scholarly journals Synaptotagmin 7 is targeted to the axonal plasma membrane through γ-secretase processing to promote synaptic vesicle docking in mouse hippocampal neurons

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Jason D Vevea ◽  
Grant F Kusick ◽  
Kevin C Courtney ◽  
Erin Chen ◽  
Shigeki Watanabe ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Synaptic Vesicle ◽  
Synaptic Vesicles ◽  
Hippocampal Neurons ◽  
Glutamate Release ◽  
Vesicle Docking ◽  
Presynaptic Function ◽  
Asynchronous Release ◽  
Vesicle Cycle ◽  
Activity Dependent

Synaptotagmin 7 (SYT7) has emerged as a key regulator of presynaptic function, but its localization and precise role in the synaptic vesicle cycle remain the subject of debate. Here, we used iGluSnFR to optically interrogate glutamate release, at the single-bouton level, in SYT7KO-dissociated mouse hippocampal neurons. We analyzed asynchronous release, paired-pulse facilitation, and synaptic vesicle replenishment and found that SYT7 contributes to each of these processes to different degrees. ‘Zap-and-freeze’ electron microscopy revealed that a loss of SYT7 diminishes docking of synaptic vesicles after a stimulus and inhibits the recovery of depleted synaptic vesicles after a stimulus train. SYT7 supports these functions from the axonal plasma membrane, where its localization and stability require both γ-secretase-mediated cleavage and palmitoylation. In summary, SYT7 is a peripheral membrane protein that controls multiple modes of synaptic vesicle (SV) exocytosis and plasticity, in part, through enhancing activity-dependent docking of SVs.

scholarly journals Synaptotagmin 7 is enriched at the plasma membrane through γ-secretase processing to promote vesicle docking and control synaptic plasticity in mouse hippocampal neurons

2021 ◽  
Author(s):  
Jason D. Vevea ◽  
Grant F. Kusick ◽  
Erin Chen ◽  
Kevin C. Courtney ◽  
Shigeki Watanabe ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Synaptic Vesicle ◽  
Synaptic Vesicles ◽  
Hippocampal Neurons ◽  
Synaptic Function ◽  
List Type ◽  
Paired Pulse ◽  
Paired Pulse Facilitation ◽  
Asynchronous Release ◽  
Vesicle Cycle

Abstract Synaptotagmin (SYT) 7 has emerged as key regulator of presynaptic function, but its localization and precise function in the synaptic vesicle cycle remain unclear. Here, we used iGluSnFR to optically and directly interrogate glutamate release, at the single bouton level, in SYT7 KO dissociated mouse hippocampal neurons. We analyzed asynchronous release, paired pulse facilitation, and synaptic vesicle replenishment, and found that SYT7 contributes to each of these processes to different degrees. ‘Zap-and-freeze’ electron microscopy revealed that loss of SYT7 impairs the docking of synaptic vesicles after a stimulus and the recovery of depleted synaptic vesicles after a stimulus train. To execute these functions, SYT7 must be targeted to the plasma membrane via γ-secretase-mediated cleavage of the amino terminus, followed by palmitoylation. The complex sorting itinerary of SYT7 endows this Ca2+-sensor with the ability to control crucial forms of synaptic function and plasticity. SYT7 mediated asynchronous release, paired pulse facilitation, and synaptic vesicle replenishment was observed optically at individual hippocampal synapses Localization, trafficking, and stability of SYT7 is dependent on processing by γ-secretase Short term plasticity defects arise in SYT7KOs due to decreased docking of synaptic vesicles after stimulation SYT7 promotes paired-pulse facilitation and asynchronous release via distinct mechanisms

scholarly journals The synaptic vesicle cycle: a single vesicle budding step involving clathrin and dynamin.

