Synaptic Vesicles Interchange Their Membrane Proteins with a Large Surface Reservoir during Recycling

Fernández-Alfonso, Tomás; Kwan, Ricky; Ryan, Timothy A.

doi:10.1016/j.neuron.2006.06.008

Cited by 186 publications

(220 citation statements)

References 24 publications

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“…Two recent studies found evidence for the existence of a pool of synaptic vesicle proteins on the plasma membrane that is selectively and readily retrieved after brief or mild stimuli (49,50). Partially preassembled endocytic structures waiting to be recovered after a stimulus could, in principle, account for the rapid endocytosis elicited under some conditions.…”

Section: Discussionmentioning

confidence: 99%

Inhibition of dynamin completely blocks compensatory synaptic vesicle endocytosis

Newton

Kirchhausen

Murthy

2006

Proc. Natl. Acad. Sci. U.S.A.

222

190

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The ability of synapses to sustain signal propagation relies on rapid recycling of transmitter-containing presynaptic vesicles. Clathrinand dynamin-mediated retrieval of vesicular membrane has an undisputed role in synaptic vesicle recycling. There is also evidence for other modes of vesicle retrieval, including bulk retrieval and the so-called kiss-and-run recycling. Whether dynamin in required for these other modes of synaptic vesicle endocytosis remains unclear. Here, we have tested the role of dynamin in synaptic vesicle endocytosis by using a small molecule called dynasore, which rapidly inhibits the GTPase activity of dynamin with high specificity. Endocytosis after sustained or brief stimuli was completely and reversibly blocked by dynasore in cultured hippocampal neurons expressing the fluorescent tracer synaptopHluorin. By contrast, dynasore had no effect on exocytosis. In the presence of dynasore, low-frequency stimulation led to sustained accumulation of synaptopHluorin and other vesicular proteins on the surface membrane at a rate predicted from net exocytosis. These vesicular components remained on surface membranes even after the stimulus was terminated, suggesting that all endocytic events rely on dynamin during low-frequency activity as well as in the period after it. Ultrastructural analysis revealed a reduction in the density of synaptic vesicles and the presence of endocytic structures only at synapses that were stimulated in the presence of dynasore. In sum, our data indicate that dynamin is essential for all forms of compensatory synaptic vesicle endocytosis including any kiss-andrun events.clathrin ͉ hippocampal ͉ vesicle recycling ͉ kiss-and-run D espite substantial progress in understanding the mechanistic basis of the synaptic vesicle cycle, several fundamental questions remain. Evidence for a ''classical'' pathway of vesicle recycling through clathrin-mediated endocytosis comes from a variety of studies and techniques, and many molecular players in this process have been identified (1-4). There is also evidence for other modes of vesicle retrieval, including bulk retrieval (5, 6) and the so-called kiss-and-run recycling (7-12). Mechanistic studies of kiss-and-run recycling have been difficult, in part, because of a lack of agreement on the exact definition of this process.The final mechanical step of the separation of a budding clathrincoated vesicle from the plasma membrane relies on the GTPase dynamin (13,14). Studies using a temperature-sensitive dynamin ortholog shibire in Drosophila have indicated that dynamin function is critical for synaptic vesicle endocytosis (15, 16). If and how dynamin is involved in alternate modes of vesicle recycling such as kiss-and-run is unclear (17-22), especially at small central synapses where these modes have been proposed (8,12,23,24). At several giant synapses, the role of dynamin in synaptic vesicle endocytosis has been assayed by using nonhydrolyzable analogs of GTP such as guanosine 5Ј-[␥-thio]triphosphate (GTP␥S) or peptides that interfere wit...

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Section: Discussionmentioning

confidence: 99%

Inhibition of dynamin completely blocks compensatory synaptic vesicle endocytosis

Newton

Kirchhausen

Murthy

2006

Proc. Natl. Acad. Sci. U.S.A.

222

190

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“…These proteins are mobile and intermix with newly exocytosed synaptic vesicle proteins 44,45 . By contrast, super-resolution stimulated emission depletion (STED) microscopy (BOX 4) has suggested that newly exocytosed synaptic vesicle proteins are clustered at the plasma membrane 46 .…”

Section: Box 1 | Presynaptic Organization -Exocytic and Endocytic Zonesmentioning

confidence: 99%

“…This apparent contradiction could be explained by differences between pHluorin-tagged synaptic vesicle membrane proteins and their native endogenous counterparts 47 . Alternatively, synaptic vesicle proteins may undergo postexocytic reclustering within the periactive zone 44,45 (FIG. 4), a process that could become rate-limiting under conditions of sustained activity (see below).…”

Section: Box 1 | Presynaptic Organization -Exocytic and Endocytic Zonesmentioning

confidence: 99%

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Protein scaffolds in the coupling of synaptic exocytosis and endocytosis

