Brain circuitry outside the synaptic cleft

A growing body of experimental evidence suggests that astroglia, and possibly microglia, play an important part in regulating synaptic networking of the brain. It has also emerged that extracellular matrix (ECM) structures that enwrap synaptic connections can generate molecular signals affecting both neuronal and glial activity. Thus it appears that the mechanism of information processing in the brain, which has hitherto been associated almost exclusively with neural circuits, could also involve informative signal exchange outside the synaptic cleft. In this Theme Issue, research teams including leading experts on astroglia–neuron communication and on ECM signalling report their recent findings, share their views and discuss future conceptual advances in the field. Potential implications for drug development and new therapeutic targets with regard to some common neurological conditions are discussed throughout the issue.     

Comparison of fast and slow synaptic terminals in lobster muscle

Summary Synaptic terminals of fast (FCE) and slow (SCE) excitatory neurons were physiologically identified on separate fibres of one muscle, the closer muscle in lobster claws. The innervation by these identified fibers was demonstrated over long distances (7–21 m) by examining serial thin sections at periodic intervals. The ultrastructure of each type of innervation was consistent both qualitatively and quantitatively in two separate samples. The FCE innervation is relatively simple in having consistently small-diameter terminals each forming a single long synapse, with few synaptic vesicles, and little if any postsynaptic apparatus. The SCE innervation is more complex in having larger-diameter but more variable terminals forming several short synapses, with many synaptic vesicles and an extensive postsynaptic apparatus. These differences in the size of the synapses and the number of synaptic vesicles parallel differences in transmitter release and fatigue sensitivity characteristic of the two types of innervation. The degree of elaboration of the postsynaptic apparatus may reflect differences in the amount of transmitter taken up after release. Our data reveal for the first time in a single muscle differences between FCE and SCE innervation previously reported in different muscles and in different species.Supported by grants from NIH (NINCDS) to A.G. Humes and the late Fred Lang and from NSERC and Muscular Dystrophy Assoc. of Canada to C.K. GovindWe thank Lena Hill for her technical expertise and critical evaluation of the study, and Dr. A.G. Humes for providing research facilities     

The role of microglia in synaptic stripping and synaptic degeneration: a revised perspective

Chronic neurodegenerative diseases of the CNS (central nervous system) are characterized by the loss of neurons. There is, however, growing evidence to show that an early stage of this process involves degeneration of presynaptic terminals prior to the loss of the cell body. Synaptic plasticity in CNS pathology has been associated with microglia and the phenomenon of synaptic stripping. We review here the evidence for the involvement of microglia in synaptic stripping and synapse degeneration and we conclude that this is a case of guilt by association. In disease models of chronic neurodegeneration, there is no evidence that microglia play an active role in either synaptic stripping or synapse degeneration, but the degeneration of the synapse and the envelopment of a degenerating terminal appears to be a neuron autonomous event. We highlight here some of the gaps in our understanding of synapse degeneration in chronic neurodegenerative disease.     

Distinct molecular mechanisms control levels of synaptic F‐actin

Polymerization of filamentous (F)‐actin at the neuronal synapse plays an important role in neuronal function. However, the regulatory mechanisms controlling the levels of synaptic actin remain incompletely understood. Here, I used established pharmacological blockers to acutely disrupt the function of actin polymerization machinery, then quantified their effect on synaptic F‐actin levels. Synaptic F‐actin was modestly decreased by inhibition of Arp2/3‐dependent actin branching. Blockade of formin‐dependent actin elongation resulted in an Arp2/3‐dependent increase in synaptic actin that could be mimicked by limited actin depolymerization. Limited actin depolymerization was also sufficient to reverse a decrease in synaptic F‐actin caused by prolonged blockade of synaptic NMDA‐type glutamate receptors. These results suggest that interplay between different actin polymerization pathways may regulate synaptic actin dynamics.     