1996 ◽  
Vol 133 (6) ◽  
pp. 1237-1250 ◽  
Author(s):  
K Takei ◽  
O Mundigl ◽  
L Daniell ◽  
P De Camilli
Keyword(s):  
Plasma Membrane ◽  
Synaptic Vesicle ◽  
Synaptic Vesicles ◽  
Hippocampal Neurons ◽  
Coated Vesicles ◽  
Coated Vesicle ◽  
Nerve Terminals ◽  
Vesicle Budding ◽  
Vesicle Cycle ◽  
Single Vesicle

Strong evidence implicates clathrin-coated vesicles and endosome-like vacuoles in the reformation of synaptic vesicles after exocytosis, and it is generally assumed that these vacuoles represent a traffic station downstream from clathrin-coated vesicles. To gain insight into the mechanisms of synaptic vesicle budding from endosome-like intermediates, lysed nerve terminals and nerve terminal membrane subfractions were examined by EM after incubations with GTP gamma S. Numerous clathrin-coated budding intermediates that were positive for AP2 and AP180 immunoreactivity and often collared by a dynamin ring were seen. These were present not only on the plasma membrane (Takei, K., P.S. McPherson, S.L.Schmid, and P. De Camilli. 1995. Nature (Lond.). 374:186-190), but also on internal vacuoles. The lumen of these vacuoles retained extracellular tracers and was therefore functionally segregated from the extracellular medium, although narrow connections between their membranes and the plasmalemma were sometimes visible by serial sectioning. Similar observations were made in intact cultured hippocampal neurons exposed to high K+ stimulation. Coated vesicle buds were generally in the same size range of synaptic vesicles and positive for the synaptic vesicle protein synaptotagmin. Based on these results, we suggest that endosome-like intermediates of nerve terminals originate by bulk uptake of the plasma membrane and that clathrin- and dynamin-mediated budding takes place in parallel from the plasmalemma and from these internal membranes. We propose a synaptic vesicle recycling model that involves a single vesicle budding step mediated by clathrin and dynamin.

scholarly journals Synaptophysin I Controls the Targeting of VAMP2/Synaptobrevin II to Synaptic Vesicles

2003 ◽  
Vol 14 (12) ◽  
pp. 4909-4919 ◽  
Author(s):  
Maria Pennuto ◽  
Dario Bonanomi ◽  
Fabio Benfenati ◽  
Flavia Valtorta
Keyword(s):  
Plasma Membrane ◽  
Membrane Protein ◽  
Synaptic Vesicle ◽  
Cell Body ◽  
Synaptic Vesicles ◽  
Hippocampal Neurons ◽  
Direct Interaction ◽  
Diffuse Component ◽  
Synaptotagmin I ◽  
Dose Dependent

Synaptic vesicle (SV) proteins are synthesized at the level of the cell body and transported down the axon in membrane precursors of SVs. To investigate the mechanisms underlying sorting of proteins to SVs, fluorescent chimeras of vesicle-associated membrane protein (VAMP) 2, its highly homologous isoform VAMP1 and synaptotagmin I (SytI) were expressed in hippocampal neurons in culture. Interestingly, the proteins displayed a diffuse component of distribution along the axon. In addition, VAMP2 was found to travel in vesicles that constitutively fuse with the plasma membrane. Coexpression of VAMP2 with synaptophysin I (SypI), a major resident of SVs, restored the correct sorting of VAMP2 to SVs. The effect of SypI on VAMP2 sorting was dose dependent, being reversed by increasing VAMP2 expression levels, and highly specific, because the sorting of the SV proteins VAMP1 and SytI was not affected by SypI. The cytoplasmic domain of VAMP2 was found to be necessary for both the formation of VAMP2-SypI hetero-dimers and for VAMP2 sorting to SVs. These data support a role for SypI in directing the correct sorting of VAMP2 in neurons and demonstrate that a direct interaction between the two proteins is required for SypI in order to exert its effect.

scholarly journals How do you recognize and reconstitute a synaptic vesicle after fusion?

F1000Research ◽  
2017 ◽  
Vol 6 ◽  
pp. 1734 ◽  
Author(s):  
Natali L. Chanaday ◽  
Ege T. Kavalali
Keyword(s):  
Plasma Membrane ◽  
Synaptic Vesicle ◽  
Synaptic Vesicles ◽  
Membrane Composition ◽  
Synaptic Vesicle Recycling ◽  
Vesicle Recycling ◽  
Molecular Identity ◽  
Daunting Task ◽  
Vesicle Biogenesis ◽  
Vesicle Cycle

Synaptic vesicle recycling is essential for sustained and reliable neurotransmission. A key component of synaptic vesicle recycling is the synaptic vesicle biogenesis process that is observed in synapses and that maintains the molecular identity of synaptic vesicles. However, the mechanisms by which synaptic vesicles are retrieved and reconstituted after fusion remain unclear. The complex molecular composition of synaptic vesicles renders their rapid biogenesis a daunting task. Therefore, in this context, kiss-and-run type transient fusion of synaptic vesicles with the plasma membrane without loss of their membrane composition and molecular identity remains a viable hypothesis that can account for the fidelity of the synaptic vesicle cycle. In this article, we discuss the biological implications of this problem as well as its possible molecular solutions.

