Conical Tomography of a Ribbon Synapse: Structural Evidence for Vesicle Fusion

Zampighi, Guido A.; Schietroma, Cataldo; Zampighi, Lorenzo; Woodruff, Michael L.; Wright, Ernest M.; Brecha, Nicholas C.

doi:10.1371/journal.pone.0016944

Cited by 59 publications

(66 citation statements)

References 115 publications

(164 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…6G–H and 7D). The aforementioned fracture pattern is consistent solely with “hemi-fusion,” which involves merging the external leaflet of the plasma membrane with the luminal leaflet of the synaptic vesicle at the contact region [25], [31]–[32]. Thus, our observations of rapidly frozen cortical neuropils without glutaraldehyde cross-linking are consistent with the concept that secretion of neurotransmitters involves the fusion-through-hemi-fusion hypothesis [33].…”

Section: Resultssupporting

confidence: 83%

A Novel Synaptic Vesicle Fusion Path in the Rat Cerebral Cortex: The “Saddle” Point Hypothesis

2014

Self Cite

View full text Add to dashboard Cite

We improved freeze-fracture electron microscopy to study synapses in the neuropil of the rat cerebral cortex at ∼2 nm resolution and in three-dimensions. In the pre-synaptic axon, we found that “rods” assembled from short filaments protruding from the vesicle and the plasma membrane connects synaptic vesicles to the membrane of the active zone. We equated these “connector rods” to protein complexes involved in “docking” and “priming” vesicles to the active zone. Depending on their orientation, the “rods” define two synaptic vesicle-fusion paths: When parallel to the plasma membrane, the vesicles hemi-fuse anywhere (“randomly”) in the active zone following the conventional path anticipated by the SNARE hypothesis. When perpendicular to the plasma membrane, the vesicles hemi-fuse at the base of sharp crooks, called “indentations,” that are spaced 75–85 nm center-to-center, arranged in files and contained within gutters. They result from primary and secondary membrane curvatures that intersect at stationary inflection (“saddle”) points. Computer simulations indicate that this novel vesicle-fusion path evokes neurotransmitter concentration domains on the post-synaptic spine that are wider, shallower, and that reach higher average concentrations than the more conventional vesicle fusion path. In the post-synaptic spine, large (∼9× ∼15 nm) rectangular particles at densities of 72±10/ µm2 (170–240/spine) match the envelopes of the homotetrameric GluR2 AMPA-sensitive receptor. While these putative receptors join clusters, called the “post-synaptic domains,” the overwhelming majority of the rectangular particles formed bands in the “non-synaptic” plasma membrane of the spine. In conclusion, in the neuropil of the rat cerebral cortex, curvatures of the plasma membrane define a novel vesicle-fusion path that preconditions specific regions of the active zone for neurotransmitter release. We hypothesize that a change in the hybridization of the R-SNARE synaptobrevin from parallel to antiparallel swings the synapse into this novel vesicle-fusion path.

show abstract

Section: Resultssupporting

confidence: 83%

A Novel Synaptic Vesicle Fusion Path in the Rat Cerebral Cortex: The “Saddle” Point Hypothesis

2014

Self Cite

View full text Add to dashboard Cite

show abstract

“…To provide an anatomical correlate for our physiological estimates of vesicle pools, we inventoried synaptic vesicle populations using ultrastructural methods, assuming that the preferential site of exocytosis is located near synaptic ribbons (Zenisek et al, 2000; Zampighi et al, 2011). We first quantified the number and size of ribbon synapses per hair cell by serial section electron microscopy (Figure 4).…”

Section: Resultsmentioning

confidence: 99%

Sharp Ca²⁺Nanodomains beneath the Ribbon Promote Highly Synchronous Multivesicular Release at Hair Cell Synapses

Graydon

Cho²,

Li³

et al. 2011

J. Neurosci.

152

212

View full text Add to dashboard Cite

Hair cell ribbon synapses exhibit several distinguishing features. Structurally, a dense body, or ribbon, is anchored to the presynaptic membrane and tethers synaptic vesicles; functionally, neurotransmitter release is dominated by large EPSC events produced by seemingly synchronous multivesicular release. However, the specific role of the synaptic ribbon in promoting this form of release remains elusive. Using complete ultrastructural reconstructions and capacitance measurements of bullfrog amphibian papilla hair cells dialyzed with high concentrations of a slow Ca2+ buffer (10 mM EGTA), we found that the number of synaptic vesicles at the base of the ribbon correlated closely to those vesicles that released most rapidly and efficiently, while the rest of the ribbon-tethered vesicles correlated to a second, slower pool of vesicles. Combined with the persistence of multivesicular release in extreme Ca2+ buffering conditions (10 mM BAPTA), our data argues against the Ca2+-dependent compound fusion of ribbon-tethered vesicles at hair cell synapses. Moreover, during hair cell depolarization, our results suggest that elevated Ca2+ levels enhance vesicle pool replenishment rates. Finally, using Ca2+ diffusion simulations, we propose that the ribbon and its vesicles define a small cytoplasmic volume where Ca2+ buffer is saturated, despite 10 mM BAPTA conditions. This local buffer saturation permits fast and large Ca2+ rises near release sites beneath the synaptic ribbon that can trigger multiquantal EPSCs. We conclude that, by restricting the available presynaptic volume, the ribbon may be creating conditions for the synchronous release of a small cohort of docked vesicles.

