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

项目摘要/摘要 这个项目的目标是确定气味诱导的钙荧光信号和 在形态上不同类型的昆虫嗅觉感受器神经元(ON)上的棘波反应。百万 每年都有成千上万的生命受到毁灭性的虫媒疾病的威胁--如疟疾、登革热和西尼罗河 热病--通过严重依赖嗅觉寻找宿主的媒介传播。 相应地，越来越多的努力致力于更好地了解昆虫的嗅觉，以开发工具和 干扰寄主寻找行为的策略。随着CRISPR基因组编辑技术的出现， 由于昆虫种类繁多，体内钙成像已成为识别响应的Orn的首选方法 来寄生气味。然而，钙诱导的荧光信号可能很难解释，因为响应 幅度和动态范围不仅取决于受体的激活程度，还取决于 神经元的固有电紧张性，这种特性可以在不同的神经元类型之间有明显的变化。例如，10% 荧光信号增加可对应于一个人的高频、接近饱和的尖峰响应 On类型，但可能代表另一种剂量曲线上升阶段的低尖峰反应。 为了准确地解释钙成像数据，了解峰值与钙的关系是必不可少的。 与单个神经元类型相关。为了实现这一目标，这项提议利用了强大的基因 果蝇的工具包和易驾驭的嗅觉系统。与其他昆虫物种类似，果蝇角是 根据包裹其感觉树突的感受器类型被归类于形态分类。 每个感受器通常含有两到四个感受器，它们表现出独特的细胞外尖峰幅度， 反映了分隔的神经元之间的大小差异。鉴于电子紧张性的性质 感觉神经元受其形态和形态特征的影响，任何这样的差异 角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反应的关系在不同的ORN形态类型中有所不同-将是 同时进行活体跨角质层荧光成像和单感受器电生理检测 正在录音。在这项提议中，目标1将确定钙反应的动态范围和尖峰荧光 代表所有形态分类的选择Orn中的关系。目标2将侧重于比较 分隔化的兽人，通常表现出独特的形态特征。成功执行了 该提案将产生丰富的数据集，以便于对钙诱导的荧光信号进行有意义的解释 记录自果蝇的ORN类型。重要的是，这些信息应该可以推广到其他 昆虫，包括病媒和农业害虫，因为这些ORN形态分类在 昆虫物种。此外，这项系统的调查将为今后确定 感觉神经元钙反应的机械基础。

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.