scholarly journals BDNF mobilizes synaptic vesicles and enhances synapse formation by disrupting cadherin–β-catenin interactions

2006 ◽  
Vol 174 (2) ◽  
pp. 289-299 ◽  
Author(s):  
Shernaz X. Bamji ◽  
Beatriz Rico ◽  
Nikole Kimes ◽  
Louis F. Reichardt
Keyword(s):  
Synaptic Vesicle ◽  
Synaptic Vesicles ◽  
Hippocampal Neurons ◽  
Molecular Mechanisms ◽  
Synapse Formation ◽  
Time Lapse ◽  
Synapse Density ◽  
Vesicle Mobility ◽  
Small Clusters ◽  
Vertebrate Central Nervous System

Neurons of the vertebrate central nervous system have the capacity to modify synapse number, morphology, and efficacy in response to activity. Some of these functions can be attributed to activity-induced synthesis and secretion of the neurotrophin brain-derived neurotrophic factor (BDNF); however, the molecular mechanisms by which BDNF mediates these events are still not well understood. Using time-lapse confocal analysis, we show that BDNF mobilizes synaptic vesicles at existing synapses, resulting in small clusters of synaptic vesicles “splitting” away from synaptic sites. We demonstrate that BDNF's ability to mobilize synaptic vesicle clusters depends on the dissociation of cadherin–β-catenin adhesion complexes that occurs after tyrosine phosphorylation of β-catenin. Artificially maintaining cadherin–β-catenin complexes in the presence of BDNF abolishes the BDNF-mediated enhancement of synaptic vesicle mobility, as well as the longer-term BDNF-mediated increase in synapse number. Together, this data demonstrates that the disruption of cadherin–β-catenin complexes is an important molecular event through which BDNF increases synapse density in cultured hippocampal neurons.

scholarly journals Counting the Number of Glutamate Molecules in Single Synaptic Vesicles

2019 ◽  
Author(s):  
Yuanmo Wang ◽  
Hoda fathali ◽  
devesh mishra ◽  
Thomas Olsson ◽  
Jacqueline Keighron ◽  
...  
Keyword(s):  
Synaptic Vesicle ◽  
Synaptic Vesicles ◽  
High Speed ◽  
Glutamate Release ◽  
Dependent Manner ◽  
Excitatory Neurotransmitter ◽  
Quantitative Measurements ◽  
Sensor Technique ◽  
Drug Treatments ◽  
Analytical Tools

<div><p>Analytical tools for direct quantitative measurements of glutamate, the principal excitatory neurotransmitter in brain, are lacking. Here, we introduce a new enzyme-based amperometric sensor technique for direct counting of the number of glutamate molecules stored inside single synaptic vesicles. An ultra-fast enzyme-based glutamate sensor is placed into a solution of isolated synaptic vesicles, which stochastically rupture at the sensor surface in a potential dependent manner by applying a constant negative potential. High-speed (10 kHz) amperometry is used to record sub-millisecond current spikes, which represent glutamate release from single vesicles that burst open. Glutamate quantification is achieved by a calibration curve that is based on measurements of glutamate release from vesicles pre-filled with various concentrations of glutamate. Our measurements show that a single synaptic vesicle encapsulates about 8000 glutamate molecules, which is comparable to the measured exocytotic quantal glutamate release in the nucleus accumbens of mouse brain tissue. Hence, this new methodology introduces the means to quantify ultra-small amounts of glutamate and to study synaptic vesicle physiology, pathogenesis and drug treatments for neuronal disorders where glutamate is involved.</p></div>

Recent Breakthroughs in Neurotransmitter Release: Paradigm for Regulated Exocytosis?

Physiology ◽  
1995 ◽  
Vol 10 (1) ◽  
pp. 42-46
Author(s):  
G Thiel
Keyword(s):  
Synaptic Vesicle ◽  
Brain Function ◽  
Neurotransmitter Release ◽  
Synaptic Vesicles ◽  
Vesicle Trafficking ◽  
Regulated Exocytosis ◽  
Intracellular Vesicle ◽  
Vesicle Cycle ◽  
Synaptic Vesicle Cycle

Synaptic vesicles play a fundamental role in brain function by mediating the release of neurotransmitters. Neurons do not use an entirely unique secretion apparatus but rather a modification of the general secretion machinery. Moreover, the synaptic vesicle cycle has many similarities with intracellular vesicle trafficking pathways.

