Analysis of PDF-dependent microcircuits in the circadian clock of the Madeira cockroach

神经肽色素分散因子（PDF）是昆虫体内一种专性的昼夜节律偶联信号，它调节分布在昆虫体内的生物钟。它与哺乳动物中最重要的昼夜节律偶联因子血管活性肠肽（VIP）在分子和细胞水平上的信号传导有着惊人的相似性。PDF和VIP信号通过cAMP升高，并且对于与外部光暗周期同步的生理和行为中的大幅度昼夜节律的表达是必不可少的。尽管如此，我们的知识PDF/VIP依赖的神经元回路和他们的控制，通过light. We更新的建议进一步集中在细胞分析的PDF依赖的昼夜节律电路的时钟的夜间活跃，长寿马德拉蟑螂，一个既定的细胞模型的昼夜节律研究。细胞内和膜片钳电生理分析结合钙离子成像，多标记免疫细胞化学，和单细胞MALDI-TOF我们研究光输入途径PDF神经元。以前的PDF注射阶段延迟的运动活动节律在黄昏和阶段提前在黎明期间cAMP和光刺激。此外，在蟑螂的昼夜节律钟中测量cAMP浓度的峰值，一个在黄昏，一个在黎明。因此，我们提出并寻找两个PDF依赖的神经元电路在蟑螂时钟锁定黄昏或黎明限制运动活动的节奏，而光周期在一年中的过程中不断变化的夜晚。

The neuropeptide pigment-dispersing factor (PDF) is an obligatory circadian coupling signal in insects which synchronizes circadian clocks distributed throughout the insect body. It shares astounding similarities in signaling on the molecular and cellular level with vasoactive intestinal polypeptide (VIP) the most important circadian coupling factor in mammals. Both, PDF and VIP signal via cAMP rises and are essential for the expression of large amplitude circadian rhythms in physiology and behavior in synchrony with external light-dark cycles. Nevertheless, our knowledge about the PDF/VIP-dependent neuronal circuits and their control via light is still limited.Our renewal proposal focuses further on cellular analysis of PDF-dependent circadian circuits in the clock of the night-active, long-lived Madeira cockroach, an established cellular model of circadian rhythm research. With intracellular and patch clamp electrophysiological analysis combined with Ca2+ imaging, multiple-label immunocytochemistry, and single-cell MALDI-TOF we examine the light input pathways to PDF neurons. Previous PDF-injections phase delayed locomotor activity rhythms during dusk and phase advanced during dawn comparably to cAMP- and light-stimuli. Furthermore, peaks in cAMP concentrations were measured in the cockroach circadian clock one at dusk and one at dawn. Thus, we propose and search for two PDF-dependent neuronal circuits in the cockroach clock locking either onto dusk or dawn to restrict locomotor activity rhythms to the night while photoperiod is changing during the course of the year.