Small vesicle bouton synapses on the distal half of the lateral dendrite of the goldfish Mauthner cell: freeze-fracture and thin section study

To understand principles of synaptic integration, it is necessary to define the types of synapses on a particular neuron and their distribution. Thin sectioning and double replica freeze-fracture techniques were employed to characterize the small vesicle bouton (SVB) synapses on the distal half of the Mauthner (M) cell lateral dendrite, which probably mediate a remote dendritic inhibition. Three morphologically distinct SVB synapses, types A, B, and C, were found. These three SVB synapses form roughly 90% of the synapses on the distal half of the lateral dendrite, with types A and B being most common. The SVB A synapse is characterized by mostly oval and round synaptic vesicles, a discrete presynaptic active zone with a highly variable shape, and a postsynaptic active zone with no apparent particle aggregate in either the E or P face. At the SVB B synapse, most of the synaptic vesicles are flat. A very high particle density is present throughout the presynaptic P face, and vesicle attachment sites are dispersed over much of the presynaptic membrane. Postsynaptic P face particle aggregates are subjacent to the presynaptic vesicle attachment sites, and are often large and anastomosing. The SVB C synapse is characterized by synaptic vesicle profiles that vary from flattened to round. The SVB C cytoplasm was unclouded by the flocculent material that characterized SVBs A and B. The presynaptic active zones at the SVB C synapse are discrete, and macular or oblong. No particle aggregates are apparent in the postsynaptic active zone. Small, macular particle aggregates were found in nonactive zone regions of the postsynaptic E face of all three types of SVBs. Small subsurface cisterns were also observed underlying the M cell membrane at all three types of SVB synapses. Neither the postsynaptic E face aggregates nor the subsurface cisterns were ever observed directly subjacent to presynaptic active zones, but were often seen adjacent to active zones. Short, straight rows of particles and short cylinders were often seen in both pre- and postsynaptic surrounding zone regions of SVB A and C synapses. These structures are thought to represent tight junctions.     

Neuronal gap junctions and morphologically mixed synapses in the spinal cord of a teleost, Sternarchus albifrons (gymnotoidei)

Spinal cord motoneurons in the gymnotid, Sternarchus albifrons, were studied electron microscopically with special reference to the freeze-fracture method. Two types of motoneurons were identified. Electromotor neurons are monopolar and are located in a midline column dorsal to the ventral gray. These cells have a small fraction of their surface covered by synapses from descending axons, often at nodes. The synapses have multiple gap junctions, but few presynaptic vesicles or other correlates of chemical transmission. The gap junctions have an ordinary appearance in freeze-fracture replicas and exhibit a highly ordered substructure. The not infrequent appositions between the cell bodies of electromotor neurons exhibit no junctional specializations. Ordinary motoneurons are multipolar and densely covered with axosomatic and axodendritic synapses. In thin sections these synapses can be divided into two groups according to whether the vesicles are spherical or flattened. Gap junctions occur only at the first type, thus forming ‘morphologically mixed’ synapses. In freeze-etch replicas of motoneurons, the gap junctions are often found near clusters of postsynaptic E face particles elsewhere associated with excitatory chemical transmission. In addition, vesicle attachment sites occur in the presynaptic membranes of some synapses with gap junctions. The morphological observations are consistent with dual chemical and electrical transmission at these particular synapses, i.e. electrical excitation across gap junctions and chemical excitation at active zones with spherical vesicles and post-synaptic E face particles.     

Intramembranous structure of synaptic membranes with special reference to spinules in the rat sensorimotor cortex

The intramembranous structure of the synaptic contact zone at presynaptic and postsynaptic membranes in the rat sensorimotor cortex was examined by means of the freeze-etching technique. In axospinous synapses, the synaptic contact zone is characterized by perforated and nonperforated aggregates of intramembranous particles at the extracellular half or E-face of the postsynaptic membrane. On some perforated synaptic contact zones, both synaptic membranes are marked by so called spinules. These invaginations of the postsynaptic membrane and the parallel presynaptic membrane into the axon terminal are situated at the particle free zones among the postsynaptic E-face intramembranous particle aggregates or in close proximity to it. Intramembranous characteristics of the spinules at both freeze-etched faces of presynaptic and postsynaptic membranes and their density of perforated axospinous synapses were analysed. The results are discussed in terms of plasticity at the synaptic contact zone of the axospinous synapses of the sensorimotor cortex in the rat.     