Neurotransmitter Release

2017 ◽  
Author(s):  
Peggy Mason
Keyword(s):  
Plasma Membrane ◽  
Synaptic Vesicle ◽  
Active Zone ◽  
Neurotransmitter Release ◽  
Synaptic Vesicles ◽  
Synaptic Cleft ◽  
Fusion Pore ◽  
Synaptic Membrane ◽  
Specialized Region ◽  
Synaptic Vesicle Fusion

The biochemical and physiological processes of neurotransmitter release from an active zone, a specialized region of synaptic membrane, are examined. Synaptic vesicles containing neurotransmitters are docked at the active zone and then primed for release by SNARE complexes that bring them into extreme proximity to the plasma membrane. Entry of calcium ions through voltage-gated calcium channels triggers synaptic vesicle fusion with the synaptic terminal membrane and the consequent diffusion of neurotransmitter into the synaptic cleft. Release results when the fusion pore bridging the synaptic vesicle and plasma membrane widens and neurotransmitter from the inside of the synaptic vesicle diffuses into the synaptic cleft. Membrane from the active zone membrane is endocytosed, and synaptic vesicle proteins are then reassembled into recycled synaptic vesicles, allowing for more rounds of neurotransmitter release.

Optical Mesurements of Presynaptic Function: What Keeps Vesicle Traffic Going

1998 ◽  
Vol 4 (S2) ◽  
pp. 1020-1021
Author(s):  
Timothy A. Ryan
Keyword(s):  
Nervous System ◽  
Synaptic Vesicles ◽  
Hippocampal Neurons ◽  
Brain Cells ◽  
Membrane Recycling ◽  
Large Collection ◽  
Vesicle Traffic ◽  
Presynaptic Function ◽  
Average Total Number ◽  
Chemical Synapses

The nervous system has evolved to make use of a variety of mechanisms that allow information to flow and be processed among a large collection of individual cells. The communication between individual brain cells occurs largely at chemical synapses. In these compartments, chemical messengers are packaged into small vesicles that fuse with the cell membrane upon stimulation, releasing neurotransmitter.. The average total number of synaptic vesicles in a typical central nervous system synapse is only a few hundred and as a result an efficient local recycling mechanism operates in order to replenish this pool during periods of even modest neuronal activity. Without this membrane recycling, synapses quickly become depleted of vesicles, and soon fail to communicate information between cells.We make use of optical techniques to follow the trafficking of synaptic vesicles at synapses formed between hippocampal neurons grown in culture. Recycling synaptic vesicles can be readily labeled using the fluorescent amphipathic membrane dye FM 1-43.

Dynamin Mediates Membrane Vesiculation

1998 ◽  
Vol 4 (S2) ◽  
pp. 1022-1023
Author(s):  
Sharon M. Sweitzer ◽  
Jenny E. Hinshaw
Keyword(s):  
Drosophila Melanogaster ◽  
Plasma Membrane ◽  
Synaptic Vesicle ◽  
Mechanism Of Action ◽  
Synaptic Vesicles ◽  
Dominant Negative ◽  
Coated Pits ◽  
Vesicle Recycling ◽  
Membrane Vesiculation ◽  
Helical Tubes

Dynamin, a 100 kDa GTPase, is essential for receptor mediated endocytosis and synaptic vesicle recycling; however its mechanism of action is unknown. The requirement for dynamin was first elucidated by the discovery that the shibire gene product in Drosophila melanogaster was homologous to mammalian dynamin-1 (1,2). The shibire flies exhibit a depletion of synaptic vesicles and an accumulation of collared clathrin-coated pits at the plasma membrane of their nerve termini (3). It was later demonstrated that endocytosis was inhibited by the overexpression of dominant negative mutants of dynamin (4,5), and that purified dynamin can self-associate to form spirals which resemble the collars of shibire and structures seen in synaptosomes treated with GTPγS (6,7). These observations led to the speculation that dynamin pinches the clathrin-coated bud from the plasma membrane. In support of this hypothesis, we show that purified recombinant dynamin can bind to a lipid bilayer in a regular and repeating pattern to form helical tubes which vesiculate upon the addition of GTP.

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