Colocalization of synapsin and actin during synaptic vesicle recycling

It has been hypothesized that in the mature nerve terminal, interactions between synapsin and actin regulate the clustering of synaptic vesicles and the availability of vesicles for release during synaptic activity. Here, we have used immunogold electron microscopy to examine the subcellular localization of actin and synapsin in the giant synapse in lamprey at different states of synaptic activity. In agreement with earlier observations, in synapses at rest, synapsin immunoreactivity was preferentially localized to a portion of the vesicle cluster distal to the active zone. During synaptic activity, however, synapsin was detected in the pool of vesicles proximal to the active zone. In addition, actin and synapsin were found colocalized in a dynamic filamentous cytomatrix at the sites of synaptic vesicle recycling, endocytic zones. Synapsin immunolabeling was not associated with clathrin-coated intermediates but was found on vesicles that appeared to be recycling back to the cluster. Disruption of synapsin function by microinjection of antisynapsin antibodies resulted in a prominent reduction of the cytomatrix at endocytic zones of active synapses. Our data suggest that in addition to its known function in clustering of vesicles in the reserve pool, synapsin migrates from the synaptic vesicle cluster and participates in the organization of the actin-rich cytomatrix in the endocytic zone during synaptic activity.     

The Drosophila epsin 1 is required for ubiquitin-dependent synaptic growth and function but not for synaptic vesicle recycling

The ubiquitin-proteasome system plays an important role in synaptic development and function. However, many components of this system, and how they act to affect synapses, are still not well understood. In this study, we use the Drosophila neuromuscular junction to study the in vivo function of Liquid facets (Lqf), a homolog of mammalian epsin 1. Our data show that Lqf plays a novel role in synapse development and function. Contrary to prior models, Lqf is not required for clathrin-mediated endocytosis of synaptic vesicles. Lqf is required to maintain bouton size and shape and to sustain synapse growth by acting as a specific substrate of the deubiquitinating enzyme Fat facets. However, Lqf is not a substrate of the Highwire (Hiw) E3 ubiquitin ligase; neither is it required for synapse overgrowth in hiw mutants. Interestingly, Lqf converges on the Hiw pathway by negatively regulating transmitter release in the hiw mutant. These observations demonstrate that Lqf plays distinct roles in two ubiquitin pathways to regulate structural and functional plasticity of the synapse.     

Substantial changes in synaptic firing frequencies induced by glial ATP hysteresis

Recent experimental studies strongly suggest the influence of glial purinergic transmission in the modulation of synaptic dynamics. By releasing adenosine triphosphate (ATP), which accumulates as adenosine, astrocytes tonically suppressed synaptic transmission. The delayed multi-step feedback of the glial ATP with the neuron suggests the existence of a discrete hysteresis phenomena. By integration of this hysteretic behavior into a delayed leaky integrate-and-fire model for the tripartite synapses, a significant sensitivity of the pre- and postsynaptic firing frequency patterns to the adenosine feedback-delays is observed that might be of importance for adenosine-related neurological deficits, such as sleep disorders.     

The synaptic action of Degenerin/Epithelial sodium channels

Degenerin/Epithelial Sodium Channels (DEG/ENaCs) are a large family of animal-specific non-voltage gated ion channels, with enriched expression in neuronal and epithelial tissues. While neuronal DEG/ENaCs were originally characterized as sensory receptor channels, recent studies indicate that several DEG/ENaC family members are also expressed throughout the central nervous system. Human genome-wide association studies have linked DEG/ENaC-coding genes with several neurologic and psychiatric disorders, including epilepsy and panic disorder. In addition, studies in rodent models further indicate that DEG/ENaC activity in the brain contributes to many behaviors, including those related to anxiety and long-term memory. Although the exact neurophysiological functions of DEG/ENaCs remain mostly unknown, several key studies now suggest that multiple family members might exert their neuronal function via the direct modulation of synaptic processes. Here, we review and discuss recent findings on the synaptic functions of DEG/ENaCs in both vertebrate and invertebrate species, and propose models for their possible roles in synaptic physiology.     

