The molecular mechanisms of aminergic release at “en-passent” type II synaptic terminals of the fruit fly (Drosophila melanogaster).

这项提议将对驱动I型和II型胺能突触终末之间相互作用的结构和功能可塑性产生深刻的见解。我们特别试图了解从II型终末发出的神经调制如何影响I型终末的快速突触囊泡释放，也称为神经肌肉连接(NMJ)。由于它们的外围位置，这些突触很容易接触到，因此，在较难获得的大脑中，突触的调节是学习过程的主要手段，因此，这些突触是一个理想的例子。在果蝇幼虫中，神经调节神经元本身的突触被称为II型突触终末，与被称为I型终末(或NMJ，神经肌肉连接)的运动神经元突触相比，研究要少得多。特别是，由酪胺能/八胺能不成对的正中神经元轴突形成的II型突触终末的胞吐机制是我们实验的重点。这些II型终末形成“传入型”突触，含有致密的核心小泡，并调节NMJ的传递效果。然而，这些II型终末是否使用支架蛋白来形成活动区类型的细胞毒作用尚不清楚。这个问题将通过结合免疫细胞化学、超微结构分析(包括免疫金标记法)以及更重要的是超分辨率光学显微镜，按照西格里斯特实验室建立的良好方案进行严格探索。作为更深入理解的一个重要方面，将寻求研究I型和II型终末之间的发育相互作用，特别是在变态期间，迄今仍是一种未知的陆地。特别是，运动和神经调节神经元及其终末在变态过程中的相互作用是否必要，以便在成年肌肉中建立全功能的“适当的”NMJ和“传入性”突触(II型终末)，是这里关注的焦点。在整个过程中，我们将使用功能重要性的行为读数：(I)幼虫爬行和(Ii)成虫静止飞行。

The proposal will yield insights into the structural and functional plasticity driving the interplay between type I and aminergic type II synaptic terminals. We particularly seek to understand how neuromodulation emanating from type II terminals impacts on the fast synaptic vesicle release of type I terminals also known as neuromuscular junctions (NMJ).Due to their peripheral location, these synapses are easily accessible and, thus, serve as an ideal example for similar interactions in the less accessible brain where modulation of synapses is a major means for learning processes. Synapses of the neuromodulatory neurons themselves, in Drosophila larvae known as type II synaptic terminals are much less studied compared to synapses of motor neurons called type I terminals (or NMJ, neuromuscular junction). In particular, the mechanism of exocytosis of the type II synaptic terminals formed by the axons of tyraminergic/octopaminergic unpaired median neurons are in the focus of our experiments. These type II terminals form synapses of the “en-passent”-type, contain dense core vesicles and modulate the efficacy of transmission by NMJs. Whether these type II-terminals use scaffold-proteins to form active zone-type cytomatrices remains unknown, however. This question will be rigorously explored by a combination of immunocytochemistry, ultrastructural analysis including immunogold labeling and, importantly, super resolution light microscopy, following protocols well established in the Sigrist lab. As an essential aspect for deeper understanding, will seek to study the developmental interplay between of type I and II terminals, particularly during metamorphosis, so far a terra incognita. Particularly, whether interactions between motor and neuromodulatory neurons and their terminals during metamorphosis are necessary in order to establish fully functional “proper” NMJs and “en-passant”-synapses (type II terminals) in the adult muscle is in the focus here. Throughout, we shall use a behavioral read out for functional importance: (i) larval crawling and (ii) adult stationary flight.