2011

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Communication between nerve cells largely occurs at chemical synapses -specialized sites of cell-cell contact where electrical signals trigger the exocytic release of neurotransmitter, which in turn activates postsynaptic receptor channels. The efficacy with which such chemical signals are transmitted is crucial for the functioning of the nervous system. Modulation of neurotransmission enables nerve cells to respond to a vast range of stimulus patterns. Synaptic activity can also be tremendously diverse; from the fast synapses of the hippocampus, to slow neuropeptide-containing synapses. Indeed, some signalling pathways are active in the absence of stimulation, such as those involved in the processing of sensory information, which transmit tonically at high rates of up to 100 Hz or more 1,2 .Intriguing questions arise from these facts, including how high rates of neurotransmission can be maintained over long periods of time, and what limits the ability of a given synapse to release neurotransmitter during sustained periods of activity. Recent data indicate that in many synapses, exocytosis of neurotransmitter is coupled to endocytosis, and that synapses have evolved a specialized apparatus of scaffolding proteins to comply with these demands. Such scaffolds aid the temporal and spatial coupling of exocytosis and endocytosis, and are crucial for maintaining rapid neurotransmission during sustained activity 3 .Exocytosis of neurotransmitter is triggered by stimulusinduced calcium influx into the nerve terminal 4,5 and the subsequent fusion of synaptic vesicles with the presynaptic plasma membrane at specialized regions called active zones (BOX 1). Exocytosed synaptic vesicle membrane proteins and lipids in turn are recycled at the endocytic or periactive zone (BOX 1) that surrounds the release site, to restore functional synaptic vesicle pools for reuse and to ensure long-term functionality of the synapse 6-8 (FIG. 1). Depending on the type of synapse and the stimulation frequency, several modes of endocytosis with different time constants -for example, fast and slow endocytosisseem to operate 7,9,10 . Clathrin-mediated endocytosis arguably represents the main pathway of synaptic vesicle endocytosis 6,8,11 , although other modes -for example, fast kiss-and-run exocytosis and endocytosis -could operate in parallel.Although the machineries for exocytic membrane fusion 12,13 and for endocytic retrieval of synaptic vesicle membranes have been studied separately in some detail 6,14 , comparatively little is known about the coupling between exocytosis and endocytosis. Here we synthesize recent data from physiological, morphological and biochemical studies in several model systems into hypothetical models for scaffold-based mechanisms underlying exocytic-endocytic coupling.The synaptic vesicle cycle Synaptic vesicle pools. Some defining features of chemical synapses are the presence of one or several clusters of synaptic vesicles in the presynaptic nerve terminal, and ultrastructural specializations at the pre-and postsy...

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“…This construct has been extensively used to investigate synaptic vesicle properties within hippocampal neurons in vitro Ryan, 2000, 2001;Fernandez-Alfonso et al, 2006) and at Drosophila (Poskanzer et al, 2003;Reiff et al, 2005) and mouse (Tabares et al, 2007) neuromuscular junctions in vivo. The pH shift that occurs during exocytosis, when the acidic vesicle interior is exposed to the neutral extracellular space, leads to an increase in spH fluorescence.…”

Section: Introductionmentioning

confidence: 99%

Heterogeneity in Synaptic Vesicle Release at Neuromuscular Synapses of Mice Expressing SynaptopHluorin

Wyatt

Balice-Gordon²

2008

J. Neurosci.

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Mammalian neuromuscular junctions are useful model synapses to study the relationship between synaptic structure and function, although these have rarely been studied together at the same synapses. To do this, we generated transgenic lines of mice in which the thy1.2 promoter drives expression of synaptopHluorin (spH) as a means of optically measuring synaptic vesicle distribution and release. SpH is colocalized with other synaptic vesicle proteins in presynaptic terminals and does not alter normal synaptic function. Nerve stimulation leads to readily detectable and reproducible fluorescence changes in motor axon terminals that vary with stimulus frequency and, when compared with electrophysiological recordings, are reliable indicators of neurotransmitter release. Measurements of fluorescence intensity changes reveal a surprising amount of heterogeneity in synaptic vesicle release throughout individual presynaptic motor axon terminals. Some discrete terminal regions consistently displayed a greater rate and extent of release than others, regardless of stimulation frequency. The amount of release at a particular site is highly correlated to the relative abundance of synaptic vesicles there, indicating that a relatively constant fraction of the total vesicular pool, ϳ30%, is released in response to activity. These studies reveal previously unknown relationships between synaptic structure and function at mammalian neuromuscular junctions and demonstrate the usefulness of spH expressing mice as a tool for studying neuromuscular synapses in adults, as well as during development and diseases that affect neuromuscular synaptic function.

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Synaptic Vesicles Interchange Their Membrane Proteins with a Large Surface Reservoir during Recycling

Cited by 186 publications

References 24 publications

Inhibition of dynamin completely blocks compensatory synaptic vesicle endocytosis

Inhibition of dynamin completely blocks compensatory synaptic vesicle endocytosis

Protein scaffolds in the coupling of synaptic exocytosis and endocytosis

Heterogeneity in Synaptic Vesicle Release at Neuromuscular Synapses of Mice Expressing SynaptopHluorin

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