A mathematical model of the tripartite synapse: astrocyte-induced synaptic plasticity

In this paper, we present a biologically detailed mathematical model of tripartite synapses, where astrocytes modulate short-term synaptic plasticity. The model consists of a pre-synaptic bouton, a post-synaptic dendritic spine-head, a synaptic cleft and a peri-synaptic astrocyte controlling Ca^2 + dynamics inside the synaptic bouton. This in turn controls glutamate release dynamics in the cleft. As a consequence of this, glutamate concentration in the cleft has been modeled, in which glutamate reuptake by astrocytes has also been incorporated. Finally, dendritic spine-head dynamics has been modeled. As an application, this model clearly shows synaptic potentiation in the hippocampal region, i.e., astrocyte Ca^2 + mediates synaptic plasticity, which is in conformity with the majority of the recent findings (Perea and Araque (Science 317, 1083–1086, 2007); Henneberger et al. (Nature 463, 232–236, 2010); Navarrete et al. (PLoS Biol. 10, e1001259, 2012)).

Electronic supplementary material

The online version of this article (doi:10.1007/s10867-012-9267-7) contains supplementary material, which is available to authorized users.     

Differences in synaptic output between excitatory and inhibitory motoneurons in a crayfish muscle

A pair of antagonistic motoneurons, one excitatory and one inhibitory, innervates the distal accessory flexor muscle in the walking limb of the crayfish Procambarus clarkii. The number and size of synapses formed by these two axons on the muscle fibers (neuromuscular synapses) and on each other (axo-axonal synapses) were estimated using thin-section electron microscopy. Although profiles of nerve terminals of the two axons occur in roughly equal proportions, the frequency of occurrence of neuromuscular synapses differed markedly: 73% were excitatory and 27% were inhibitory. However, inhibitory synapses were 4–5 times larger than excitatory ones, and consequently, the total contact areas devoted to neuromuscular synapses were similar for both axons. Axo-axonal synapses were predominantly from the inhibitory axon to the excitatory axon (86%), and a few were from the excitatory axon to the inhibitory axon (14%). The role of the inhibitory axo-axonal synapse is presynaptic inhibition, but that of the excitatory axo-axonal synapse is not known. The differences in size of neuromuscular synapses between the two axons may reflect intrinsic determinants of the neuron, while the similarity in total synaptic area may reflect retrograde influences from the muscle for regulating synapse number.     

The organization of synaptic vesicles at tonically transmitting connections of locust visual interneurons

Large, second‐order neurons of locust ocelli, or L‐neurons, make some output connections that transmit small changes in membrane potential and can sustain transmission tonically. The synaptic connections are made from the axons of L‐neurons in the lateral ocellar tracts, and are characterized by bar‐shaped presynaptic densities and densely packed clouds of vesicles near to the cell membrane. A cloud of vesicles can extend much of the length of this synaptic zone, and there is no border between the vesicles that are associated with neighboring presynaptic densities. In some axons, presynaptic densities are associated with discrete small clusters of vesicles. Up to 6% of the volume of a length of axon in a synaptic zone can be occupied with a vesicle cloud, packed with 4.5 to 5.5 thousand vesicles per μm³. Presynaptic densities vary in length, from less than 70 nm to 1.5 μm, with shorter presynaptic densities being most frequent. The distribution of vesicles around short presynaptic densities was indistinguishable from that around long presynaptic densities, and vesicles were distributed in a similar way right along the length of a presynaptic density. Within the cytoplasm, vesicles are homogeneously distributed within a cloud. We found no differences in the distribution of vesicles in clouds between locusts that had been dark‐adapted and locusts that had been light‐adapted before fixation. © 2002 Wiley Periodicals, Inc. J Neurobiol 50: 93–105, 2002; DOI 10.1002/neu.10018     

PSD-95 promotes the stabilization of young synaptic contacts

Maintaining a population of stable synaptic connections is probably of critical importance for the preservation of memories and functional circuitry, but the molecular dynamics that underlie synapse stabilization is poorly understood. Here, we use simultaneous time-lapse imaging of post synaptic density-95 (PSD-95) and Ca²⁺/calmodulin-dependent protein kinase II (CaMKII) to investigate the dynamics of protein composition at axodendritic (AD) contacts. Our data reveal that this composition is highly dynamic, with both proteins moving into and out of the same synapse independently, so that synapses cycle rapidly between states in which they are enriched for none, one or both proteins. We assessed how PSD-95 and CaMKII interact at stable and transient AD sites and found that both phospho-CaMKII and PSD-95 are present more often at stable than labile contacts. Finally, we found that synaptic contacts are more stable in older neurons, and this process can be mimicked in younger neurons by overexpression of PSD-95. Taken together, these data show that synaptic protein composition is highly variable over a time-scale of hours, and that PSD-95 is probably a key synaptic protein that promotes synapse stability.     

