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Before fusion, synaptic vesicles (SVs) pause at discrete release/docking sites. During repetitive stimulation, the probability of site occupancy changes following SV fusion and replenishment. The occupancy probability is considered to be one of the crucial determinants of synaptic strength, but it is difficult to estimate separately because it usually blends with other synaptic parameters. Thus, the contribution of site occupancy to synaptic function, particularly to synaptic depression, remains elusive. Here, we directly estimated the occupancy probability at the hippocampal mossy fibre‐CA3 interneuron synapse showing synaptic depression, using statistics of counts of vesicular events detected by deconvolution. We found that this synapse had a particularly high occupancy (∼0.85) with a high release probability of a docked SV (∼0.8) under 3 mm external calcium conditions. Analyses of quantal amplitudes and SV counts indicated that quantal size reduction decreased the amplitudes of all responses in a train to a similar degree, whereas release/docking site number was unchanged during trains, suggesting that quantal size and release/docking site number had little influence on the extent of synaptic depression. Model simulations revealed that the initial occupancy with high release probability and slow replenishment determined the time course of synaptic depression. Consistently, decreasing external calcium concentration reduced both the occupancy and release probability, and the reductions in turn produced less depression. Based on these results, we suggest that the occupancy probability is a crucial determinant of short‐term synaptic depression at glutamatergic synapses in the hippocampus. Key points: The occupancy probability of a release/docking site by a synaptic vesicle at presynaptic terminals is considered to be one of the crucial determinants of synaptic strength, but it is difficult to estimate separately from other synaptic parameters.Here, we directly estimate the occupancy probability at the hippocampal mossy fibre‐interneuron synapse using statistics of vesicular events detected by deconvolution.We show that the synapses have particularly high occupancy (0.85) with high release probability (0.8) under high external calcium concentration ([Ca2+]o) conditions, and that both parameter values change with [Ca2+]o, shaping synaptic depression.Analyses of the quantal amplitudes and synaptic vesicle counts suggest that quantal sizes and release/docking site number have little influence on the extent of synaptic depression.The results suggest that the occupancy probability is a crucial determinant of short‐term synaptic depression at glutamatergic synapses in the hippocampus. [ABSTRACT FROM AUTHOR]

When activated at low frequencies (0.1–1 Hz), second postnatal week synapses onto the most distal part of the apical dendritic tree (stratum lacunosum‐moleculare) of rat hippocampal CA1 pyramidal cells display a frequency‐dependent synaptic depression not observed for the more proximal (stratum radiatum) synapses. Depression in this frequency range is thought of as a possible contributor to behavioural habituation. In fact, in contrast to the proximal synapses, the distal synapses provide more direct sensory information from the entorhinal cortex as well as from thalamic nuclei. The use of antagonists showed that the activation of GABAA, GABAB, NMDA, mGlu, kainate, adenosine, or endocannabinoid receptors was not directly involved in the depression, indicating it to be intrinsic to the synapses themselves. While the depression affected paired‐pulse plasticity in a manner indicating a decrease in vesicle release probability, the depression could not be explained by a stimulus‐dependent decrease in calcium influx. Despite affecting the synaptic response evoked by brief high‐frequency stimulation (10 impulses, 20 Hz) in a manner indicating vesicle depletion, the depression was unaffected by large variations in release probability. The depression was found not only to affect the synaptic transmission at low frequencies (0.1–1 Hz) but also to contribute to the depression evolving during brief high‐frequency stimulation (10 impulses, 20 Hz). We propose that a release‐independent process directly inactivating release sites with a fast onset (ms) and long duration (up to 20 s) underlies this synaptic depression. [ABSTRACT FROM AUTHOR]

Synaptic active zone (AZ) contains multiple specialized release sites for vesicle fusion. The utilization of release sites is regulated to determine spatiotemporal organization of the two main forms of synchronous release, uni-vesicular (UVR) and multi-vesicular (MVR). We previously found that the vesicle-associated molecular motor myosin V regulates temporal utilization of release sites by controlling vesicle anchoring at release sites in an activity-dependent manner. Here we show that acute inhibition of myosin V shifts preferential location of vesicle docking away from AZ center toward periphery, and results in a corresponding spatial shift in utilization of release sites during UVR. Similarly, inhibition of myosin V also reduces preferential utilization of central release sites during MVR, leading to more spatially distributed and temporally uniform MVR that occurs farther away from the AZ center. Using a modeling approach, we provide a conceptual framework that unites spatial and temporal functions of myosin V in vesicle release by controlling the gradient of release site release probability across the AZ, which in turn determines the spatiotemporal organization of both UVR and MVR. Thus myosin V regulates both temporal and spatial utilization of release sites during two main forms of synchronous release.