show abstract

“…S1, and Table S1; all P values <0.05), indicating that the VM and PM had been fixed during the formation of a single bilayer characteristic of hemifusion, an intermediate state of membrane fusion that precedes fusion pore formation (33). VM-PM hemifusion, as assessed visually, has been reported for CNS synapses (34,35). Extent of VM-PM contact areas.…”

Section: Vm-pm Contact Sites In Terminals Fixed During Repetitive Evokedmentioning

confidence: 94%

Variable priming of a docked synaptic vesicle

Jung

Szule

Marshall

et al. 2016

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

119

View full text Add to dashboard Cite

The priming of a docked synaptic vesicle determines the probability of its membrane (VM) fusing with the presynaptic membrane (PM) when a nerve impulse arrives. To gain insight into the nature of priming, we searched by electron tomography for structural relationships correlated with fusion probability at active zones of axon terminals at frog neuromuscular junctions. For terminals fixed at rest, the contact area between the VM of docked vesicles and PM varied >10-fold with a normal distribution. There was no merging of the membranes. For terminals fixed during repetitive evoked synaptic transmission, the normal distribution of contact areas was shifted to the left, due in part to a decreased number of large contact areas, and there was a subpopulation of large contact areas where the membranes were hemifused, an intermediate preceding complete fusion. Thus, fusion probability of a docked vesicle is related to the extent of its VM-PM contact area. For terminals fixed 1 h after activity, the distribution of contact areas recovered to that at rest, indicating the extent of a VM-PM contact area is dynamic and in equilibrium. The extent of VM-PM contact areas in resting terminals correlated with eccentricity in vesicle shape caused by force toward the PM and with shortness of active zone material macromolecules linking vesicles to PM components, some thought to include Ca 2+ channels. We propose that priming is a variable continuum of events imposing variable fusion probability on each vesicle and is regulated by force-generating shortening of active zone material macromolecules in dynamic equilibrium.synapse | hemifusion | priming | active zone material | electron tomography S ynaptic vesicles (SVs) move toward and dock on (are held in contact with) the presynaptic plasma membrane (PM) of a neuron's axon terminal before fusing with the PM and releasing their neurotransmitter into the synaptic cleft to mediate synaptic transmission (1, 2). Docking is achieved by force-generating interactions of the vesicle membrane (VM) protein synaptobrevin with the PM proteins syntaxin and SNAP25 (3, 4). These interactions, which produce force by forming a coiled coil called the SNARE core complex, are regulated by auxiliary proteins (1, 5-7). Such force on the VM-PM contact site may also play a role in their fusion (8). At typical synapses, docking and fusion take place at structurally specialized regions along the PM called active zones (9, 10). Several lines of evidence suggest that the formation of the SNARE core complex occurs in the macromolecules composing the common active zone organelle, active zone material (AZM) (2,(11)(12)(13)(14), which is positioned near Ca 2+ channels concentrated in the PM at active zones (15-17). Influx of Ca 2+ through the channels after the arrival of a nerve impulse triggers fusion of the VM of docked SVs with the PM.Priming is a step in synaptic transmission between the docking of an SV on, and fusion with, the PM and accounts for the observation that relatively few docked SVs fuse with the PM...

show abstract

Conical Tomography of a Ribbon Synapse: Structural Evidence for Vesicle Fusion

Cited by 59 publications

References 115 publications

A Novel Synaptic Vesicle Fusion Path in the Rat Cerebral Cortex: The “Saddle” Point Hypothesis

A Novel Synaptic Vesicle Fusion Path in the Rat Cerebral Cortex: The “Saddle” Point Hypothesis

Sharp Ca²⁺Nanodomains beneath the Ribbon Promote Highly Synchronous Multivesicular Release at Hair Cell Synapses

Variable priming of a docked synaptic vesicle

Contact Info

Product

Resources

About

Conical Tomography of a Ribbon Synapse: Structural Evidence for Vesicle Fusion

Cited by 59 publications

References 115 publications

A Novel Synaptic Vesicle Fusion Path in the Rat Cerebral Cortex: The “Saddle” Point Hypothesis

A Novel Synaptic Vesicle Fusion Path in the Rat Cerebral Cortex: The “Saddle” Point Hypothesis

Sharp Ca2+Nanodomains beneath the Ribbon Promote Highly Synchronous Multivesicular Release at Hair Cell Synapses

Variable priming of a docked synaptic vesicle

Contact Info

Product

Resources

About

Sharp Ca²⁺Nanodomains beneath the Ribbon Promote Highly Synchronous Multivesicular Release at Hair Cell Synapses