Structure of the synaptic membranes in the inner plexiform layer of the retina: A freeze-fracture study in monkeys and rabbits

The internal structure of the synaptic membranes in the inner plexiform layer (IPL) of the retina of monkeys and rabbits was studied with the freeze-fracturing technique. In ribbon synapses, the presynaptic active zone is characterized by an aggregate of P-face particles, images of synaptic vesicle exocytosis, and forming coated vesicles which occupy distinct, contiguous membrane domains from apex to base of the synaptic ridge. The postsynaptic membrane contains a prominent aggregate of homogeneous particles which remain associated with the E-face. In the presynaptic membrane of conventional synapses, images of synaptic vesicle exocytosis are intermingled with large P-face particles, whereas forming coated vesicles surround the active zone. Three types of internal organization characterize the postsynaptic membrane of conventional synapses. Usually, the postsynaptic membrane exhibits the same internal structure as the surrounding nonjunctional plasmalemma. A second, less common type of conventional synapse contains a loose aggregate of heterogeneous particles which remain associated with the P-face. Finally, synapses were exceptionally found which are macular in shape and contain an aggregate of E-face particles within the postsynaptic membrane. The freeze-fracture evidence suggests that the axonal endings of bipolar cells—or at least some of them—make excitatory synapses, whereas the vast majority of amacrine cell dendrites make inhibitory synapses. Additional specializations of the cell surface in the IPL include gap junctions, puncta adhaerentia, subsurface cisterns, and cell corner aggregates.     

Ultrastructure of the electrotonic and chemical components of the lateral-to-motor and medial-to-motor synapses in crayfish nerve cord

Lateral-to-motor and medial-to-motor synapses in crayfish nerve cords are composed of an electrical and a chemical component. The presynaptic terminals showed localized clusters of synaptic vesicles, electron-dense areas, coated pits, and coated vesicles. In thin sections, active zones were defined by electron-dense regions where synaptic vesicles attached and, in freeze-fracture replicas, by clusters of intramembrane particles localized in bands with vesicle openings on the sides of these bands. The cytoplasmic surface of the postsynaptic membrane opposite the active zones was coated with electron-dense material that in freeze-fracture replicas was seen as an increase in intramembrane particles located in the external leaflet (EF-face). This specialization of the postsynaptic membrane may correspond to the neurochemical receptor. Also, pre- and postsynaptic membranes were separated by a wider extracellular gap than those of adjacent nonsynaptic regions and electrical synapses or gap junctions. Synaptic vesicles were located exclusively at the synaptic regions by means of a cytoskeleton that was different for the electrical and the chemical components. The vesicles associated with the electrical component were anchored to a cytoskeleton composed of a beaded layer of densities located parallel to the membrane. This cytoskeleton maintained the synaptic vesicles separated from the presynaptic membrane by a distance of 13 +/- 2 nm. The synaptic vesicles associated with the chemical component were anchored to electron-dense regions formed by filaments arranged in bundles, anchored to the presynaptic membrane. Vesicles lined both sides positioned to discharge their contents into the extracellular space and to replace the discharged vesicles.     

Internal organization of membranes at end bulbs of Held in the anteroventral cochlear nucleus.

The end of bulb of Held in the rostral ventral cochlear nucleus of the chinchilla and guinea pig was studied with the freeze-fracture technique. The end bulb has multiple, small active zones which are uniformly distributed within the calyceal portion of this terminal. Single or small groups of active zones are surrounded by enlarged channels of extracellular space often containing processes of astrocytes. Small plasmalemmal deformations occur at these active zones. The number of these deformations is thought to be indicative of exocytotic transmitter release because they are more frequent in animals fixed in a noisy environment compared to animals fixed in a quiet environment. Thus, our study provides a basis for the quantitative study of changes in transmitter secretion at a central nervous system synapse driven by a controllable natural stimulus. The postsynaptic active zone at end bulbs resembles other excitatory synapses in the central nervous system in having an aggregate of large particles on the external membrane leaflet. This junctional aggregate of particles is coextensive with the presynaptic active zone and with the postsynaptic density seen in thin sections. Several perisynaptic aggregates of particles are deployed around each active zone on the external membrane leaflet. These irregularly-shaped aggregates occur preferentially opposite the channels of enlarged extracellular space and along the edge of the end bulb and are not components of intercellular junctions or plasmalemmal contacts with cytoplasmic organelles. Although the function of the different particle aggregates on the postsynaptic membrane is not clear, our findings provide a basis for studying the factors controlling and maintaining their structure as well as more evidence that a consistent relationship exists between types of synaptic action and structure of the postsynaptic membrane.     

