Number Of Active Zones Research Articles

Maintenance of a central high frequency synapse in the absence of synaptic activity.

Activity has long been considered essential for circuit formation and maintenance. This view has recently been challenged by proper synaptogenesis and only mildly affected synapse maintenance in the absence of synaptic activity in forebrain neurons. Here, we investigated whether synaptic activity is necessary for the development and maintenance of the calyx of Held synapse. This giant synapse located in the auditory brainstem is highly specialized to maintain high frequency, high-fidelity synaptic transmission for prolonged times and thus shows particularly high synaptic activity. We expressed the protease tetanus toxin light chain (TeNT) exclusively in bushy cells of the ventral cochlear nucleus (VCN) of juvenile mice. Since globular bushy cells give rise to the calyx of Held, expression of TeNT in these cells specifically abolished synaptic transmission at the calyx without impairing general functionality of the central auditory system. Calyces lacked synaptic activity after two weeks of TeNT expression. However, this did not lead to major changes in presynaptic morphology, the number of active zones (AZs) or the composition of postsynaptic AMPA-type glutamate receptors (GluAs). Moreover, the fenestration of the calyx of Held, a hallmark of structural maturation, occurred normally. We thus show that the maintenance of a specialized high frequency synapse in the auditory brainstem occurs in a hardwired, probably genetically encoded, manner with little dependence on synaptic activity.

Simulations of active zone structure and function at mammalian NMJs predict that loss of calcium channels alone is not sufficient to replicate LEMS effects.

Lambert-Eaton myasthenic syndrome (LEMS) is an autoimmune-mediated neuromuscular disease thought to be caused by autoantibodies against P/Q-type voltage-gated calcium channels (VGCCs), which attack and reduce the number of VGCCs within transmitter release sites (active zones; AZs) at the neuromuscular junction (NMJ), resulting in neuromuscular weakness. However, patients with LEMS also have antibodies to other neuronal proteins, and about 15% of patients with LEMS are seronegative for antibodies against VGCCs. We hypothesized that a reduction in the number of P/Q-type VGCCs alone is not sufficient to explain LEMS effects on transmitter release. Here, we used a computational model to study a variety of LEMS-mediated effects on AZ organization and transmitter release constrained by electron microscopic, pharmacological, immunohistochemical, voltage imaging, and electrophysiological observations. We show that models of healthy AZs can be modified to predict the transmitter release and short-term facilitation characteristics of LEMS and that in addition to a decrease in the number of AZ VGCCs, disruption in the organization of AZ proteins, a reduction in AZ number, a reduction in the amount of synaptotagmin, and the compensatory expression of L-type channels outside the remaining AZs are important contributors to LEMS-mediated effects on transmitter release. Furthermore, our models predict that antibody-mediated removal of synaptotagmin in combination with disruption in AZ organization alone could mimic LEMS effects without the removal of VGCCs (a seronegative model). Overall, our results suggest that LEMS pathophysiology may be caused by a collection of pathological alterations to AZs at the NMJ, rather than by a simple loss of VGCCs.NEW & NOTEWORTHY We used a computational model of the active zone (AZ) in the mammalian neuromuscular junction to investigate Lambert-Eaton myasthenic syndrome (LEMS) pathophysiology. This model suggests that disruptions in presynaptic active zone organization and protein content (particularly synaptotagmin), beyond the simple removal of presynaptic calcium channels, play an important role in LEMS pathophysiology.

Activity-regulated growth of motoneurons at the neuromuscular junction is mediated by NADPH oxidases.

