Fine Structure Of Synapses Research Articles

High abundance of BDNF within glutamatergic presynapses of cultured hippocampal neurons.

In the mammalian brain, the neurotrophin brain-derived neurotrophic factor (BDNF) has emerged as a key factor for synaptic refinement, plasticity and learning. Although BDNF-induced signaling cascades are well known, the spatial aspects of the synaptic BDNF localization remained unclear. Recent data provide strong evidence for an exclusive presynaptic location and anterograde secretion of endogenous BDNF at synapses of the hippocampal circuit. In contrast, various studies using BDNF overexpression in cultured hippocampal neurons support the idea that postsynaptic elements and other dendritic structures are the preferential sites of BDNF localization and release. In this study we used rigorously tested anti-BDNF antibodies and achieved a dense labeling of endogenous BDNF close to synapses. Confocal microscopy showed natural BDNF close to many, but not all glutamatergic synapses, while neither GABAergic synapses nor postsynaptic structures carried a typical synaptic BDNF label. To visualize the BDNF distribution within the fine structure of synapses, we implemented super resolution fluorescence imaging by direct stochastic optical reconstruction microscopy (dSTORM). Two-color dSTORM images of neurites were acquired with a spatial resolution of ~20 nm. At this resolution, the synaptic scaffold proteins Bassoon and Homer exhibit hallmarks of mature synapses and form juxtaposed bars, separated by a synaptic cleft. BDNF imaging signals form granule-like clusters with a mean size of ~60 nm and are preferentially found within the fine structure of the glutamatergic presynapse. Individual glutamatergic presynapses carried up to 90% of the synaptic BDNF immunoreactivity, and only a minor fraction of BDNF molecules was found close to the postsynaptic bars. Our data proof that hippocampal neurons are able to enrich and store high amounts of BDNF in small granules within the mature glutamatergic presynapse, at a principle site of synaptic plasticity.

A cell adhesion molecule mimetic, FGL peptide, induces alterations in synapse and dendritic spine structure in the dentate gyrus of aged rats: a three‐dimensional ultrastructural study

The FGL peptide is a neural cell adhesion molecule (NCAM) mimetic comprising a 15-amino-acid-long sequence of the FG loop region of the second fibronectin type III module of NCAM. It corresponds to the binding site of NCAM for the fibroblast growth factor receptor 1. FGL improves cognitive function through enhancement of synaptic function. We examined the effect of FGL on synaptic and dendritic structure in the brains of aged (22-month-old) rats that were injected subcutaneously (8 mg/kg) at 2-day intervals until 19 days after the start of the experiment. Animals were perfused with fixative, brains removed and coronal sections cut at 50 microm. The hippocampal volume was measured, tissue embedded and ultrathin sections viewed in a JEOL 1010 electron microscope. Analyses were made of synaptic and dendritic parameters following three-dimensional reconstruction via images from a series of approximately 100 serial ultrathin sections. FGL affected neither hippocampal volume nor spine or synaptic density in the middle molecular layer of the dentate gyrus. However, it increased the ratio of mushroom to thin spines, number of multivesicular bodies and also increased the frequency of appearance of coated pits. Three-dimensional analysis showed a significant decrease in both post-synaptic density and apposition zone curvature of mushroom spines following FGL treatment, whereas for thin spines the convexity of the apposition zone increased. These data indicate that FGL induces large changes in the fine structure of synapses and dendritic spines in hippocampus of aged rats, complementing data showing its effect on cognitive processes.

Improved methods for ultracryotomy of CNS tissue for ultrastructural and immunogold analyses

We examined each step of the protocol for ultracryotomy for central nervous system tissue in order to define and overcome some of the methodological difficulties. The following three steps emerged as critical for the method's success: (1) pretreatment of grids to render them hydrophilic immediately before use; (2) careful collection of ultrathin cryosections during ultracryotomy; (3) removal of the appropriate amount of excess poly(vinyl alcohol)-uranyl acetate (PVA-UA) prior to drying after staining with PVA-UA. By taking account of the three critical steps described above, we succeeded in obtaining ultrathin cryosections, including serial sections, with excellent preservation of ultrastructure, as well as semithin cryosections which are useful for evaluating the quality of the samples and for selecting areas of interest for ultrastructural analysis. Cytoplasmic organelles in neurons and glial cells, and the fine structure of synapses and myelinated fibers were well preserved. The localization of gold particles after immunostaining for astrocytic glutamate transporter (GLAST), metabotropic glutamate receptor 1 (mGluR1) and neurofilament protein was consistent with previous reports and ultrastructure was well-preserved in all cases. These findings should be helpful to researchers wishing to carry out ultrastructural and immunogold analyses of cryosections of nervous tissue.