Ultrastructural correlates of electrical-chemical synaptic transmission in goldfish cranial motor nuclei

The output connections of the cranial relay neurons, part of the Mauthner cell network, were examined in goldfish with light and electron microscopic techniques. Either lucifer yellow or horseradish peroxidase (HRP) was injected into cranial relay neuron axons to demonstrate that they diverge to several motor nuclei and to many motoneurons within one nucleus. Retrograde transport of the enzyme from injections of mandibular muscles was used to label the trigeminal motoneurons. In the electron microscope, cranial relay neuron processes were distinguished by the granular appearance of the electron-opaque polymer formed enzymatically by HRP, while the retrogradely labeled motoneurons had the polymer enclosed in lysosomes. The cranial relay neuron terminals contained many presynaptic vesicles which concentrated the HRP reaction product. Active zones and synaptic clefts were evident. At some synapses, both gap junctions and presynaptic vesicles were found. The mechanism of synaptic transmission was investigated by simultaneous recording with two intracellular microelectrodes from cranial relay neuron-motoneuron pairs. Composite postsynaptic potentials in a trigeminal motoneuron were evoked by intracellular stimulation of a cranial relay neuron axon. The earliest excitatory postsynaptic potential (EPSP) component had a latency of 0.25 msec and had a peak amplitude that was not depressed by repetitive stimulation. A second component had larger peak amplitudes which were reduced easily by repetitive stimulation. Antidromic action potentials were not transmitted from motoneurons to the cranial relay neuron axons. Thus, both electrical and chemical transmission probably occur at the cranial relay neuron-motoneuron synapses. Since the cranial relay neurons fire synchronously and receive excitatory chemical synapses, the function of the gap junctions and electrical transmission is unclear. Perhaps the importance of these gap junctions is more for transport of small molecules than for impulse transmission.     

Electrical synapse formation disrupts calcium-dependent exocytosis, but not vesicle mobilization

Electrical coupling exists prior to the onset of chemical connectivity at many developing and regenerating synapses. At cholinergic synapses in vitro, trophic factors facilitated the formation of electrical synapses and interfered with functional neurotransmitter release in response to photolytic elevations of intracellular calcium. In contrast, neurons lacking trophic factor induction and electrical coupling possessed flash-evoked transmitter release. Changes in cytosolic calcium and postsynaptic responsiveness to acetylcholine were not affected by electrical coupling. These data indicate that transient electrical synapse formation delayed chemical synaptic transmission by imposing a functional block between the accumulation of presynaptic calcium and synchronized, vesicular release. Despite the inability to release neurotransmitter, neurons that had possessed strong electrical coupling recruited secretory vesicles to sites of synaptic contact. These results suggest that the mechanism by which neurotransmission is disrupted during electrical synapse formation is downstream of both calcium influx and synaptic vesicle mobilization. Therefore, electrical synaptogenesis may inhibit synaptic vesicles from acquiring a readily releasable state. We hypothesize that gap junctions might negatively interact with exocytotic processes, thereby diminishing chemical neurotransmission.     

Differences in membrane structure between excitatory and inhibitory components of the reciprocal synapse in the olfactory bulb

The reciprocal dendro-dendritic synapse between granule and mitral or tufted dendrites in the external plexiform layer of the olfactory bulb consists of an excitatory mitral-to-granule synaptic contact and an adjacent inhibitory granule-to-mitral synaptic contact. The pre- and postsynaptic membranes of both synaptic contacts were identified in replicas of freeze-fractured external plexiform layer in rabbits, mice, and chinchillas. At the excitatory synaptic contact there is a prominent specialization in the postsynaptic memberane, represented by an aggregate of homogeneous particles associated with the external half of the membrane. In contrast, the postsynaptic membrane at the inhibitory granule-to-mitral synaptic contact lacks evident internal specializations, and the distribution of particles on both fracture faces resembles that at non-synaptic regions. Less marked differences in particle distribution characterized the cytoplasmic half of the presynaptic membranes. These differences probably reflect diversity in the nature or distribution of membrane proteins at excitatory and inhibitory synapses. Protuberances on the external half of the presynaptic membrane, possibly sites of vesicle interaction with the plasma membrane, surrounded but were not coextensive with both types of synaptic contact. A few gap junctions connected proximal dendrites of mitral or tufted cells with granule cell dendrites.     