Neurons respond to changes in the levels of activity they experience in a variety of ways, including structural changes at pre- and postsynaptic terminals. An essential plasticity signal required for such activity-regulated structural adjustments are reactive oxygen species (ROS). To identify sources of activity-regulated ROS required for structural plasticity in vivo we used the Drosophila larval neuromuscular junction as a highly tractable experimental model system. For adjustments of presynaptic motor terminals, we found a requirement for both NADPH oxidases, Nox and dual oxidase (Duox), that are encoded in the Drosophila genome. This contrasts with the postsynaptic dendrites from which Nox is excluded. NADPH oxidases generate ROS to the extracellular space. Here, we show that two aquaporins, Bib and Drip, are necessary ROS conduits in the presynaptic motoneuron for activity regulated, NADPH oxidase dependent changes in presynaptic motoneuron terminal growth. Our data further suggest that different aspects of neuronal activity-regulated structural changes might be regulated by different ROS sources: changes in bouton number require both NADPH oxidases, while activity-regulated changes in the number of active zones might be modulated by other sources of ROS. Overall, our results show NADPH oxidases as important enzymes for mediating activity-regulated plasticity adjustments in neurons.

EFFECT OF GLYCOLYSIS SUPRESSION ON ULTRASTRUCTURE OF THE HIPPOCAMPAL GIANT SYNAPSES

Suppression of glycolysis in the rat hippocampal formations was induced by daily intracerebroventricular injection of 2-deoxy-D-glucose for 10 days. Using electron microscopy, a disturbance of morphological interactions of axonal terminals of granular neurons with perisynaptic astrocytic processes was found. At the same time, both the determinant ultrastructural characteristics of giant synapses and two types of functional contacts (asymmetric chemical active zones with dendritic spines and symmetric adhesive junctions with dendrites) were retained. However, they showed morphological signs of a weakening of synaptic processes, which were expressed in a decrease in the number of intraterminal active zones, clustering of synaptic vesicles, and their removal from neurotransmitter release sites. Compared to the norm, an increase in the number of small mitochondria with an electron-dense matrix and the evidence of initiation of glycogen synthesis in the form of glycosomes were found in the giant terminals. The data obtained indicate that the giant synapses of the hippocampal mossy fibers are a plastic, self-correcting system that is able to function by adapting its own bioenergetic metabolism, when brain glycolysis is suppressed.

Identification of genes regulating stimulus-dependent synaptic assembly in Drosophila using an automated synapse quantification system.

Neural activity-dependent synaptic plasticity is an important physiological phenomenon underlying environmental adaptation, memory and learning. However, its molecular basis, especially in presynaptic neurons, is not well understood. Previous studies have shown that the number of presynaptic active zones in the Drosophila melanogaster photoreceptor R8 is reversibly changed in an activity-dependent manner. During reversible synaptic changes, both synaptic disassembly and assembly processes were observed. Although we have established a paradigm for screening molecules involved in synaptic stability and several genes have been identified, genes involved in stimulus-dependent synaptic assembly are still elusive. Therefore, the aim of this study was to identify genes regulating stimulus-dependent synaptic assembly in Drosophila using an automated synapse quantification system. To this end, we performed RNAi screening against 300 memory-defective, synapse-related or transmembrane molecules in photoreceptor R8 neurons. Candidate genes were narrowed down to 27 genes in the first screen using presynaptic protein aggregation as a sign of synaptic disassembly. In the second screen, we directly quantified the decreasing synapse number using a GFP-tagged presynaptic protein marker. We utilized custom-made image analysis software, which automatically locates synapses and counts their number along individual R8 axons, and identified cirl as a candidate gene responsible for synaptic assembly. Finally, we present a new model of stimulus-dependent synaptic assembly through the interaction of cirl and its possible ligand, ten-a. This study demonstrates the feasibility of using the automated synapse quantification system to explore activity-dependent synaptic plasticity in Drosophila R8 photoreceptors in order to identify molecules involved in stimulus-dependent synaptic assembly.

The human cognition-enhancing CORD7 mutation increases active zone number and synaptic release.

