DELIVERY AND EXOCYTOSIS OF NEUROPEPTIDE VESICLES AT THE NERVE TERMINAL

Neuropeptides, which control emotion, behavior and body homeostasis, are released at nerve terminals through the exocytosis of dense-core vesicles (DCVs). Because neuropeptides are synthesized and packaged exclusively at the cell body, accurate and timely delivery of DCV to the distant nerve terminal, as well as tight regulation of neuropeptide release should be ensured to prevent the deprivation of releasable DCVs during activity. Traditionally, it was believed that DCVs are delivered to the nerve terminal exclusively through anterograde fast axonal transport. However, it is now known that DCVs continuously transit bidirectionally at nerve terminals that are highly branched with release sites located along strings of varicosities called en passant boutons. Therefore, it is not clear how DCV logistics operates to support neurosecretion. In addition, little is known about the release properties of neuropeptides in fully differentiated neurons.\ud To address these issues, GFP-tagged neuropeptides were imaged in Drosophila melanogaster neuromuscular junction (NMJ) to study the routing and exocytosis behavior of DCVs in vivo. By using a newly developed optical technique, simultaneous photobleaching and imaging (SPAIM), to track single DCVs, it was found that DCVs circulate between the most distal bouton and the proximal axon. Neuropeptides are constantly delivered to en passant boutons via sporadic bidirectional capture of DCVs. These newly arrived DCVs can become releasable within minutes. Surprisingly, neuropeptides are discharged only partially and slowly from DCVs, suggesting that “kiss-and-run” mode of exocytosis predominates over full fusion. Interestingly, these partially emptied DCVs can undergo multiple rounds of exocytosis, allowing some local recycling of DCVs and minimizing the need for the supply of new DCVs. Finally, it was found that DCVs are reloaded into en passant boutons after depolarization evoked-neuropeptide release by activity-dependent capture that is regulated through the ryanodine receptor-CamKII signaling pathway. \ud Altogether, this thesis describes some elegant mechanisms utilized by the neuron that efficiently maintain neuropeptide stores at the distant and morphologically complex nerve terminal. \u