Analysis of olfactory transduction mechanisms in the behaving hawkmoth Manduca sexta

昆虫气味的传导和编码仍存在争议。在昆虫嗅觉受体神经元（ORN）中，普遍存在的Orco受体被认为与嗅觉受体（ORs）多聚化形成气味门控离子通道。这种离子型机制的功能影响正在争论中。在天蛾Manduca sexta，我们提出了一个新的假设，这表明，旁边的代谢级联泄漏OR-Orco离子通道的时间编码信息素刺激。他们建议产生阈下膜电位振荡，控制自发活动。从而确定了气味传递的阈值和时间过程。我们假设，Orco功能作为一个依赖于昆虫的起搏器电流，调节细胞内Ca 2+和环核苷酸浓度的变化依赖于昆虫的行为状态。在这里，我们研究信息素依赖的转导级联在天蛾在不同的行为状态。重点是挑战我们的新假设的MsexOrco在时间编码的信息素的作用。与膜片钳研究，药理学，和Ca 2+成像在异源表达系统，以及与原代细胞培养的ORN在体外，我们研究各自的贡献离子型机制和代谢级联。用酶联免疫吸附试验和尖端记录法对黑腹小蠊的完整感器进行了研究。sexta在原位，我们进一步挑战我们的新假设，并分析是否依赖于不同的行为背景和生理状态是基于环核苷酸水平的昼夜变化。

Insect odor transduction and encoding is still under debate. In insect olfactory receptor neurons (ORNs) an ubiquitous Orco receptor is suggested to multimerize with olfactory receptors (ORs) to form an odor-gated ion channel. The functional implications of this ionotropic mechanism are under debate. In the hawkmoth Manduca sexta we propose a new hypothesis which suggests that next to metabotropic cascades leaky OR-Orco ion channels are employed in temporal encoding of pheromone stimuli. They are suggested to generate subthreshold membrane potential oscillations which control spontaneous activity. Thereby, threshold and time course of odor transduction are determined. We hypothesize that Orco functions as a hormone-dependent pacemaker current, regulated via changes in the intracellular Ca2+ and cyclic nucleotide concentrations dependent on the behavioral state of the insect. Here, we examine pheromone-dependent transduction cascades in the hawkmoth during different behavioral states. Focus is to challenge our new hypothesis of the role of MsexOrco in temporal encoding of pheromones. With patch clamp studies, pharmacology, and Ca2+-imaging in a heterologous expression system as well as with primary cell cultures of ORNs in vitro we examine the respective contribution of ionotropic mechanisms and metabotropic cascades. With ELISAs and tip recordings of intact sensilla of M. sexta in situ we further challenge our new hypothesis and analyze whether dependence on different behavioral contexts and physiological states is based upon circadian changes in cyclic nucleotide levels.