Humans carrying the CORD7 (cone-rod dystrophy 7) mutation possess increased verbal IQ and working memory. This autosomal dominant syndrome is caused by the single-amino acid R844H exchange (human numbering) located in the 310 helix of the C2A domain of RIMS1/RIM1 (Rab3-interacting molecule 1). RIM is an evolutionarily conserved multi-domain protein and essential component of presynaptic active zones, which is centrally involved in fast, Ca2+-triggered neurotransmitter release. How the CORD7 mutation affects synaptic function has remained unclear thus far. Here, we established Drosophila melanogaster as a disease model for clarifying the effects of the CORD7 mutation on RIM function and synaptic vesicle release. To this end, using protein expression and X-ray crystallography, we solved the molecular structure of the Drosophila C2A domain at 1.92 Å resolution and by comparison to its mammalian homologue ascertained that the location of the CORD7 mutation is structurally conserved in fly RIM. Further, CRISPR/Cas9-assisted genomic engineering was employed for the generation of rim alleles encoding the R915H CORD7 exchange or R915E, R916E substitutions (fly numbering) to effect local charge reversal at the 310 helix. Through electrophysiological characterization by two-electrode voltage clamp and focal recordings we determined that the CORD7 mutation exerts a semi-dominant rather than a dominant effect on synaptic transmission resulting in faster, more efficient synaptic release and increased size of the readily releasable pool but decreased sensitivity for the fast calcium chelator BAPTA. In addition, the rim CORD7 allele increased the number of presynaptic active zones but left their nanoscopic organization unperturbed as revealed by super-resolution microscopy of the presynaptic scaffold protein Bruchpilot/ELKS/CAST. We conclude that the CORD7 mutation leads to tighter release coupling, an increased readily releasable pool size and more release sites thereby promoting more efficient synaptic transmitter release. These results strongly suggest that similar mechanisms may underlie the CORD7 disease phenotype in patients and that enhanced synaptic transmission may contribute to their increased cognitive abilities.

Enhancement of wavefront measurement sensitivity in a zonal wavefront sensor without curtailing the sensing speed

In this paper we propose a zonal wavefront sensing scheme that facilitates wavefront measurement with enhanced sensitivity at the standard video rate. We achieve this enhanced sensitivity by implementing a sequential display of binary holograms described over each zone sampling the incident wavefront with the help of a ferroelectric liquid crystal spatial light modulator. By keeping the number of active zones as 24 and using a camera with an imaging frame rate equal to the binary hologram display rate of the spatial light modulator, we are able to reach a sensing frame rate of 60 Hz. In addition to enhanced sensitivity, the proposed scheme facilitates zone-wise tuning of binary holograms and eliminates the possibility of any crosstalk between adjacent zones. We perform a proof-of-principle experiment that validates the proposed zonal wavefront sensing scheme and demonstrates its advantages.

The ubiquitin ligase Ariadne-1 regulates neurotransmitter release via ubiquitination of NSF

Ariadne-1 (Ari-1) is an E3 ubiquitin-ligase essential for neuronal development, but whose neuronal substrates are yet to be identified. To search for putative Ari-1 substrates, we used an in vivo ubiquitin biotinylation strategy coupled to quantitative proteomics of Drosophila heads. We identified 16 candidates that met the established criteria: a significant change of at least twofold increase on ubiquitination, with at least two unique peptides identified. Among those candidates, we identified Comatose (Comt), the homologue of the N-ethylmaleimide sensitive factor (NSF), which is involved in neurotransmitter release. Using a pull-down approach that relies on the overexpression and stringent isolation of a GFP-fused construct, we validate Comt/NSF to be an ubiquitination substrate of Ari-1 in fly neurons, resulting in the preferential monoubiquitination of Comt/NSF. We tested the possible functional relevance of this modification using Ari-1 loss-of-function mutants, which displayed a lower rate of spontaneous neurotransmitter release due to failures at the presynaptic side. By contrast, evoked release in Ari-1 mutants was enhanced compared with controls in a Ca2+-dependent manner without modifications in the number of active zones, indicating that the probability of release per synapse is increased in these mutants. This phenotype distinction between spontaneous and evoked release suggests that NSF activity may discriminate between these two types of vesicle fusion. Our results thus provide a mechanism to regulate NSF activity in the synapse through Ari-1-dependent ubiquitination.

