The effect of light stimulation in the pre- and postnatal periods on the synaptic ultrastructure in the chick hyperstriatum

Morphometric analysis of synapses in the medial parts of the ventral and accessorium hyperstriatum in the right and left hemispheres in chicks has been performed after rhythmic optic stimulation from the 18th day of incubation, flickering stimulation at the 11-12th hour after hatching, as well as in chicks reared under normal illumination and those kept in darkness within the first two days after hatching. Numeric density of synapses, mean dimensions of postsynaptic thickenings, the number of synaptic vesicles per active zone in presynapses and mean length of the whole axodendritic contact were determined in ultrathin sections. The data obtained indicate that early visual experience significantly affects the synaptic structures in both parts of the brain. A discussion is made of the plasticity of synapses during stimulation, morphological and functional relationships between hyperstriatal areas in relation to different aspects of processing and storage of visual information.     

视皮层LTP维持阶段的突触形态计量学研究

本实验使用１８～２０ｄ的幼年大鼠视皮层脑片标本,在ＬＴＰ出现后３ｈ取局部微脑片固定进行ＬＴＰ维持阶段超微结构的研究。分别与孵育相同时间而未予任何刺激的脑片和仅给予测试刺激的脑片作比较。运用图像分析仪分别对三组电镜结果进行以下参数的测量：（１）突触间隙的宽度;（２）突触后致密物（ＰＳＤ）的厚度;（３）活性区的长度;和（４）突触界面曲率。用双盲法对突触数目进行计量,并用立体计量学方法对各种突触类型进行定量,所得数据用方差分析进行统计学处理。结果显示：（１）ＬＴＰ形成后１５ｈ左右,其反应达到峰值,然后维持在最高水平一直到３ｈ仍无下降趋势;（２）突触间隙的宽度较两个对照组明显增宽;（３）ＰＳＤ的厚度也明显增厚;（４）活性区的面密度及突触界面曲率明显增加;（５）总突触数目和棘突触数目的数密度较空白对照明显增高;（６）穿孔性突触的数密度与对照组相比明显增加。结果提示：活性区面密度的增加及突触界面曲率的增大可能是ＬＴＰ维持的形态学基础。穿孔性突触的形成与ＬＴＰ的维持密切相关。     

A morphometric study of postnatal synaptogenesis in the neocortex of precocial and altricial rodents]

Quantitative electronmicroscopic studies have been made on the development of synapses in two modally different areas of the brain (V-VI layers of the visual and auditory cortex) in the rat and mouse Acomys cahirinus within first two weeks of their postnatal life. The density of synapses as well the relative amount of different types of synapses (symmetrical, asymmetrical, axo-spinal and synapses with large amounts of synaptic vesicles) were measured. It was shown that only in rats the development of synapses in the visual area usually is faster than in the auditory one.     

Quantitative Ultrastructural Investigations on Synaptogenesis in the Cerebellum and the Optic Tectum of Light-Reared and Dark-Reared Rainbow Trout (Salmo gairdneri Rich.)

The morphogenetic differentiation of synapses in the cerebellum and the optic tectum of darkand light- reared rainbow trout was investigated at critical stages of development. During normal differentiation the cerebellum is characterized by the appearance of 'indented', spinelike synapses. This type of synapses increases with age and prevails from day 60 on. At the same time the number of 'flat' synapses decreases. In the cerebellum the highest synaptic density (123 ± 12 synapses/1,000 μm²) is reached 30 days after hatching when the larvae begin to swim. The optic tectum is characterized by a preponderance of flat synapses in early postnatal and adult life; maximal synaptic density (66 ± 5 synapses/1,000 μm²) is reached 60 days after hatching when the larvae have reached optimal visual acuity.
Light deprivation causes a considerable and significant reduction in the number of synapses per unit area in the cerebellum and the optic tectum. The length of synaptic contacts do not change. If light-deprived, the density of synaptic vesicles decreases significantly in the optic tectum of a 25-day-old trout (74 ± 3 instead of 132 ± 7 vesicles/μm²). In the cerebellum this effect is absent.     

