suggesting that an odorant-binding event has a very low probability of signaling downstream. In other words, the unitary response appears to reflect the effect of a single activated Golf/adenylyl cyclase complex. This phenomenon is apparently due to a very short odorant dwell-time on the OR molecule (i.e., rapid dissociation of the odorant from the OR). This short odorant dwell-time likewise dominates the termination of olfactory transduction. Hence, the traditionally believed determinants for terminating G-protein-coupled receptor (GPCR) signaling, namely, receptor phosphorylation and subsequent arrestin binding, are unimportant at least at the level of the unitary response, although they may still be important with intense and prolonged odorant stimulation. So far, the above results have been obtained from amphibian ORNs and in low-Ca2+ solution (in order to boost the unitary-response amplitude). In this application, we propose to continue experiments with amphibian ORNs but at the same time to use also mouse ORNs, which offer the distinct advantage of genetic manipulations. Aim 1 is to obtain the best estimate of the olfactory unitary-response amplitude in frog ORNs in physiological conditions (i.e., normal external Ca2+), to characterize its spatial spread along the olfactory cilium, to dissect its membrane-current components (cyclic-nucleotide-gated current versus Cl current), and to estimate the number of unitary responses required for bringing the ORN to firing threshold. Aim 2 is to launch a similar detailed study of the unitary response in mouse ORNs, which will serve as the groundwork for examining available genetic mouse lines for olfactory transduction. Aim 3 is to study/dissect Ca-dependent and Ca-independent adaptation by ORNs in more detail, largely by making use of genetic mouse lines. Aim 4 is to study constitutive OR activity in the absence of odorants, which so far has received little attention in vertebrate olfactory transduction. We have noticed this phenomenon in the course of previous experimentation. The experimental approach will involve predominantly suction-pipette recording from single ORNs of frog and WT or genetically engineered mice. PUBLIC HEALTH RELEVANCE: PROJECT NARRATIVE The studies proposed in this application will enhance our understanding of olfactory transduction in vertebrate olfactory receptor neurons in the nose. Any new information derived from these studies will also be highly relevant to disease states affecting odorant detection by the nose.

描述（由申请人提供）： 项目摘要该提案的长期目标是在定量详细了解脊椎动物嗅觉受体神经元（ORNS）中的转导机制。在当前赠款期间，我们已经能够解决由ORN上的单个气味结合事件触发的响应（所谓的“单一响应”）。令人惊讶的是，我们发现随机遇到的ORN（因此随机遇到的气味受体或ORS）都给出了类似振幅的单一响应，这表明一个气味结合事件的下游信号传导的可能性很低。换句话说，单一响应似乎反映了单个激活的高尔夫/腺苷酸环化酶复合物的作用。这种现象显然是由于在或分子上存在非常短的气味停留时间（即，气味与OR的快速分离）。这种短气味的住所同样主导了嗅觉转导的终止。因此，传统上认为是终止G蛋白偶联受体（GPCR）信号传导的决定因素，即受体磷酸化和随后的抑制蛋白结合，至少在单位反应的水平上并不重要，尽管它们可能仍然具有强烈的强度和延长的ODORNONS刺激。到目前为止，已从两栖动物ORN和低CA2+溶液中获得了上述结果（为了提高单一响应幅度）。在此应用程序中，我们建议继续使用两栖动物ORN进行实验，但同时也使用鼠标ORN，这为遗传操作提供了明显的优势。 Aim 1 is to obtain the best estimate of the olfactory unitary-response amplitude in frog ORNs in physiological conditions (i.e., normal external Ca2+), to characterize its spatial spread along the olfactory cilium, to dissect its membrane-current components (cyclic-nucleotide-gated current versus Cl current), and to estimate the number of unitary responses required for bringing the ORN to firing 临界点。 AIM 2是对小鼠ORN中的单一响应进行类似的详细研究，这将是检查可用的遗传小鼠线以进行嗅觉转导的基础。 AIM 3是通过使用遗传小鼠系来详细研究/剖析ORN的CA依赖性和与CA非依赖性的适应。 AIM 4是在没有气味的情况下研究构成或活性，到目前为止，这种气味很少关注脊椎动物嗅觉转导。在先前的实验过程中，我们注意到了这种现象。实验方法将主要涉及从青蛙和WT或遗传工程小鼠的单个ORN的吸入式录音。公共卫生相关性：该应用程序中提出的研究将增强我们对鼻子脊椎动物嗅觉受体神经元嗅觉转导的理解。从这些研究中得出的任何新信息也将与影响鼻子的气味检测的疾病状态高度相关。