Synaptic Organization of the Human Temporal Lobe Neocortex as Revealed by High-Resolution Transmission, Focused Ion Beam Scanning, and Electron Microscopic Tomography.

Modern electron microscopy (EM) such as fine-scale transmission EM, focused ion beam scanning EM, and EM tomography have enormously improved our knowledge about the synaptic organization of the normal, developmental, and pathologically altered brain. In contrast to various animal species, comparably little is known about these structures in the human brain. Non-epileptic neocortical access tissue from epilepsy surgery was used to generate quantitative 3D models of synapses. Beside the overall geometry, the number, size, and shape of active zones and of the three functionally defined pools of synaptic vesicles representing morphological correlates for synaptic transmission and plasticity were quantified. EM tomography further allowed new insights in the morphological organization and size of the functionally defined readily releasable pool. Beside similarities, human synaptic boutons, although comparably small (approximately 5 µm), differed substantially in several structural parameters, such as the shape and size of active zones, which were on average 2 to 3-fold larger than in experimental animals. The total pool of synaptic vesicles exceeded that in experimental animals by approximately 2 to 3-fold, in particular the readily releasable and recycling pool by approximately 2 to 5-fold, although these pools seemed to be layer-specifically organized. Taken together, synaptic boutons in the human temporal lobe neocortex represent unique entities perfectly adapted to the “job” they have to fulfill in the circuitry in which they are embedded. Furthermore, the quantitative 3D models of synaptic boutons are useful to explain and even predict the functional properties of synaptic connections in the human neocortex.

Variants in SCAF4 Cause a Neurodevelopmental Disorder and Are Associated with Impaired mRNA Processing

RNA polymerase II interacts with various other complexes and factors to ensure correct initiation, elongation, and termination of mRNA transcription. One of these proteins is SR-related CTD-associated factor 4 (SCAF4), which is important for correct usage of polyA sites for mRNA termination. Using exome sequencing and international matchmaking, we identified nine likely pathogenic germline variants in SCAF4 including two splice-site and seven truncating variants, all residing in the N-terminal two thirds of the protein. Eight of these variants occurred de novo, and one was inherited. Affected individuals demonstrated a variable neurodevelopmental disorder characterized by mild intellectual disability, seizures, behavioral abnormalities, and various skeletal and structural anomalies. Paired-end RNA sequencing on blood lymphocytes of SCAF4-deficient individuals revealed a broad deregulation of more than 9,000 genes and significant differential splicing of more than 2,900 genes, indicating an important role of SCAF4 in mRNA processing. Knockdown of the SCAF4 ortholog CG4266 in the model organism Drosophila melanogaster resulted in impaired locomotor function, learning, and short-term memory. Furthermore, we observed an increased number of active zones in larval neuromuscular junctions, representing large glutamatergic synapses. These observations indicate a role of CG4266 in nervous system development and function and support the implication of SCAF4 in neurodevelopmental phenotypes. In summary, our data show that heterozygous, likely gene-disrupting variants in SCAF4 are causative for a variable neurodevelopmental disorder associated with impaired mRNA processing.

Synaptic Plasticity Induced by Differential Manipulation of Tonic and Phasic Motoneurons in Drosophila.