Neurotrophin-3 is required for appropriate establishment of thalamocortical connections

Ca2+-permeable AMPARs are inwardly rectifying due to block by intracellular polyamines. Neuronal activity regulates polyamine synthesis, yet whether this affects Ca2+-AMPAR-mediated synaptic transmission is unknown. We test whether 4 hr of increased visual stimulation regulates glutamatergic retino-tectal synapses in Xenopus tadpoles. Tectal neurons containing Ca2+-AMPARs form a gradient along the rostro-caudal developmental axis. These neurons had inwardly rectifying AMPAR-mediated EPSCs. Four hours of visual stimulation or addition of intracellular spermine increased rectification in immature neurons. Polyamine synthesis inhibitors blocked the effect of visual stimulation, suggesting that visual activity regulates AMPARs via the polyamine synthesis pathway. This modulation resulted in changes in the integrative properties of tectal neurons. Regulation of polyamine synthesis by physiological stimuli is a novel form of modulation of synaptic transmission important for understanding the short-term effects of enhanced sensory experience during development.     

Ultrastructural changes in the interneuronal contacts of the upper layers of the cerebral cortex in cats when aminazine is applied

The fine structure of upper layer of axodendritic synapses of the cat cerebral cortex was studied during 10, 45 and 90 minute applications of aminazine, and also 2.5 and 5 hours after the cessation of the 90 minute application. During the 10 and 45 minute applications no clear-cut changes are observed in the ultrastructure of the synapses. 90 minutes after the onset of application osmophilic changes are enhanced in the pre- and postsynaptic areas. During the drug application, the number of synaptic vesicles and the area of the axonal terminals decrease. The above morphological changes are reversible: restoration of the number of synaptic vesicles commences, but the area of the terminal is still shrunken.     

Information given by interneuronal junctions of the rat cerebral cortex during the post-asphyxia period

Using PTA technique, the organization of subsynaptic units (paramembrane specialized formations of the cytoskeleton) was revealed in the molecular layer of the rat neocortex during post-asphyxia period. The density and summarized length of the active contact zone (ACZ) with different degree of maturity of subsynaptic units were calculated and the information was evaluated. It was revealed that the total length of ACZ synapses was restored to the control level on the 7th day after asphyxia and the information of the synaptic pool was 30.1% on the 14th day, in spite of a significant deficiency in the synapse density. The information of the synaptic pool was restored mainly due to hypertrophy of interneuronal contacts with well developed presynaptic grating.     

Veratridine-stimulated central synapses in culture: A quantitative ultrastructural analysis

Synapses in explant cultures of fetal rat neocortex at day 18 in vitro were stimulated by veratridine (10^?4M) for 20 min. The cultures were subsequently processed for electron microscopy and the synapses were analyzed by quantitative techniques, incorporating set mathematical treatment. The mean values of area, perimeter, and form factor of the presynaptic elements significantly increased following veratridine stimulation, compared to the values of control synapses. The length of the postsynaptic thickening also increased, while synaptic curvature did not change significantly in the veratridine group. A fivefold reduction was observed in the mean number of synaptic vesicles per presynaptic element and in the vesicle-terminal area ratio, following veratridine stimulation. The cytoplasm-terminal area ratio and the occurrence of vacuoles/cisternae significantly increased after veratridine application. Planar measurement of membranes (boundary length) of different presynaptic organelles revealed that the total membrane did not change significantly in the veratridine group. The data indicated an increase in volume and swelling of the pre- and postsynaptic elements, considerable depletion of synaptic vesicles, and preservation of the total presynaptic membrane following veratridine stimulation in nerve tissue culture.     

N-Cadherin Relocalizes from the Periphery to the Center of the Synapse after Transient Synaptic Stimulation in Hippocampal Neurons