Rapid neural circuit switching mediated by synaptic plasticity during neural morphallactic regeneration

The aquatic oligochaete, Lumbriculus variegatus (Lumbriculidae), undergoes a rapid regenerative transformation of its neural circuits following body fragmentation. This type of nervous system plasticity, called neural morphallaxis, involves the remodeling of the giant fiber pathways that mediate rapid head and tail withdrawal behaviors. Extra- and intracellular electrophysiological recordings demonstrated that changes in cellular properties and synaptic connections underlie neurobehavioral plasticity during morphallaxis. Sensory-to-giant interneuron connections, undetectable prior to body injury, emerged within hours of segment amputation. The appearance of functional synaptic transmission was followed by interneuron activation, coupling of giant fiber spiking to motor outputs and overt segmental shortening. The onset of morphallactic plasticity varied along the body axis and emerged more rapidly in segments closer to regions of sensory field overlap between the two giant fiber pathways. The medial and lateral giant fibers were simultaneously activated during a transient phase of network remodeling. Thus, synaptic plasticity at sensory-to-giant interneuron connections mediates escape circuit morphallaxis in this regenerating annelid worm.     

C-terminal synaptic targeting elements for postsynaptic density proteins ProSAP1/Shank2 and ProSAP2/Shank3

Synapses are specialized contact sites mediating communication between neurons. Synaptogenesis requires the specific assembly of protein clusters at both sides of the synaptic contact by mechanisms that are barely understood. We studied the synaptic targeting of multi-domain proteins of the ProSAP/Shank family thought to serve as master scaffolding molecules of the postsynaptic density. In contrast to Shank1, expression of green-fluorescent protein (GFP)-tagged ProSAP1/Shank2 and ProSAP2/Shank3 deletion constructs in hippocampal neurons revealed that their postsynaptic localization relies on the integrity of the C-termini. The shortest construct that was perfectly targeted to synaptic sites included the last 417 amino acids of ProSAP1/Shank2 and included the C-terminal sterile alpha motif (SAM) domain. Removal of 54 residues from the N-terminus of this construct resulted in a diffuse distribution in the cytoplasm. Altogether, our data delineate a hitherto unknown targeting signal in both ProSAP1/Shank2 and ProSAP2/Shank3 and provide evidence for an implication of these proteins and their close homologue, Shank1, in distinct molecular pathways.     

Population density regulates Drosophila synaptic morphology in a Fasciclin‐II‐dependent manner

Genetic analysis of the Drosophila larval neuromuscular junction has identified some of the key molecules that regulate synaptic plasticity. Among these molecules, the expression level of Fasciclin II (FasII), a homophilic cell adhesion molecule, is critically important for determining the final form of the neuromuscular junction. Genetic reduction of FasII expression by 50% yields more elaborate nerve terminals, while a greater reduction in expression, to 10% of wild‐type, yields a substantial reduction in the nerve terminal morphology. Importantly, regulation of FasII expression seems to be the final output for several genetic manipulations that transform NMJ morphology. In an effort to understand the importance of this regulatory pathway in the normal animal, we have undertaken studies to identify environmental cues that might be important for initiating FasII‐dependent changes in synaptic plasticity. Here we report on the relationship between larval population density and synaptic morphology, synaptic strength, and FasII levels. We raised Drosophila larvae under conditions of increasing population density and found an inverse exponential relationship between population density and the number of synaptic boutons, the number of branches, and the length of branches. We also observed population‐dependent alteration in FasII levels, with lower densities having less FasII at the synapse. The correlation between density and morphological change was abrogated in larvae constitutively expressing FasII, and in wild‐type larvae grown on soft culture medium. Together these data show that environmental cues can induce regulation of FasII. Interestingly, however, the quantal content of synaptic transmission was not different among the different population densities, suggesting that other factors contribute to maintaining synaptic strength at a defined level. © 2004 Wiley Periodicals, Inc. J Neurobiol, 2004     