Structural and functional plasticity induced by neuronal competition is a common feature of developing nervous systems. However, the rules governing how postsynaptic cells differentiate between presynaptic inputs are unclear. In this study, we characterized synaptic interactions following manipulations of tonic Ib or phasic Is glutamatergic motoneurons that coinnervate postsynaptic muscles of male or female Drosophila melanogaster larvae. After identifying drivers for each neuronal subtype, we performed ablation or genetic manipulations to alter neuronal activity and examined the effects on synaptic innervation and function at neuromuscular junctions. Ablation of either Ib or Is resulted in decreased muscle response, with some functional compensation occurring in the Ib input when Is was missing. In contrast, the Is terminal failed to show functional or structural changes following loss of the coinnervating Ib input. Decreasing the activity of the Ib or Is neuron with tetanus toxin light chain resulted in structural changes in muscle innervation. Decreased Ib activity resulted in reduced active zone (AZ) number and decreased postsynaptic subsynaptic reticulum volume, with the emergence of filopodial-like protrusions from synaptic boutons of the Ib input. Decreased Is activity did not induce structural changes at its own synapses, but the coinnervating Ib motoneuron increased the number of synaptic boutons and AZs it formed. These findings indicate that tonic Ib and phasic Is motoneurons respond independently to changes in activity, with either functional or structural alterations in the Ib neuron occurring following ablation or reduced activity of the coinnervating Is input, respectively.SIGNIFICANCE STATEMENT Both invertebrate and vertebrate nervous systems display synaptic plasticity in response to behavioral experiences, indicating that underlying mechanisms emerged early in evolution. How specific neuronal classes innervating the same postsynaptic target display distinct types of plasticity is unclear. Here, we examined whether Drosophila tonic Ib and phasic Is motoneurons display competitive or cooperative interactions during innervation of the same muscle, or compensatory changes when the output of one motoneuron is altered. We established a system to differentially manipulate the motoneurons and examined the effects of cell type-specific changes to one of the inputs. Our findings indicate Ib and Is motoneurons respond differently to activity mismatch or loss of the coinnervating input, with the Ib subclass responding robustly compared with Is motoneurons.

CTGF/CCN2 facilitates LRP4-mediated formation of the embryonic neuromuscular junction.

At the neuromuscular junction (NMJ), lipoprotein-related receptor 4 (LRP4) mediates agrin-induced MuSK phosphorylation that leads to clustering of acetylcholine receptors (AChRs) in the postsynaptic region of the skeletal muscle. Additionally, the ectodomain of LRP4 is necessary for differentiation of the presynaptic nerve terminal. However, the molecules regulating LRP4 have not been fully elucidated yet. Here, we show that the CT domain of connective tissue growth factor (CTGF/CCN2) directly binds to the third beta-propeller domain of LRP4. CTGF/CCN2 enhances the binding of LRP4 to MuSK and facilitates the localization of LRP4 on the plasma membrane. CTGF/CCN2 enhances agrin-induced MuSK phosphorylation and AChR clustering in cultured myotubes. Ctgf-deficient mouse embryos (Ctgf-/- ) have small AChR clusters and abnormal dispersion of synaptic vesicles along the motor axon. Ultrastructurally, the presynaptic nerve terminals have reduced numbers of active zones and mitochondria. Functionally, Ctgf-/- embryos exhibit impaired NMJ signal transmission. These results indicate that CTGF/CCN2 interacts with LRP4 to facilitate clustering of AChRs at the motor endplate and the maturation of the nerve terminal.

Proportional Downscaling of Glutamatergic Release Sites by the General Anesthetic Propofol atDrosophilaMotor Nerve Terminals

Propofol is the most common general anesthetic used for surgery in humans, yet its complete mechanism of action remains elusive. In addition to potentiating inhibitory synapses in the brain, propofol also impairs excitatory neurotransmission. We use electrophysiological recordings from individual glutamatergic boutons in male and female larval Drosophila melanogaster motor nerve terminals to characterize this effect. We recorded from two bouton types, which have distinct presynaptic physiology and different average numbers of release sites or active zones. We show that a clinically relevant dose of propofol (3 μm) impairs neurotransmitter release similarly at both bouton types by decreasing the number of active release sites by half, without affecting release probability. In contrast, an analog of propofol has no effect on glutamate release. Coexpressing a truncated syntaxin1A protein in presynaptic boutons completely blocked this effect of propofol. Overexpressing wild-type syntaxin1A in boutons also conferred a level of resistance by increasing the number of active release sites to a physiological ceiling set by the number of active zones or T-bars, and in this way counteracting the effect of propofol. These results point to the presynaptic release machinery as a target for the general anesthetic. Proportionally equivalent effects of propofol on the number of active release sites across the different bouton types suggests that glutamatergic circuits that involve smaller boutons with fewer release sites may be more vulnerable to the presynaptic effects of the drug.