Development of synaptic junctions in cerebellar glomeruli

In the glomerular synapses of developing mouse cerebellar cortex, 2 components of the synaptic junctions assemble independently in the immature granule cell dendrites, and then combine. 'Initial' junctions between mossy fiber axons and immature granule cell dendrites have presynaptic and postsynaptic electron-dense fuzz and a widened synaptic cleft, but lack the aggregate of intramembrane particles associated with the extracellular half of the postsynaptic membrane which characterizes mature synaptic junctions. In the vicinity of 'initial junctions' there are particle aggregates which resemble those at mature synaptic junctions, but which are less densely packed and which are not associated with the other features of a junction. The constituent particles of these aggregates move to the sites of 'initial junctions' to combine with them and form 'immature synaptic junctions'. Many of these immature junctions are larger in area than mature synaptic junctions. The immature junctions accumulate a fairly uniform complement of intramembrane particles, which increase in packing density as the junctions decrease in area and attain smaller, adult size.     

Freeze-fracture study of the postsynaptic membrane of the cerebellar mossy fiber synapse in the frog

We have examined the postsynaptic membrane of the synaptic junctions of frog cerebellar mossy fibers by electron microscopy of freeze-fracture replicas and thin sections. The intramembranous particles (imps) in the E fracture face of the postsynaptic membrane are approximately 10 nm in size and form conspicuous aggregates which we classified as macular, annular, or anastomotic in form, according to the occurrence and placement of imp-free ‘windows’ within the aggregate. The size and shape of the aggregates appear related in that the area of macular aggregates is consistently smaller than the area of annular or anastomotic aggregates. Measurements of aggregate area range from 0.06 to 0.75 μm². The variable size and shape of the imp aggregate in the postsynaptic membrane sets it apart from other excitatory synapses in the central nervous system, where macular aggregates are usually described. Examination of serial thin sections suggests that the shape of the postsynaptic density is equivalent to that of the imp aggregate observed in the postsynaptic membrane by freeze-fracture. This supports the notion that the region of postsynaptic membrane associate with the postsynaptic density in thin sections correponds to the particle-rich regions of E face membrane observed in freeze-fracture replicas.     

Postnatal development of axosomatic synapses in the rat visual cortex: morphogenesis and quantitative evaluation

Postnatal development of axosomatic synapses was studied in the rat visual cortex in order to obtain experimental data that may explain how the unequal distribution of asymmetric and symmetric synapses evolves on the soma of cortical neurons. Three types of synaptic junctions were identified: asymmetric or type 1 synapses, with postsynaptic densities greater than or equal to 20 nm, symmetric type 2 synapses, and symmetric synapses with an intermediate structure. The third synapse type had a structure similar to that of type 1 synapses, although the postsynaptic densities were thinner than 20 nm. Type 1 synapses developed in three phases. In phase 1, the first postnatal week, there were many free postsynaptic thickenings and immature synapses whereby a higher degree of postsynaptic differentiation was visible in comparison to the presynaptic elements. During the following 10 days, phase 2, type 1 synapses containing thin postsynaptic densities and intermediate synapses temporarily increased in number. Intermediate synapses are interpreted as precursors of type 1 synapses that have relatively immature postsynaptic elements. Toward the end of synaptogenesis, phase 3, the free postsynaptic thickenings reappeared while type 1 synapses containing well developed postsynaptic elements prevailed. Throughout the whole postnatal period, the numerical density of axosomatic type 1 synapses remained very low and the ratio of asymmetric to symmetric synapses at the neuronal somata was inversely proportional to that at the dendrites. Also, there was a significant decrease in the numerical density of type 1 synapses between postnatal days (P) 17 and 30. Data normalized according to cortical growth suggest that this is probably due to a decrease in the number of axosomatic type 1 synapses. This corresponds to the observation that in layers III and V a few type 1 synapses were found on pyramid-like cells up to P10 which then disappeared in later stages. Axosomatic type 2 synapses appear to be formed by two different presynaptic processes. The first presynaptic type contains flocculent material with glycogen granules and resembles axonal growth cones. These junctions contain multiple adhesion patches, intermediate junctions, one or more active zones, narrow synaptic clefts, and small pleomorphic vesicles. All of these are structural features of adult type 2 synapses. The growth-cone-like presynaptic elements disappeared after about 3 weeks. The second presynaptic type is smaller in size and also forms contacts with a structure similar to adult type 2 synapses.(ABSTRACT TRUNCATED AT 400 WORDS)     

