The fine structure of freeze-fractured presynaptic membranes

Pfenninger, Karl H.; Akert, K.; Moor, H.; Sandri, C.

doi:10.1007/bf01099180

Cited by 185 publications

(73 citation statements)

References 28 publications

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“…Although the polyhedral structure of this cage is first noted here, the existence of these units is supported by a wealth of published information, indicating their presence in a variety of central synapses (Gray, 1963;Ackert et al, 1972;Pfenninger et al, 1972;Peters et al, 1991;Zampighi and Fisher, 1997). Furthermore, a possible role of the polyhedral cage in docking and fusion is suggested by the strong presence of vesicles in their corona that are hemifused or fully fused to the plasma membrane (Fig.…”

Section: Discussionmentioning

confidence: 66%

“…This hypothesis originated with Gray' seminal studies showing that dense particles link vesicles to depressions in the active zone, called synaptopores Pfenninger et al, 1972), the dimensions of which increase on electrical stimulation (Triller and Korn, 1985). Additional studies supporting this hypothesis include the observation that the emergence of these dense particles correlates precisely with the maturation of central synapses (Bloom and Aghajanian, 1966) and that their chemical composition involves a myriad of proteins involved in cell adhesion, membrane fusion, and endocytosis (Phillips et al, 2001).…”

Section: Introductionmentioning

confidence: 99%

“…Previous studies indicate that the structure of the cytomatrix depends on the type of synapse and animal species under study. In mammalian central synapses, ϳ60 nm diameter "dense" particles that self-assemble into arrays, called the "presynaptic grid," comprise the cytomatrix (Gray, 1963;Ackert et al, 1972;Pfenninger et al, 1972). In neuromuscular junctions of frog, it appears as an arrangements of "ribs," "beams," and "pegs" are referred to as the "active zone material" (Harlow et al, 2001), whereas in neuromuscular junctions of Drosophila, the active zone consists of "T bars" (Meinerzhagen, 1996;Meinerzhagen et al, 1998).…”

Section: Introductionmentioning

confidence: 99%

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Conical Electron Tomography of a Chemical Synapse: Polyhedral Cages Dock Vesicles to the Active Zone

Zampighi¹,

Fain²,

Zampighi³

et al. 2008

J. Neurosci.

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In this study, we tested the hypothesis that the structure of the active zone of chemical synapses has remained uncertain because of limitations of conventional electron microscopy. To resolve these limitations, we reconstructed chemical synapses of rat neocortex, the archetypical "average" synapse, by conical electron tomography, a method that exhibits an isotropic in plane resolution of ϳ3 nm and eliminates the need to impose symmetry or use averaging methods to increase signal-to-noise ratios. Analysis of 17 reconstructions by semiautomatic density segmentation indicated that the active zone was constructed of a variable number of distinct "synaptic units" comprising a polyhedral cage and a corona of approximately seven vesicles. The polyhedral cages measured ϳ60 nm in diameter, with a density of ϳ44/m 2 and were associated with vesicles at the active zone ("first tier"). Vesicles in this first-tier position represented ϳ7.5% of the total number of vesicles in the terminal and were contiguous, hemifused (ϳ4% of total), or fully fused (ϳ0.5% of total) to the plasma membrane. Our study supports the hypothesis that rat neocortical synapses are constructed of variable numbers of distinct synaptic units that facilitate the docking of vesicles to the active zone and determine the number of vesicles available for immediate release.

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Section: Discussionmentioning

confidence: 66%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Conical Electron Tomography of a Chemical Synapse: Polyhedral Cages Dock Vesicles to the Active Zone

Zampighi¹,

Fain²,

Zampighi³

et al. 2008

J. Neurosci.

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“…Mem (20). As yet, no such grouping of intramembranous particles has been found associated with exocytosis in mammalian cells (21)(22)(23)(24). In our previous reported study of unstimulated mast cells, no organized arrays suggestive of predetermined loci for secretion were found (8) Fig.…”

Section: Resultsmentioning

confidence: 94%

Freeze-fracture study of mast cell secretion.