Production of modified basalt fibre for heat-insulating products manufacturing

As a result of the research flexible heat-insulating materials were developed based on basalt fiber with increased effectiveness, which can be achieved due to directed fiber microstructure forming through fusion modification. It is known that chemical composition of initial fusion not identically influences on physical and chemical and mechanical properties of basaltic fibres. Structural descriptions of fusion and basaltic fibre got from him appear main factors the nearer, than high speed of cooling. This index mainly depends on such constituents: a) ambient temperatures; b) coefficient of heat conducting; c) heat conducting of fusion; d) areas of surface. By researches influence of temperature of basaltic fusion was shown on the structure of fibres. Structural characteristics of the basalt fiber (number of active zones, coefficient of its distribution on the basalt fiber surface, as well as a correlation of three groups of active zones) depend on rheological properties of basal fusion, speed of drawing through the die plate and cooling speed.

The p75NTR neurotrophin receptor is required to organize the mature neuromuscular synapse by regulating synaptic vesicle availability

The coordinated movement of organisms relies on efficient nerve-muscle communication at the neuromuscular junction. After peripheral nerve injury or neurodegeneration, motor neurons and Schwann cells increase the expression of the p75NTR pan-neurotrophin receptor. Even though p75NTR targeting has emerged as a promising therapeutic strategy to delay peripheral neuronal damage progression, the effects of long-term p75NTR inhibition at the mature neuromuscular junction have not been elucidated. We performed quantitative neuroanathomical analyses of the neuromuscular junction in p75NTR null mice by laser confocal and electron microscopy, which were complemented with electromyography, locomotor tests, and pharmacological intervention studies. Mature neuromuscular synapses of p75NTR null mice show impaired postsynaptic organization and ultrastructural complexity, which correlate with altered synaptic function at the levels of nerve activity-induced muscle responses, muscle fiber structure, force production, and locomotor performance. Our results on primary myotubes and denervated muscles indicate that muscle-derived p75NTR does not play a major role on postsynaptic organization. In turn, motor axon terminals of p75NTR null mice display a strong reduction in the number of synaptic vesicles and active zones. According to the observed pre and postsynaptic defects, pharmacological acetylcholinesterase inhibition rescued nerve-dependent muscle response and force production in p75NTR null mice. Our findings revealing that p75NTR is required to organize mature neuromuscular junctions contribute to a comprehensive view of the possible effects caused by therapeutic attempts to target p75NTR.

Neurons regulate synaptic strength through homeostatic scaling of active zones.

How neurons stabilize their overall synaptic strength following conditions that alter synaptic morphology or function is a key question in neuronal homeostasis. In this issue, Goel et al. (2019. J. Cell Biol. https://doi.org/10.1083/jcb.201807165) find that neurons stabilize synaptic output despite disruptions in synapse size, active zone number, or postsynaptic function by controlling the delivery of active zone material and active zone size.

Homeostatic scaling of active zone scaffolds maintains global synaptic strength.

Synaptic terminals grow and retract throughout life, yet synaptic strength is maintained within stable physiological ranges. To study this process, we investigated Drosophila endophilin (endo) mutants. Although active zone (AZ) number is doubled in endo mutants, a compensatory reduction in their size homeostatically adjusts global neurotransmitter output to maintain synaptic strength. We find an inverse adaptation in rab3 mutants. Additional analyses using confocal, STED, and electron microscopy reveal a stoichiometric tuning of AZ scaffolds and nanoarchitecture. Axonal transport of synaptic cargo via the lysosomal kinesin adapter Arl8 regulates AZ abundance to modulate global synaptic output and sustain the homeostatic potentiation of neurotransmission. Finally, we find that this AZ scaling can interface with two independent homeostats, depression and potentiation, to remodel AZ structure and function, demonstrating a robust balancing of separate homeostatic adaptations. Thus, AZs are pliable substrates with elastic and modular nanostructures that can be dynamically sculpted to stabilize and tune both local and global synaptic strength.