N-cadherin is a cell adhesion molecule which is enriched at synapses. Binding of N-cadherin molecules to each other across the synaptic cleft has been postulated to stabilize adhesion between the presynaptic bouton and the postsynaptic terminal. N-cadherin is also required for activity-induced changes at synapses, including hippocampal long term potentiation and activity-induced spine expansion and stabilization. We hypothesized that these activity-dependent changes might involve changes in N-cadherin localization within synapses. To determine whether synaptic activity changes the localization of N-cadherin, we used structured illumination microscopy, a super-resolution approach which overcomes the conventional resolution limits of light microscopy, to visualize the localization of N-cadherin within synapses of hippocampal neurons. We found that synaptic N-cadherin exhibits a spectrum of localization patterns, ranging from puncta at the periphery of the synapse adjacent to the active zone to an even distribution along the synaptic cleft. Furthermore, the N-cadherin localization pattern within synapses changes during KCl depolarization and after transient synaptic stimulation. During KCl depolarization, N-cadherin relocalizes away from the central region of the synaptic cleft to the periphery of the synapse. In contrast, after transient synaptic stimulation with KCl followed by a period of rest in normal media, fewer synapses have N-cadherin present as puncta at the periphery and more synapses have N-cadherin present more centrally and uniformly along the synapse compared to unstimulated cells. This indicates that transient synaptic stimulation modulates N-cadherin localization within the synapse. These results bring new information to the structural organization and activity-induced changes occurring at synapses, and suggest that N-cadherin relocalization may contribute to activity dependent changes at synapses.     

Activity-independent prespecification of synaptic partners in the visual map of Drosophila

Specifying synaptic partners and regulating synaptic numbers are at least partly activity-dependent processes during visual map formation in all systems investigated to date . In Drosophila, six photoreceptors that view the same point in visual space have to be sorted into synaptic modules called cartridges in order to form a visuotopically correct map . Synapse numbers per photoreceptor terminal and cartridge are both precisely regulated . However, it is unknown whether an activity-dependent mechanism or a genetically encoded developmental program regulates synapse numbers. We performed a large-scale quantitative ultrastructural analysis of photoreceptor synapses in mutants affecting the generation of electrical potentials (norpA, trp;trpl), neurotransmitter release (hdc, syt), vesicle endocytosis (synj), the trafficking of specific guidance molecules during photoreceptor targeting (sec15), a specific guidance receptor required for visual map formation (Dlar), and 57 other novel synaptic mutants affecting 43 genes. Remarkably, in all these mutants, individual photoreceptors form the correct number of synapses per presynaptic terminal independently of cartridge composition. Hence, our data show that each photoreceptor forms a precise and constant number of afferent synapses independently of neuronal activity and partner accuracy. Our data suggest cell-autonomous control of synapse numbers as part of a developmental program of activity-independent steps that lead to a "hard-wired" visual map in the fly brain.     

The role of the synaptic vesicles in transmission at the insect nerve-muscle junction

The ultrastructure of nerve-muscle synapses on red and white fibres of locust ($\text{Schistocerca}$$\text{gregaria}$) retractor unguis muscle fibres was examined before and after stimulation. At a stimulation frequency of 15Hz, the synapses on the white fibres fatigued completely; those on the red fibres also fatigued, but during prolonged stimulation they showed intermittent periods of recovery. Synaptic vesicles at both types of fatigued synapses were reduced in volume compared with controls and at fatigued synapses on white fibres the vesicles had irregular outlines. Aggregation of vesicles were observed at fatigued synapses on red fibres and the synaptic cleft width at these synapses was less than at control synapses on red fibres. These results are discussed in the light of the vesicle hypothesis for synaptic transmission.     

Participation of puncta adherentia junctions in the formation of synaptic connections between a dentate fascia graft and the host brain

The fetal dentate fascia of Wistar rats on the 20th day of gestation was heterotopically grafted into the somatosensory neocortex of adult rats. Granule cells of a graft projected their axons (mossy fibers) to the host brain and established synaptic contacts with inappropriate targets. The organization of ectopic mossy fiber synapses was studied by electron microscopy. It was shown that ectopic synapses reproduce the structural determinants of hippocampal giant synapses and induce a subcellular reorganization of postsynaptic neocortex dendrites. Using morphometric analysis, a significant increase was found in the number of discrete puncta adherentia junctions and their total length in ectopic synapses as compared with the control group. The data obtained indicate that puncta adherentia contacts participate in the structural and chemical adaptation of neuronal targets to alien axons growing from transplants.     

