The relationship between spike response and calcium fluorescent signal in insect olfactory receptor neurons

项目概要/摘要 本项目的目标是确定气味诱导的钙荧光信号之间的关系， 在形态上不同类型的昆虫嗅觉受体神经元（ORN）的棘波反应。数百万 每年都有2000万人的生命受到毁灭性虫媒疾病的威胁，如疟疾、登革热和西尼罗河病毒 发烧-这是由严重依赖嗅觉寻找宿主的病媒传播的。 相应地，越来越多的努力致力于更好地理解昆虫嗅觉，以开发工具和工具。 策略来干扰宿主寻找行为。随着CRISPR基因组编辑技术的出现， 广泛的昆虫，在体内钙成像已成为一种选择的方法，用于确定ORN的反应， 来承载气味然而，钙诱导的荧光信号可能难以解释，因为响应 振幅和动态范围不仅由受体激活的程度决定，而且还由 神经元的内在电紧张特性，其可在神经元类型之间显著变化。例如，10% 荧光信号增加可对应于高频、接近饱和的尖峰响应 ORN类型，但可以替代地表示在另一种类型的剂量曲线的上升阶段处的低尖峰响应。 为了准确地解释钙成像数据，必须了解尖峰-钙关系 与单个神经元类型相关。为了实现这一目标，该提案利用了强大的遗传 工具包和易于处理的果蝇嗅觉系统。与其他昆虫物种相似，果蝇的ORN是 根据封装它们的感觉树突的感觉器类型分类。 每个感器通常包含两到四个ORN，它们表现出独特的细胞外尖峰幅度， 反映了分隔神经元之间的大小差异。考虑到电紧张性， 感觉神经元受其形态学和形态测量学特征的影响， ORN的形态类型可能会影响它们的穗钙关系。一般的假设是， 气味诱导的尖峰和钙响应关系在不同的ORN形态类型中有所不同， 通过同时在体内跨表皮荧光成像和单感器电生理测试 录制.在该提案中，目标1将确定钙响应动态范围和尖峰荧光 代表所有形态学类别的选择ORN中的关系。目标2将侧重于比较 典型地表现出独特的形态特征的区室化的ORN。成功执行 该提案将产生丰富的数据集，以促进对钙诱导荧光信号的有意义的解释 果蝇ORN类型的记录。重要的是，这一信息预计将推广到其他 昆虫，包括病媒和农业害虫，因为这些ORN形态类别在世界各地都有发现。 昆虫种类此外，这一系统的调查将奠定基础，为今后的研究，以确定 感觉神经元钙反应的机制基础。

PROJECT SUMMARY/ABSTRACT The goal of this project is to determine the relationships between odor-induced calcium fluorescence signals and spike responses across morphologically distinctive types of insect olfactory receptor neurons (ORNs). Millions of lives are threatened annually by devastating insect-borne diseases—such as malaria, dengue, and West Nile fever—which are transmitted by vectors that heavily rely on their sense of smell for host-seeking. Correspondingly, growing efforts are dedicated to better understanding insect olfaction to develop tools and strategies to interfere with host-seeking behavior. With the advent of CRISPR genome editing technology in a wide range of insects, in vivo calcium imaging has become a method of choice for identifying ORNs that respond to host odors. However, calcium-induced fluorescence signals may be difficult to interpret, because response amplitudes and dynamic ranges are determined not only by the degree of receptor activation, but also by the intrinsic electrotonic property of neurons which can vary markedly across neuronal types. For example, a 10% increase in fluorescence signal may correspond to a high-frequency, near-saturating spike response for one ORN type, but may instead represent a low spike response at the rising phase of the dosage curve for another. In order to accurately interpret calcium imaging data, it is imperative to understand the spike-calcium relationship associated with individual neuronal types. To achieve this goal, this proposal leverages the powerful genetic toolkit and tractable olfactory system of Drosophila. Similar to other insect species, Drosophila ORNs are categorized under morphological classes based on the sensillum types that encapsulate their sensory dendrites. Each sensillum typically contains two to four ORNs, which exhibit distinctive extracellular spike amplitudes that reflect the size differences between compartmentalized neurons. Given that the electrotonic properties of sensory neurons are influenced by their morphological and morphometric features, any such differences across ORN morphological types will likely impact their spike-calcium relationship. The general hypothesis—that the odor-induced spike and calcium response relationship vary across distinctive ORN morphological types—will be tested via simultaneous in vivo trans-cuticle fluorescence imaging and single-sensillum electrophysiological recording. In this proposal, Aim 1 will determine the calcium response dynamic range and spike-fluorescence relationship in select ORNs representing all morphological classes. Aim 2 will focus on the comparison between compartmentalized ORNs which typically exhibit distinctive morphometric features. Successful execution of the proposal will yield a rich dataset to facilitate meaningful interpretation of calcium-induced fluorescence signals recorded from Drosophila ORN types. Importantly, this information is expected to be generalizable to other insects, including disease vectors and agricultural pests, as these ORN morphological classes are found across insect species. Moreover, this systematic survey will lay the foundation for future studies to determine the mechanistic underpinnings of sensory neurons’ calcium responses.