Emil¹,

Lagunoff²,

Koehler³

1976

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

100

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ABS1RACr Within seconds after exposure of rat peritoneal mast cells to polymyxin B, bulges appear on the surface of the cells. Freeze-fracture electron microscopy reveals that each bulge overlies a mast cell granule. In contrast to the even distribution of intramembranous particles in the plasma membrane of unstimulated cells, the intramembranous particles in the stimulated cells are unevenly distributed in the membrane of the bulges with large patches of membrane lacking intramembranous particles. The membranes over the most prominent bulges are entirely free of intramembranous particles, and in some instances there is an increased concentration of intramembranous particles at the margins of the bulges. Perigranule membranes exhibit the same changes in distribution of intramembranous particles. Electron microscopy of thin sections of rapidly fixed, stimulated mast cells shows a peculiar structure of the membrane overlying some bulges; instead of the pentalaminar membranes previously demonstrated, the membrane at these sites of presumptive fusion of perigranule and plasma membrane assumes the form of a single dense lamina with a fine fuzzy coating on either side. It seems possible that membrane fusion and subsequent pore formation proceed in the stimulated mast cell through a stage of flight of intramembranous particles and molecular rearrangement of the other membrane components.It is well established that specific membrane-bound cytoplasmic granules represent the storage form of histamine and heparin in mast cells (1-3). The granules are extruded from the cell under the influence of a variety of agents. Induced granule secretion has been shown to occur by exocytosis (4), in which apposing membranes of granules and cell surface interact to establish an opening (5). Involvement of membranes surrounding other granules produces a series of channels that penetrate well into the cell domain (6, 7).The extent of membrane interaction occurring during secretion would be expected to make the mast cell a particularly favorable object for the investigation of secretory membrane events. We have previously reported on the appearance of normal unstimulated mast cells with the freeze-fracture technique (8). The application of freeze-fracture technique to the study of mast cell secretion provides information complementary to that available from the previously used modes of morphologic analysis (5). MATERIALS AND METHODSPeritoneal cells, including mast cells, were collected from the peritoneal cavities of 2-to 4-month-old male CDF rats obtained from Charles River Breeding Laboratories Inc., Wilmington, Mass., or bred in our departmental animal facilities from CDF stock. In each of the degranulation experiments performed, six to eight rats were used. Six to ten million mast cells were separated from the other peritoneal cells by centrifugation through 35% albumin (9), so that mast cells constituted at least 80% by number of the final cell suspension. Mast cells were then washed once and suspended in balanced salt solution (9) at...

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“…Studies using phosphotungstic acid staining showed regularly spaced dense projections at the AZ with docked SVs nestled between them (Gray, 1963;Pfenninger et al, 1972;Triller and Korn, 1985;Peters et al, 1991;Phillips et al, 2001;Zhai and Bellen, 2004). However, aldehyde fixatives used in these studies aggregate filaments (Gulley and Reese, 1981;Tatsuoka and Reese, 1989;Gotow et al, 1991).…”

Section: Introductionmentioning

confidence: 99%

Three-Dimensional Architecture of Presynaptic Terminal Cytomatrix

Siksou¹,

Rostaing²,

Lechaire³

et al. 2007

Journal of Neuroscience

302

364

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Presynaptic terminals are specialized for mediating rapid fusion of synaptic vesicles (SVs) after calcium influx. The regulated trafficking of SVs likely results from a highly organized cytomatrix. How this cytomatrix links SVs, maintains them near the active zones (AZs) of release, and organizes docked SVs at the release sites is not fully understood.To analyze the three-dimensional (3D) architecture of the presynaptic cytomatrix, electron tomography of presynaptic terminals contacting spines was performed in the stratum radiatum of the rat hippocampal CA1 area. To preserve the cytomatrix, hippocampal slices were immobilized using high-pressure freezing, followed by cryosubstitution and embedding. SVs are surrounded by a dense network of filaments. A given vesicle is connected to ϳ1.5 neighboring ones. SVs at the periphery of this network are also linked to the plasma membrane, by longer filaments. More of these filaments are found at the AZ. At the AZ, docked SVs are grouped around presynaptic densities. Filaments with adjacent SVs emerge from these densities. Immunogold localizations revealed that synapsin is located in the presynaptic bouton, whereas Bassoon and CAST (ERC2) are at focal points next to the AZ. In synapsin triple knock-out mice, the number of SVs is reduced by 63%, but the size of the boutons is reduced by only 18%, and the mean distance of SVs to the AZ is unchanged. This 3D analysis reveals the morphological constraints exerted by the presynaptic molecular scaffold. SVs are tightly interconnected in the axonal bouton, and this network is preferentially connected to the AZ.

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The fine structure of freeze-fractured presynaptic membranes

Cited by 185 publications

References 28 publications

Conical Electron Tomography of a Chemical Synapse: Polyhedral Cages Dock Vesicles to the Active Zone

Conical Electron Tomography of a Chemical Synapse: Polyhedral Cages Dock Vesicles to the Active Zone

Freeze-fracture study of mast cell secretion.

Three-Dimensional Architecture of Presynaptic Terminal Cytomatrix

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