Lernen und Gedächtnis: Zelluläre und molekulare Grundlagen

Learning and memory are a key issue of current neuroscience research. Scientists from several disciplines have suggested that the processes of learning and memory are encoded via activity‐dependent changes in the strength of the synapse. As a result of this focus, a huge amount of effort has been invested in the understanding of cellular and molecular mechanisms behind these changes in synaptic efficacy. One phenomenon is that after repeated paring or high‐frequency stimulation synapses can be potentiated and that this enhancement of synaptic strength can last from hours to days (the so called long term potentiation, LTP). Apart from these functional changes it has recently been shown that structural changes at a synapse or in the number of synapses can be correlated with activity‐dependent processes involved in long‐term memory storage. A promising candidate molecule to link changes in function to changes in structure is the nerve‐growth factor BDNF (brain derived neurotrophic factor)     

Age-related changes in the synapses of the rat's neostriatum

By means of transmission electron microscopy, the age-related changes in axospinous (ASS) and axodendritic (ADS) synapses in the dorsal part of the rostral neostriatum in two groups of Wistar rats: young (3-month-old), and senescent (25-month-old) were examined. The changes in different parameters, characterizing the ASS and ADS: synaptic density (SD), number of synaptic vesicles (SV), number of synaptic contact zone (SCZ), and number of dendritic spines, bearing synapses (DS) were investigated morphometrically. The SD of the ASS decreased significantly during aging, but the SD of the ADS did not changed significantly. The mean area of the synaptic boutons increased significantly during aging in two types of synapses. The mean number of vesicles per synaptic bouton increased, but the number of vesicles per microm2 of synaptic bouton, and per microm3 of the neuropil decreased. The mean SCZ length increased in both types of synapses. The total SCZ length per 1000 microm2 of the neuropil, and the total area of the SCZ per 1000 microm3 of the neuropil decreased in ASS, but the same parameters of the ADS did not changed significantly. The mean number of synaptic DS per 1000 microm2 of the neropil decreased during aging, but the mean area of the synaptic DS increased. The present results support the hypothesis that the synaptic contacts change significantly during aging, and the ASS are more vulnerable during aging than the ADS.     

Geometry and structural plasticity of synaptic connectivity

Changes in synaptic connectivity patterns through the formation and elimination of dendritic spines may contribute to structural plasticity in the brain. We characterize this contribution quantitatively by estimating the number of different synaptic connectivity patterns attainable without major arbor remodeling. This number depends on the ratio of the synapses on a dendrite to the axons that pass within a spine length of that dendrite. We call this ratio the filling fraction and calculate it from geometrical analysis and anatomical data. The filling fraction is 0.26 in mouse neocortex, 0.22-0.34 in rat hippocampus. In the macaque visual cortex, the filling fraction increases by a factor of 1.6-1.8 from area V1 to areas V2, V4, and 7a. Since the filling fraction is much smaller than 1, spine remodeling can make a large contribution to structural plasticity.     

Synaptic potential in the motor giant axon of the crayfish

Some electrical properties of the synapses between central giant axons (presynaptic) and the motor giant axon (postsynaptic) of the crayfish abdominal nerve cord have been investigated. Postsynaptic potential change in response to presynaptic volleys contains two components: a spike potential and a synaptic potential of very long time course. Amplitude of the synaptic potential is graded according to the number of active presynaptic axons. Conductance increase in the synaptic membrane endures over most of the period of potential change, and it is this rather than the "electrical time constant" of the membrane that in large measure determines the form of the synaptic potential. Temporal summation of synaptic potential occurs during repetitive presynaptic stimulation, and after such stimulation the rate of decay of synaptic potential is greatly slowed.     