Fine structure of synapses and retinal innervation of substance P and adenosin deaminase containing neurons in the superior colliculus of the rat

The fine structure of substance P (SP) and adenosine deaminase (ADA) immunoreactive structures in synaptic contacts localized to the superficial layers of the superior colliculus of the rat was investigated by means of immunoelectron microscopy. We also examined the possibility of retinal innervation of SP- and ADA- containing neurons by immunohistochemistry after degeneration of retinal terminals caused by enucleation. SP-like immunoreactive presynaptic terminals of the stratum griseum superficiale (SGS) formed both asymmetric and symmetric synaptic contacts. Presynaptic dendritelike structures were also observed. SP immunoreactive postsynaptic elements made contacts with terminals showing diverse features. ADA-like immunoreactive structures were seen only as postsynaptic elements to different kinds of nonimmunoreactive terminals and were mostly localized in the ventral third of the SGS and the dorsalmost stratum opticum (SO). After enucleation, degenerating retinal terminals were found to form synaptic contacts with SP and ADA immunoreactive structures. The highest number of such degenerating terminals on ADA immunoreactive structures was observed 2 days after retinal denervation, very few being seen after 5 days. These degenerating terminals were restricted to the ventral SGS and dorsal SO. SP immunoreactive structures postsynaptic to degenerating retinal terminals were most numerous 5 days after enucleation and mainly localized in the dorsal SGS. Occasionally, SP immunoreactive dendritelike processes forming synapses with degenerating retinal terminals were simultaneously presynaptic to other nonimmunoreactive profiles, defining, therefore, serial synapses. The present results suggest that SP-I and ADA-I collicular neurons may be part of distinct channels carrying visual information to the lateral posterior and lateral geniculate nuclei of the thalamus, respectively.

Visual classification of synapses based on a combination of features

Several methods of measuring features of the fine structure of synapses accurately on electron micrographs are now available. To calculate areas and volumes of terminals, and of the mitochondria and vesicles contained in them, and also to measure the thickness of preand postsynaptic membranes, the synaptic space, and the length of the active zones, the geo~ metrical and stereological principles of Glagolev and Saltykov [2, i0, 12] are used. Determination of the absolute number of vesicles on average per terminal and per unit area of cross section of a terminal is now widely practiced [i, 8]. Active zones are classified visually on the basis of their configuration: Straight and positively and negatively curved active zones are distinguished [6]; according to the degree of abundance of the pre~ and postsynaptic membranes after special staining of brain tissue [7]; according to the structure of the postsynaptic membrane [9]; according to relations between the preand postsynaptic membranes (symmetrical and asymmetrical active zones) [4, 5], and so on. Synaptic terminals also are classified according to the structureof the vesicles they contain [5~ llJ or the distribution of vesicles over the area of cross section of the terminal [i],

Ultrastructure of synapses and cellular relationships in the oculomotor nucleus of the rhesus monkey.

The fine structure of synapses and of cellular relations was examined in the somatic efferent portion of the oculomotor nucleus of the adult rhesus monkey. Axosomatic and axodendritic synapses are characterized by distinct synaptic clefts which usually measure 20-30 nm between pre- and postsynaptic membranes. Cytoplasmic "thickenings" of pre- and postsynaptic membranes are often observed. Subjunctional bodies are present at both axosomatic and axodendritic synapses. Somatic and dendritic spine synapses are present. Serial synapses are also found, suggesting the operation of presynaptic inhibition in this nucleus. At some synapses the extracellular gap between pre- and postsynpatic membranes is reduced to 5-9 nm. However, junctions similar to the latter are also present between neurons and glia, and at the junctions between adjacent glial elements. The present results provide no evidence for a clear morphological substrate for electrotonic transmission in the somatic efferent portion of the primate oculomotor nucleus.

