Synaptotagmins ensure speed and efficiency of inhibitory neurotransmitter release Dissertation Thesis

Author(s): Chen, Chong
Advisor(s): Jonas, Peter
Committee Chair(s): Danzl, Johann
Committee Member(s): Shigemoto, Ryuichi
Title: Synaptotagmins ensure speed and efficiency of inhibitory neurotransmitter release
Affiliation IST Austria
Abstract: Neuronal networks in the brain consist of two main types of neuron, glutamatergic principal neurons and GABAergic interneurons. Although these interneurons only represent 10–20% of the whole population, they mediate feedback and feedforward inhibition and are involved in the generation of high-frequency network oscillations. A hallmark functional property of GABAergic interneurons, especially of the parvalbumin‑expressing (PV+) subtypes, is the speed of signaling at their output synapse across species and brain regions. Several molecular and subcellular factors may underlie the submillisecond signaling at GABAergic synapses. Such as the selective use of P/Q type Ca2+ channels and the tight coupling between Ca2+ channels and Ca2+ sensors of exocytosis. However, whether the molecular identity of the release sensor contributes to these signaling properties remains unclear. Besides, these interneurons are mainly show depression in response to train of stimuli. How could they keep sufficient release to control the activity of postsynaptic principal neurons during high network activity, is largely elusive. For my Ph.D. work, we firstly examined the Ca2+ sensor of exocytosis at the GABAergic basket cell (BC) to Purkinje cell (PC) synapse in the cerebellum. Immunolabeling suggested that BC terminals selectively expressed synaptotagmin 2 (Syt2), whereas synaptotagmin 1 (Syt1) was enriched in excitatory terminals. Genetic elimination of Syt2 reduced action potential-evoked release to ~10% compared to the wild-type control, identifying Syt2 as the major Ca2+ sensor at BC‑PC synapses. Differential adenovirus-mediated rescue revealed Syt2 triggered release with shorter latency and higher temporal precision, and mediated faster vesicle pool replenishment than Syt1. Furthermore, deletion of Syt2 severely reduced and delayed disynaptic inhibition following parallel fiber stimulation. Thus, the selective use of Syt2 as the release sensor at BC–PC synapse ensures fast feedforward inhibition in cerebellar microcircuits. Additionally, we tested the function of another synaptotagmin member, Syt7, for inhibitory synaptic transmission at the BC–PC synapse. Syt7 is thought to be a Ca2+ sensor that mediates asynchronous transmitter release and facilitation at synapses. However, it is strongly expressed in fast-spiking, PV+ GABAergic interneurons and the output synapses of these neurons produce only minimal asynchronous release and show depression rather than facilitation. How could Syt7, a facilitation sensor, contribute to the depressed inhibitory synaptic transmission needs to be further investigated and understood. Our results indicated that at the BC–PC synapse, Syt7 contributes to asynchronous release, pool replenishment and facilitation. In combination, these three effects ensure efficient transmitter release during high‑frequency activity and guarantee frequency independence of inhibition. Taken together, our results confirmed that Syt2, which has the fastest kinetic properties among all synaptotagmin members, is mainly used by the inhibitory BC‑PC synapse for synaptic transmission, contributing to the speed and temporal precision of transmitter release. Furthermore, we showed that Syt7, another highly expressed synaptotagmin member in the output synapses of cerebellar BCs, is used for ensuring efficient inhibitor synaptic transmission during high activity.
Publication Title: IST Dissertation
Degree Granting Institution: IST Austria  
Degree: PhD
Degree Date: 2018-03-01
Start Page: 1
Total Pages: 110
Copyright Statement: CC BY
DOI: 10.15479/AT:ISTA:th_997
Notes: I would like to express my sincere gratitude to my supervisor Prof. Peter Jonas. He is a great scientist and a wonderful leader. He is knowledgeable and always gives useful suggestions for the projects, and finds ways to encourages us when we are frustrated by the experiments. It has actually been a great honor to work with him and I feel happy of having the chance to join in his lab. We finished two quite competitive projects together and still have one ongoing project. From Prof. Jonas one can find all the characteristics that a successful scientist must have for doing science, which is critical for us, the young generation. Finally, I want to show my appreciation to all members in Jonas lab. In chronological order: Itaru Arai, who taught me the virus injection and showed me how to do the paired recording. Hua Hu, a great colleague, gave me quite useful suggestions for patching and encouraged me when I was frustrated about patching at the beginning. Caro, a nice labmate, who always discusses literature with me and organizes events for the lab members. Rajiv, a nice and warm-hearted guy. Xiaomin Zhang, with whom I always talk about career. She also shared a lot of food with me. Ben Suter, a quite clever and nice office mate, who is trying to provide help all the time. Magdalena, another office mate, who brings so much fun to us. And also, Florian, Eva, Alois, David, Claudia, Olena ... Besides the lab members, some people at IST Austria also helped me a lot. Ruslan, Stephan, Shamsi, Zuzka, Pepa. Thanks to all of you and it has been a great pleasure to meet all of you here at IST Austria.
Open access: yes (repository)