Structural changes during transmitter release at synapses in the frog sympathetic ganglion

Interactions between synaptic vesicles and the presynaptic membrane which accompany transmitter release were examined at excitatory, cholinergic synapses in bullfrog sympathetic ganglia. Ganglia were fixed at rest or during electrical stimulation of the preganglionic axons and then were either thin-sectioned or freeze-fractured. Release of transmitter for brief periods is accompanied by selective depletion of four-fifths of the synaptic vesicles aligned at active zones, an overall loss of half of the synaptic vesicles in the terminals, and synaptic vesicle openings at active zones. These findings are consistent with the hypotheses that synaptic vesicles which are ready to be released are aligned at active zones and that these vesicles fuse with and add their membrane to the presynaptic membrane as they release transmitter. Larger vesicles with dense cores also contact and open onto the presynaptic membrane at the active zone, appearing to release their contents by exocytosis. The arrangement of intramembrane particles at fractured postsynaptic specializations resembles that at other excitatory, cholinergic synapses.     

Changes in the structure of synaptic junctions during climbing fiber synaptogenesis

Synaptogenesis between climbing fiber axons and Purkinje cells involves both an orderly translocation of synaptic junctions over the Purkinje cell surface and an elimination of all but one innervating axon. We used thin-section and freeze-fracture electron microscopic techniques to study structural changes in synaptic junctions during this interval of synapse translocation and elimination. In freeze-fractured preparations, virtually all climbing fiber synaptic junctions with the perisomatic processes and somatic spines lacked the particle aggregates that characterized the extracellular half of the postsynaptic membrane of mature synaptic junctions with dendritic spines. Some climbing fiber junctions with the dendritic shaft in the second postnatal week were associated with such aggregates, despite the fact that these junctions are transient. Thus, during the interval when Purkinje cells initially were innervated by multiple climbing fibers, and subsequently denervated of all but one climbing fiber afferent per cell, only a few of the transient synaptic junctions on the cell body and proximal dendrites have associated particles. The presence of a particle aggregate at a synaptic junction does not appear to be correlated with the permanence of that junction and probably is not correlated with the capacity to support synaptic transmission. The particle aggregates might be indicative of relatively long-lived junctions, or might occur only at junctions formed by the climbing fiber that will persist in synaptic contact with the mature Purkinje cell.     

Quantitative and morphological studies on the plasticity of mixed synapses as exemplified by clear, spherical vesicles

Large mixed axosomatic synapses (synaptic membrane with chemically "active zones" and electric "gap-junction" parts) within the region of the oculomotor nucleus of the trout were stimulated for a prolonged period of time (14 days) under physiological conditions. Concerning the great number of clear vesicles that are found in the presynaptic part of this type of synapse it can be said that they can be subdivided morphometrically into two classes with respect to the special membrane complexes of the mixed synapse (active zone, gap junction). This paper is designed to find out to what an extent the proven adaptability of the "mixed synapse" system does reflect the quantitative changes in the population of these organelles. For this purpose the number (numerical density of vesicles NVv) as well as the size (vesicular volume Vv) of the vesicles were determined by stereologic methods from segments of vesicles that were visible in the electron microscopic picture. Stimulation of the contact caused an accumulation of vesicles in the presynaptic area of the active zone as well as in the region of the terminal axonal cylinder (myelin-free presynaptic part of the axon--"marginal zone"). At the gap junctions the number of vesicles remained unchanged, but their size diminished significantly. On the one hand, the findings obtained are discussed as an expression of potential compensatory mechanisms involved in vesicle supply or formation (axonal transport, recycling) for chemical transmission, and, on the other, as indicating possible interactions between vesicles and gap junctions for electric transmission. They suggest that the behaviour of the vesicle population of mixed synapses is governed by laws of its own.     

