Sensory control of oxygen-dependent taxis behavior

描述（由申请人提供）：地球上高等生命形式的进化和生存依赖于富氧大气来驱动有效的能量代谢。生理和行为机制已经发展到维持足够的呼吸氧水平，同时限制有毒活性氧的形成。 所有生物体，无论是简单的还是复杂的，监测外部和内部氧气水平的能力是在正常环境条件下生存的关键。在患者可能面临心血管疾病、癌症或呼吸系统疾病等病理状况的临床情况下，这些机制可能会被破坏或被推到极限。基本的氧传感机制是多种多样的，但目前还知之甚少。利用广泛的实验工具，可在模型遗传系统，如C。elegans和Drosophila可以为我们进一步理解氧传感机制做出重要贡献。我们利用遗传学、分子生物学和生理学的方法，将果蝇幼虫的一种基本的趋航行为与外周感觉神经元的一个子集的激活联系起来。初步研究表明，幼虫利用氧敏感机制，以协调必要的食物出口之前，pupariation和最终生存的行为。这些幼虫的定向偏好需要一小部分外周感觉神经元特异性表达DEG/ENaC离子通道亚基Pickpocket 1（PPK 1）的功能。转基因过敏的ppk 1表达神经元导致幼虫的敏感性急剧增加，增加氧气水平。使用破伤风毒素转基因灭活表达ppk 1的神经元会导致对增加的氧水平的厌恶感丧失。我们提出了一个模型，在该模型中，PPK 1表达的神经元可以作为环境中的氧在幼虫阶段的传感器。在节肢动物如果蝇中，氧气通过复杂的气管小管系统扩散到基本上每个细胞和组织。在幼虫觅食阶段，当幼虫表现出最高的厌恶增加氧气水平，幼虫气管系统的外部开口，后气门，是幼虫突出食物表面以上的唯一部分，以允许进入大气。初步结果表明，一个单一的ppk 1表达神经元支配每个后气门。本研究的重点将放在后气门神经元及其在氧感受和趋氧行为中的潜在作用。具体目标我将利用钙敏感的G-CaMP成像技术，检查直接氧依赖性激活的PPK 1表达神经元支配后气门。具体目标II将寻求相关的功能相同的PPK 1表达后气门神经元与幼虫aerotactic行为产生单细胞超敏神经元克隆使用MARCM技术。这些目标的成功完成应建立一个通用的和生产性的遗传模型，用于未来的遗传和分子研究的环境氧传感机制。 公共卫生相关性：所有生物体，无论是简单的还是复杂的，监测外部和内部氧气水平的能力是在正常环境条件下生存的关键。在患者可能面临心血管疾病、癌症或呼吸系统疾病等病理状况的临床情况下，这些机制可能会被破坏或被推到极限。尽管氧气传感在这个星球上的生命生存中发挥着核心作用，但我们对用于平衡氧气供应与代谢需求的各种机制知之甚少。

DESCRIPTION (provided by applicant): The evolution and survival of higher life forms on Earth has relied upon an oxygen-enriched atmosphere to drive efficient energy metabolism. Physiological and behavioral mechanisms have evolved to maintain adequate levels of respiratory oxygen while at the same time limiting toxic reactive oxygen species formation. The ability of all organisms, whether simple or complex, to monitor external and internal oxygen levels is key to survival under normal environmental conditions. In clinical situations where patients may be confronted with pathological conditions such as cardiovascular disease, cancer or respiratory illness, these mechanisms can be disrupted or pushed to their limits. Essential oxygen-sensing mechanisms are diverse and as yet poorly understood. Use of the broad range of experimental tools available in model genetic systems such as C. elegans and Drosophila can contribute significantly to advancing our understanding of mechanisms involved in oxygen-sensing. We have utilized genetic, molecular and physiological approaches to relate a fundamental aerotactic behavior in Drosophila larvae to the activation of a subset of peripheral sensory neurons. Preliminary studies suggest that larvae utilize an oxygen-sensing mechanism to coordinate behaviors necessary for food exit prior to pupariation and ultimate survival. These larval aerotactic preferences require function of a small subset of peripheral sensory neurons specifically expressing the DEG/ENaC ion channel subunit, Pickpocket1 (PPK1). Transgenic hypersensitization of ppk1-expressing neurons causes a dramatic increase in larval sensitivity to increased oxygen levels. Transgenic inactivation of ppk1-expressing neurons using tetanus toxin causes a loss of aversion to increased oxygen levels. We propose a model in which PPK1-expressing neurons may function as sensors of environmental oxygen during larval stages. In arthropods such as Drosophila, oxygen is supplied to essentially every cell and tissue by diffusion of air through a complex tracheal tubule system. During larval foraging stages, when larvae display the highest aversion to increased oxygen levels, the external openings of the larval tracheal system, the posterior spiracles, are the only part of the larva projected above the surface of the food to allow access to the atmosphere. Preliminary results show that a single ppk1-expressing neuron innervates each of the posterior spiracles. The focus of this proposal will be on those posterior spiracle neurons and their potential role in oxygen-sensing and aerotactic behavior. Specific Aim I will utilize Ca2+sensitive G-CaMP imaging techniques to examine direct oxygen-dependent activation of the PPK1-expressing neurons innervating the posterior spiracles. Specific Aim II will seek to correlate the function of the same PPK1-expressing posterior spiracle neurons with larval aerotactic behavior by generating single cell hypersensitized neuronal clones using the MARCM technique. Successful completion of these aims should establish a versatile and productive genetic model for use in future genetic and molecular studies of environmental oxygen sensing mechanisms. PUBLIC HEALTH RELEVANCE: The ability of all organisms, whether simple or complex, to monitor external and internal oxygen levels is key to survival under normal environmental conditions. In clinical situations where patients may be confronted with pathological conditions such as cardiovascular disease, cancer or respiratory illness, these mechanisms can be disrupted or pushed to their limits. Despite the central role of oxygen sensing in the survival of life on this planet, we know relatively little concerning the diverse mechanisms that are utilized to balance oxygen supply with metabolic demand.