The fine structure of synapses in relation to the large spherical neurons in the anterior ventral cochlear of the cat.

The fine structure of synapses in relation to the large spherical neurons, the principal neurons in the anterior ventral cochlear nucleus (AVCN) of the cat, was studied. Axo-somatic synapses were classified into three types based on their size and shape and the distribution of synaptic vesicles. Axo-dendritic synapses were also classified into three types with respect to synaptic contact, cytoplasmic densities associated with the pre- and postsynaptic membranes and the shape of the synaptic vesicles. Mossy fibre-like endings and preterminal axons containing dense-core vesicles were observed in this study. Serial or triadic synapses were also found in cat AVCN. We occasionally found a nematosome in the cytoplasm of the principal neuron.

THE ORGANIZATION OF SYNAPTIC AXOPLASM IN THE LAMPREY (PETROMYZON MARINUS) CENTRAL NERVOUS SYSTEM

The fine structure of synapses in the central nervous system of lamprey (Petromyzon marinus) ammocoetes has been investigated. Both synapses within the neuropil and synaptic links between giant fibers (including Müller cells) and small postsynaptic units are described. The distribution of neurofilaments and microtubules in nerve profiles over a wide diameter range is described, and the possible role of these structures in intracellular transport is discussed. Electron micrographs indicate that small lucent "synaptic vesicles" occur sparsely throughout the axoplasm and in regular arrays in association with microtubules in the vicinity of synapses. Within a synaptic focus, immediately adjoining the presynaptic membrane, vesicles are randomly arranged and are not associated with microtubules. Neurofilaments are present, generally in large numbers, but these are not associated with vesicles or other particulates. The structural findings are considered in terms of current concepts of fast and slow transport in neurons and the mechanochemical control of intracellular movement of materials.

Fine structure of synapses in the hippocampus of the rabbit with special reference to dark presynaptic endings.

The stratum radiatum of h3 and h4 in the hippocampus of the rahbit, where the mossy fiber endings are distributed, was investigated under the electron microscope. These regions contain a certain number of electron dense presynaptic endings. These are characterized by highly dense synaptic vesicles and mitochondrial matrices. The dense endings are not considered as degenerated. Electron dense silver particles, substituted for zinc, occurred on the synaptic vesicles of these dense terminals as well as the mossy fiber endings after the application of Timm's histochemical method modified for electron microscopy. It is concluded that the dark synaptic endings observed might represent mossy fiber terminals in a special functional phase, or might be the result of structural alteration in the course of tissue preparation. The zinc localized in the synaptic vesicles is thought to be associated with the neurotransmitter present in these endings.

Fine structure of synapses in the dorsal nucleus of the lateral geniculate body of normal and blinded rats.

Degenerating boutons, observed from 2 to 60 days after eye enucleation, displayed decreased plasma membrane density, increased axoplasmic density, and enlarged mitochondria with deformed cristae when compared with boutons from normal animals. There was also a loss of synaptic plasma membrane specialization and the boutons abnormally indented contiguous dendrites. The number and appearance of synaptic vesicles in some degenerating boutons were notably altered. Phagocytosis of boutons in most instances appeared to be accomplished by astrocytes. When degeneration was first apparent in some boutons, the subsynaptic organelle in the adjacent dendritic cytoplasm was enlarged, somewhat less dense and was associated with small granular and circular profiles. Subsynaptic organelles in experimental animals were absent from contiguities between dendrites and other cell processes, except in a few instances when only small portions of boutons remained at their synaptic sites, suggesting that the organelles disappeared when boutons had been completely phagocytized.

The fine structure of synapses of the subfornical organ of the cat.