Cochlear Inner Hair Cell Ribbon Synapse is the Primary Target of Ototoxic Aminoglycoside Stimuli

The ribbon synapses of inner hair cells (IHCs) play an important role in sound encoding and neurotransmitter release. However, it remains unclear whether IHC ribbon synapse plasticity can be interrupted by ototoxic aminoglycoside stimuli. Here, we report that quantitative changes in the number of IHC ribbon synapses and hearing loss occur in response to gentamicin treatment in mice. Using 3D reconstruction, we were able to calculate the number of IHC ribbon synapses after ototoxic gentamicin exposure. Mice were injected intraperitoneally with a low dose of gentamicin (100 mg/kg) once a day for 14 days. Double immunostaining was used to identify IHC ribbon synapses; histopathology and scanning electron microscopy were used to observe the morphology of cochlear hair cells and spiral ganglion neurons (SGNs), the hearing threshold shifts were recorded by auditory brainstem response examinations. Our study shows that the maximal number of IHC ribbon synapses appeared at the 7th day after treatment, followed by a significant reduction after the 7th day regardless of ongoing treatment. Correspondingly, the maximal elevation of hearing threshold was observed at the 7th day after treatment. Meanwhile, additional cochlear components included OHCs, IHCs, and SGNs were unaffected, suggesting that IHC ribbon synapses are more susceptible to ototoxic aminoglycoside stimulation. Our study indicated that quantitative changes in the number of IHC ribbon synapses is critical response to lower dose of ototoxic stimulation, and may contribute to moderate hearing loss. Additionally, our data indcated that ribbon synaptic plasticity may require the quantitative changes to play self-protective role adapted to ototoxic aminoglycoside stimuli.     

Status epilepticus induced by pentylenetetrazole increases short-term synaptic facilitation in the hippocampus of juvenile rats

We studied the effect of status epilepticus (SE) on short-term synaptic plasticity. The amplitudes of field potentials in response to extracellular stimulation of the Schaffer collaterals were recorded in hippocampal slices. Subtle modifications were revealed on day 1 after SE, whereas on days 3 and 7 we did not find any differences from the control. These data show that, one day after SE, the probability of a transmitter release in hippocampal synapses decreases that serves as a compensatory mechanism, which prevents seizure activity.     

Synapses identifiable in the parietal ganglia of the snail Helix lucorum

The synaptic plasticity is a background for learning and memory. Identifiable synapses that are the synapses between individually identifiable neurons are a very convenient model for studying plasticity. Synapses between the interoceptive mechanosensory neurons and the command neurons of the withdrawal behavior were identified in the Helix lucorum brain. It was shown that synaptic plasticity estimated by the dynamics of the elementary postsynaptic potentials elicited by single presynaptic spikes differed from the synaptic plasticity estimated by the dynamics of compound synaptic responses of the same neurons to sensory stimulation. Habituation and heterosynaptic facilitation phenomena are discussed in terms of the dynamics of the elementary postsynaptic potentials.     

Increase in proportion of hippocampal spine synapses expressing neural cell adhesion molecule NCAM180 following long-term potentiation

Neural recognition molecules such as the neural cell adhesion molecule (NCAM) have been implicated in synaptic plasticity, including long-term potentiation (LTP), sensitization, and learning and memory. The major isoform of NCAM carrying the longest cytoplasmic domain of all NCAM isoforms (NCAM180) is predominantly localized in postsynaptic membranes and postsynaptic densities of hippocampal neurons, with only a proportion of synapses carrying detectable levels of NCAM180. To investigate whether this differential expression of NCAM180 may correlate with distinct states of synaptic activity, LTP was induced by high-frequency stimulation of the perforant path and the percentage of NCAM180 immunopositive spine synapses determined in the outer third of the dentate molecular layer of the dentate gyrus by immunoelectron microscopy. Twenty-four hours following induction of LTP by high-frequency stimulation, the percentage of spine synapses expressing NCAM180 increases from 37% (passive control) to 70%. This increase was inhibited by the noncompetitive N-methyl-D -aspartate receptor antagonist MK801. Following repeated LTP induction at 10 consecutive days with one tetanization each day, 60% of all spine synapses were NCAM180 immunoreactive. Compared to passive control animals, the percentage of NCAM180 expressing synapses in low-frequency stimulated animals decreased from 37% to 28%. Spine synapses in the inner part of the dentate molecular layer not contacted by the afferents of the perforant path did not change the percentage of NCAM180-expressing synapses. The results obtained by the postembedding immunogold staining technique confirmed the difference in NCAM180 expression of spine synapses between passive control and potentiated animals. These observations suggest a role for NCAM180 in synaptic remodeling accompanying LTP. © 1998 John Wiley & Sons, Inc. J Neurobiol 37: 359–372, 1998