Ultrastructural basis of strong unitary inhibition in a binaural neuron.

Neurons of the lateral superior olive (LSO) in the brainstem receive powerful glycinergic inhibition that originates from the contralateral ear, and that plays an important role in sound localization. We investigated the ultrastructural basis for strong inhibition of LSO neurons using serial block face scanning electron microscopy. The soma and the proximal dendrite of an LSO neuron are surrounded by a high density of inhibitory axons, whereas excitatory axons are much sparser. A given inhibitory axon establishes contacts via several large axonal thickenings, called varicosities, which typically elaborate several active zones (range 1-11). The number of active zones across inhibitory axon segments is variable. These data thus provide an ultrastructural correlate for the strong and multiquantal, but overall variable, unitary IPSC amplitude observed for inhibitory inputs to LSO neuron. Binaural neurons in the lateral superior olive (LSO) integrate sound information arriving from each ear, and powerful glycinergic inhibition of these neurons plays an important role in this process. In the present study, we investigated the ultrastructural basis for strong inhibitory inputs onto LSO neurons using serial block face scanning electron microscopy. We reconstructed axon segments that make contact with the partially reconstructed soma and proximal dendrite of a mouse LSO neuron at postnatal day 18. Using functional measurements and the Sr2+ method, we find a constant quantal size but a variable quantal content between 'weak' and 'strong' unitary IPSCs. A 3-D reconstruction of a LSO neuron and its somatic synaptic afferents reveals how a large number of inhibitory axons intermingle in a complex fashion on the soma and proximal dendrite of an LSO neuron; a smaller number of excitatory axons was also observed. A given inhibitory axon typically contacts an LSO neuron via several large varicosities (average diameter 3.7μm), which contain several active zones (range 1-11). The number of active zones across individual axon segments was highly variable. These data suggest that the variable unitary IPSC amplitude is caused by a variable number of active zones between inhibitory axons that innervate a given LSO neuron. The results of the present study show that relatively large multi-active zone varicosities, which can be repeated many times in a given presynaptic axon, provide the ultrastructural basis for the strong multiquantal inhibition received by LSO neurons.

Postsynaptic Syntaxin 4 negatively regulates the efficiency of neurotransmitter release

Signaling from the postsynaptic compartment regulates multiple aspects of synaptic development and function. Syntaxin 4 (Syx4) is a plasma membrane t-SNARE that promotes the growth and plasticity of Drosophila neuromuscular junctions (NMJs) by regulating the localization of key synaptic proteins in the postsynaptic compartment. Here, we describe electrophysiological analyses and report that loss of Syx4 leads to enhanced neurotransmitter release, despite a decrease in the number of active zones. We describe a requirement for postsynaptic Syx4 in regulating several presynaptic parameters, including Ca2+ cooperativity and the abundance of the presynaptic calcium channel Cacophony (Cac) at active zones. These findings indicate Syx4 negatively regulates presynaptic neurotransmitter release through a retrograde signaling mechanism from the postsynaptic compartment.

Massive quantum regions for simulations on bio-nanomaterials: synthetic ferritin nanocages.

QM/MM molecular dynamics simulations on the 4His-ΔC* protein cage have been performed using multiple active zones (up to 86 quantum regions). The regulation and nanocage stability exerted by the divalent transition metal ions in the monomer-to-cage conversion have been understood by comparing high level quantum trajectories obtained using Cu2+ and Ni2+ coordination ions.