Electron microscopic autoradiography of the uptake of [H]GABA in dispersed cell cultures of rat cerebellums. II. The development of gabaergic synapses

The formation of GABAergic synapses in dispersed cell cultures of the rat cerebellum was followed from 7 to 21 days in vitro (DIV). The majority of GABAergic synapses appeared between 10 and 14 DIV, and apparently no additional GABAergic synapses formed after 14 DIV.The first step in the development of a GABAergic synapse appeared to be the formation of a large diameter swelling in a GABAergic neuronal process. After the initial contact between the pre- and postsynaptic elements was established, both the number of synaptic vesicles and the thickness of the postsynaptic density increased, while the cross-sectional area of the presynaptic element decreased. The length of the postsynaptic density showed some increase, but no changes were noted in the synaptic cleft thickness, size of the synaptic vesicles or the shape of the synaptic vesicles.Our findings indicate that the formation of GABAergic synapses was not preceded by the formation of other types of junctions or preformed synaptic elements. In addition, the timing and the rate of formation of GABAergic synapses appears not to depend on contact with a single type of postsynaptic neuron, but rather to depend upon intrinsic properties of the development of the GABAergic neuron.     

The formation of chemical synapses between cell-cultured neuronal somata

The study of the development and plasticity of chemical synaptic connections is frequently restricted by the lack of access to the synaptic terminals. This can, in part, be overcome by plating neurons into cell culture where all regions of a neuron are made experimentally accessible. However, the small size of synaptic terminals still makes direct experimental manipulation difficult. In this study we have found in the absence of neurite extension, directly contacting cell somata (diameter 50-100 mumol) will form chemical synapses. Identified neurons B5 and B19 of Helisoma were plated into culture under conditions that promote adhesion between cell pairs. Under these conditions, neurite outgrowth was absent, but action potentials in B5 evoked inhibitory postsynaptic potentials in B19 that were reversed in sign by the injection of chloride ions and were blocked by tubocurare (10(-5) M), reduced extracellular Ca2+, and Cd2+ ions. Such synapses exhibited classical properties of chemical synapses, including the spontaneous release of neurotransmitter. Since somatic synapses represent an appropriate model of synaptic transmission, this system was utilized to study the role of mutual neuronal contact in the development of transmitter release capabilities. Future pre- and postsynaptic somata were cultured separately for 3 d, the period required for the development of synaptic transmission under conditions of maintained contact. Then, neurons were made to contact and intracellular recordings taken within 0 to 4 hours. Postsynaptic potentials were detected as early as 10 sec following contact. Thus, qualitatively the development of transmitter release capabilities does not require maintained contact.(ABSTRACT TRUNCATED AT 250 WORDS)     

Perforated and non-perforated synapses in rat neocortex: three-dimensional reconstructions

Perforated and non-perforated synapses in the molecular layer of rat parietal cortex have been assessed morphologically and quantitatively using three-dimensional reconstructions of the postsynaptic terminal. Perforated synapses were analyzed at nine ages, ranging from 0.5 to 22 months of age, and non-perforated synapses at three ages--0.5, 12, and 22 months. Examination of the reconstructions shows that perforated synapses increase in size and complexity with increasing age. This increasing complexity is reflected in a break-up of the postsynaptic density, which is punctuated by larger, branched perforations. In the most extreme cases the result is the appearance of isolated islands of postsynaptic density separated by, and also surrounded by, a synaptic contact zone. Spinules are especially prominent at around 12 months of age in perforated synapses, and the overall negative curvature of the young junctions is replaced by positively curved junctions from 4 months onwards. The non-perforated synapses are relatively small and show few changes with increasing age. Using the measurement option in the reconstruction program, the following trends emerged. All parameters of perforated synapses increased in size with increasing age, whereas the corresponding parameters of non-perforated synapses remained relatively unchanged over this age range. In addition, the percentage of the synaptic contact zone surface area occupied by the postsynaptic density decreased with increasing age in perforated synapses, but increased in non-perforated synapses. The total postsynaptic density surface area of non-perforated synapses per unit volume of molecular layer was double that of perforated synapses at 0.5 months, but the situation was reversed at 12 months. This parameter was similar in the 2 populations at 22 months. This suggests that perforated synapses contribute more to the total surface area of the postsynaptic density in mid- to late-adulthood than do non-perforated synapses, despite non-perforated synapses outnumbering perforated by 2-3:1 at these ages. These data provide more specific evidence that perforated and non-perforated synapses constitute separate synaptic populations from early in development, and that perforated synapses are responsible for the maintenance of neuronal postsynaptic density surface area from mid-adulthood onwards.     