1. The subfornical organ of the cat contains various types of morphologically different synapses. The most frequent type is that of Gray 1. In addition, Gray 2 type junctions are seen. Large, multifocal synapses occur at perikarya and large dendritic stems. The most conspicuous synapse is designated “crest synapse with subjunctional bodies”; it occurs both in axodendritic and axosomatic junctions. No spine synapses have been observed. 2. All presynaptic endings are characterized by a polymorphic vesicular material. Clear vesicles range from 200–800 A in diameter and may be spheric, elliptoid or flat. A small number of dark-cored vesicles (1,000–1,500 A) is also present in most terminals; they are often located more distally from the synapse. 3. Crest synapses differ from spine synapses with respect to the profile; they typically contain two junctions, one on either side of the crest within a distance of 1,500 A in a back-to-back position. In between, there is a lattice of postsynaptic spherules in a hexagonal or triagonal array. The spherules seem to be interconnected by a network of fine extensions which may be attached to the dense material apposed to the postsynaptic membrane. 4. The subsynaptic bodies seem to occur exclusively in crest synapses with double contacts and their as yet mysterious function may be understood in terms of either mechanical buttresses or in context with the “double plug” function of a postsynaptic crest. The hypothesis that they may represent a specific innervation is rejected since identical pre- and postsynaptic elements were found to form both crest synapses with subjunctional bodies and classical, non-profiled junctions.

The fine structure of synapses and tight junctions in the central nervous system of the blowfly larva

A description is given of the structure of synaptic contacts and tight junctions in the neuropile region of the larva of the blowfly, Phormia terrae-novae, as they appear after fixation with glutaraldehyde. The chemical synapse is formed by the apposition of the presynaptic fibre, 0·5 to 4 μ in dia., with two or more smaller postsynaptic fibres, 0·1 to 0·5 μ in dia. A cleft approximately 200 Å wide separates pre- and postsynaptic fibres. Each presynaptic aynaptic axon makes many synaptic contacts. Two presynaptic axons may synapse with the same postsynaptic axons. Serial synapses are also present. Presynaptic fibres contain vesicles which are connected by filaments to electron dense bodies that are adpressed to the inner surface of the presynaptic membrane. The synaptic vesicles are also connected to one another by structures of unit membrane appearance. Electron-dense material is deposited on the inner surface of the postsynaptic membrane and occasionally on the inner surface of the presynaptic membrane. Tight junctions are found between large diameter nerve fibres. They are five-layered membrane structures formed by the fusion of the outer dense component of apposed triple-layered unit membranes. Such structures suggest that electrical synapses may be present in the neuropile of insects.

Fine structure of synapses in the cerebral cortex

The synaptolemma of axo-dendritic synapses in the rabbit striate and peristriate cortex has been examined in the electron microscope. Observations are reported of 1) a subsynaptic organelle, in the juxta-membranous part of the dendrite cytoplasm; 2) a typical attenuation of the synaptic cleft, located at the circumference or at the center of the synaptolemma; 3) interlemmal elements of two types, designated A and B.

The Fine Structure of Synapses in the Ciliary Ganglion of the Chick

Ciliary ganglia of chick embryos and newly hatched chicks were examined in the light and electron microscopes. Particular attention was given to the fine structure of calyciform synapses, which are characteristically found in ciliary ganglia of birds. The calyciform endings are characterized by large expansions of the presynaptic axons upon ganglion cells, and the terminal processes extend over a considerable area of the cell surface. Often, indeed they appear to envelop the cell. In the electron microscope image, the appositional membranes are separated by a space about 300 to 400 A wide; i.e., the synaptic cleft. At irregularly spaced regions, the appositional membranes show areas of increased density. The presynaptic processes contain clusters of synaptic vesicles, localized at these dense regions. Thus the fine structure complex typical of other synapses is evident. The unique structural features of this synapse are as follows: (a) The calyx or presynaptic terminal derives from a single axon, does not arborize, and terminates upon a single ganglion cell. Thus, unlike the classical bouton terminal, this represents an anatomical device for firing single cells by single axons. (b) The surface area in contiguity, i.e., the area of appositional membranes, is far more extensive than the bouton terminal. The fine structure of this synapse is compared with others, for example, the classical boutons terminaux and purely electrical synapses, in an attempt to correlate fine structure with function.