Age‐related differences in synaptic plasticity following muscle unloading

The objective of the present investigation was to determine the effects of muscle unloading—a form of subtotal disuse— on the morphology of the neuromuscular junction (NMJ) in younger and aged animals. Sixteen aged (22 months) and 16 young adult (8 months) male Fischer 344 rats were assigned to control and hindlimb suspension (HS) conditions (n = 8/group). At the conclusion of the 4 week experimental period, soleus muscles were collected, and immunofluorescent procedures were used to visualize acetylcholine (ACh) vesicles and receptors, nerve terminal branching, as well as NCAM and NT‐4 expression. Quantitative analyses revealed that aged controls displayed significant (p < 0.05) reductions in area and perimeter length of ACh vesicle and receptor regions, without affecting nerve terminal branch number or length. In contrast to younger NMJs, which were resilient to the effects of unloading, NMJs of aged HS rats demonstrated significant expansion of ACh vesicle and receptor dimensions compared to aged controls. Qualitative analyses of NCAM staining indicated that aging alone somewhat increased this molecule's expression (aged controls > young controls). Among the four groups, however, the greatest amount of NCAM content was detected among aged HS muscles, matching the degree of synaptic plasticity exhibited in those muscles. Unlike NCAM, the expression of NT‐4 did not appear to differ among the treatment groups. These data suggest that although young adult muscle maintains normal NMJ structure during prolonged exposure to unloading, aged NMJs experience significant adaptation to that stimulus. © 2003 Wiley Periodicals, Inc. J Neurobiol 57: 246–256, 2003     

Syntaxin-1通过激活突触递质传递加速早期突触的形成

Syntaxin-1是一种多结构域蛋白,通过与synaptobrevin-2和SNAP-25形成SNARE复合体调节囊泡融合.然而,syntaxin-1在突触形成过程中是否发挥作用,目前尚不清楚.本研究显示syntaxin-1的表达水平与突触形成过程高度相关.Syntaxin-1的R151A和I155A突变影响其在突触形成中的促进作用,而Habc结构域或跨膜结构域在突触形成中无显著作用.结果表明,syntaxin-1通过激活突触囊泡释放来加速突触的形成.     

pH changes in the invaginating synaptic cleft mediate feedback from horizontal cells to cone photoreceptors by modulating Ca2+ channels

Feedback from horizontal cells (HCs) to cone photoreceptors plays a key role in the center-surround-receptive field organization of retinal neurons. Recordings from cone photoreceptors in newt retinal slices were obtained by the whole-cell patch-clamp technique, using a superfusate containing a GABA antagonist (100 microM picrotoxin). Surround illumination of the receptive field increased the voltage-dependent calcium current (ICa) in the cones, and shifted the activation voltage of ICa to negative voltages. External alkalinization also increased cone ICa and shifted its activation voltage toward negative voltages. Enrichment of the pH buffering capacity of the extracellular solution increased cone ICa, and blocked any additional increase in cone ICa by surround illumination. Hyperpolarization of the HCs by a glutamate receptor antagonist-augmented cone ICa, whereas depolarization of the HCs by kainate suppressed cone ICa. From these results, we propose the hypothesis that pH changes in the synaptic clefts, which are intimately related to the membrane voltage of the HCs, mediate the feedback from the HCs to cone photoreceptors. The feedback mediated by pH changes in the synaptic cleft may serve as an additional mechanism for the center-surround organization of the receptive field in the outer retina.     

Emerging roles for semaphorins and VEGFs in synaptogenesis and synaptic plasticity

Synapse formation, maintenance and plasticity are critical for the correct function of the nervous system and its target organs. During development, these processes enable the establishment of appropriate neural circuits. During adulthood, they allow adaptation to both physiological and environmental changes. In this review, we discuss emerging roles for two families of classical axon and vascular guidance cues in synaptogenesis and synaptic plasticity, the semaphorins and the vascular endothelial growth factors (VEGFs). Their contribution to synapse formation and function add a new facet to the spectrum of overlapping and complementary roles for these molecules in development, adulthood and disease.