Development of the rhesus monkey retina. I. Emergence of the inner plexiform layer and its synapses

The emergence and differentiation of the inner plexiform layer (IPL) and the establishment of its synapses were analyzed in retinae of 18 rhesus monkeys ranging in age from the 55th embryonic day (E55) to 10 years. The IPL becomes recognizable by E65 as a thin acellular zone consisting of immature neurites and growth cones scattered within large extracellular spaces. In each specimen, apposing paired membrane specializations were classified as junctions without synaptic vesicles, conventional synapses, ribbon synapses, or gap junctions. Initially, at E65, the IPL consists of variety of immature cell processes that are interconnected exclusively by junctions without synaptic vesicles, at a density of 4.7/100 microns2. By E73, the IPL becomes more distinct and wider and contains 7.8 such junctions/100 microns2. Conventional synapses develop by the addition of vesicles to initially vesicle-free junctions. The first conventional synapses appear at E78. They increase in density from 1.5 to 3.2/100 microns2 between E78 and E84 and reach a density of 7.9/100 microns2 by E99. A rapid burst in synaptogenesis occurs in the IPL between E99 and E114; a density of 16.5/100 microns2 is reached, mainly due to accretion of conventional synapses. Ribbon synapses first become recognizable at E99, almost 3 weeks after the emergence of conventional synapses. By E114 they account for about 7% of all synaptic contacts in the IPL. The rate of synaptogenesis slows down during the last quarter of gestation; the adult level of about 24 contacts/100 microns2 is reached between E130 and E149. Of these, 72.2% are of conventional type, 15.4% are ribbon synapses, and 12.4% are junctions without vesicles. However, in the adult the density of junction without vesicles is only about one-half that found at E149. Gap junctions are absent during the initial and rapid phases of synaptogenesis; they appear abruptly, between E130 and E149, only after the density of chemical synapses in the IPL has reached the adult level. In the rhesus monkey, synaptogenesis begins several weeks later in the IPL than in its primary targets--the dorsal lateral geniculate nucleus and the superior colliculus (Hendrickson and Rakic, '77; Cooper and Rakic, '83). However, the rapid increase in density of conventional synapses in the IPL coincides with the segregation of retinal projections from right and left eyes in the geniculate nucleus (Rakic, '76) and with the elimination of the large surplus of retinal ganglion cell axons (Rakic and Reley, '83).     

Motor nerve terminals on abdominal muscles in larval flesh flies,Sarcophaga bullata: Comparisons with Drosophila

Motor nerve terminals on abdominal body-wall muscles 6A and 7A in larval flesh flies were investigated to establish their general structural features with confocal microscopy, transmission electron microscopy, and freeze-fracture procedures. As in Drosophila and other dipterans, two motor axons supply these muscles, and two morphologically different terminals were discerned with confocal microscopy: thin terminals with relatively small varicosities (Type Is), and thicker terminals with larger varicosities (Type Ib). In serial electron micrographs, Type Ib terminals were distinguished from Type Is terminals by their larger cross-sectional area, more extensive subsynaptic reticulum, more mitochondrial profiles, and more clear synaptic vesicles. Type Ib terminals possessed larger synapses and more synaptic contact area per unit terminal length. Although presynaptic dense bars of active zones were similar in mean length for the two terminal types, there were almost twice as many dense bars per synapse for Type Ib terminals. Freeze-fractures through the presynaptic membrane showed particle-free areas indicative of synapses on the P-face, within which were localized aggregations of large intramembranous particles indicative of active zones. These particles were similar in number to those found at active zones of several other arthropod neuromuscular junctions. In general, synaptic structural parameters strongly paralleled those of the anatomically homologous muscles in Drosophila melanogaster. In live preparations, simultaneous focal recording from identified varicosities and intracellular recording indicated that the two terminals produced excitatory junction potentials of similar amplitude in a physiological solution similar to that used for Drosophila. J. Comp. Neurol. 402:197–209, 1998. © 1998 Wiley-Liss, Inc.