The continuing decline and loss of biodiversity has caused an increase in the use of interventionist conservation tools, such as translocation. However, many translocation attempts fail to establish viable populations, with poor release site selection often flagged as an inhibitor of success. We used species distribution models (SDMs) to predict the climate suitability of 102 release sites for amphibians, reptiles, and terrestrial insects and compared suitability predictions between successful and failed attempts. We then quantified the importance of climate suitability relative to 5 other variables frequently considered in the literature as important determinants of translocation success: number of release years, number of individuals released, life stage released, origin of the source population, and position of the release site relative to the species' range. Probability of translocation success increased as predicted climate suitability increased and this effect was the strongest among the variables we considered, accounting for 48.3% of the variation in translocation outcome. These findings should encourage greater consideration of climate suitability when selecting release sites for conservation translocations and we advocate the use of SDMs as an effective way to do this. Article impact statement: Species distribution models effectively identify climates more likely to lead to successful establishment of translocated populations. [ABSTRACT FROM AUTHOR]

Central mammalian synapses release synaptic vesicles in dedicated structures called docking/release sites. It has been assumed that when voltage-dependent calcium entry is sufficiently large, synaptic output attains a maximum value of one synaptic vesicle per action potential and per site. Here we use deconvolution to count synaptic vesicle output at single sites (mean site number per synapse: 3.6). When increasing calcium entry with tetraethylammonium in 1.5 mM external calcium concentration, we find that synaptic output saturates at 0.22 vesicle per site, not at 1 vesicle per site. Fitting the results with current models of calcium-dependent exocytosis indicates that the 0.22 vesicle limit reflects the probability of docking sites to be occupied by synaptic vesicles at rest, as only docked vesicles can be released. With 3 mM external calcium, the maximum output per site increases to 0.47, indicating an increase in docking site occupancy as a function of external calcium concentration.

Following action potential invasion in presynaptic terminals, synaptic vesicles are released in a stochastic manner at release sites (docking sites). Since neurotransmission occurs at frequencies up to 1 kHz, the mechanisms underlying consecutive vesicle releases at a docking site during high frequency bursts is a key factor for understanding the role and strength of the synapse. Particularly new vesicle recruitment at the docking site during neuronal activity is thought to be crucial for short-term plasticity. However current studies have not reached a unified docking site model for central synapses. Here I review newly developed analyses that can provide insight into docking site models. Quantal analysis using counts of vesicular release events provide a wealth of information not only to monitor the number of docking sites, but also to distinguish among docking site models. The stochastic properties of cumulative release number during bursts allow us to estimate the total number of releasable vesicles and to deduce the features of vesicle recruitment at docking sites and the change of release probability during bursts. This analytical method may contribute to a comprehensive understanding of release/replenishment mechanisms at a docking site.

Synaptic vesicle fusion occurs at specialized release sites at the active zone. How refilling of release sites with new vesicles is regulated in central synapses remains poorly understood. Using nanoscale-resolution detection of individual release events in rat hippocampal synapses we found that inhibition of myosin V, the predominant vesicle-associated motor, strongly reduced refilling of the release sites during repetitive stimulation. Single-vesicle tracking revealed that recycling vesicles continuously shuttle between a plasma membrane pool and an inner pool. Vesicle retention at the membrane pool was regulated by neural activity in a myosin V dependent manner. Ultrastructural measurements of vesicle occupancy at the plasma membrane together with analyses of single-vesicle trajectories during vesicle shuttling between the pools suggest that myosin V acts as a vesicle tether at the plasma membrane, rather than a motor transporting vesicles to the release sites, or directly regulating vesicle exocytosis.

Following action potential invasion in presynaptic terminals, synaptic vesicles are released in a stochastic manner at release sites (docking sites). Since neurotransmission occurs at frequencies up to 1 kHz, the mechanisms underlying consecutive vesicle releases at a docking site during high frequency bursts is a key factor for understanding the role and strength of the synapse. Particularly new vesicle recruitment at the docking site during neuronal activity is thought to be crucial for short-term plasticity. However current studies have not reached a unified docking site model for central synapses. Here I review newly developed analyses that can provide insight into docking site models. Quantal analysis using counts of vesicular release events provide a wealth of information not only to monitor the number of docking sites, but also to distinguish among docking site models. The stochastic properties of cumulative release number during bursts allow us to estimate the total number of releasable vesicles and to deduce the features of vesicle recruitment at docking sites and the change of release probability during bursts. This analytical method may contribute to a comprehensive understanding of release/replenishment mechanisms at a docking site. [ABSTRACT FROM AUTHOR]

Action potentials (spikes) can trigger the release of a neurotransmitter at chemical synapses between neurons. Such release is uncertain, as it occurs only with a certain probability. Moreover, synaptic release can occur independently of an action potential (asynchronous release) and depends on the history of synaptic activity. We focus here on short-term synaptic facilitation, in which a sequence of action potentials can temporarily increase the release probability of the synapse. In contrast to the phenomenon of short-term depression, quantifying the information transmission in facilitating synapses remains to be done. We find rigorous lower and upper bounds for the rate of information transmission in a model of synaptic facilitation. We treat the synapse as a two-state binary asymmetric channel, in which the arrival of an action potential shifts the synapse to a facilitated state, while in the absence of a spike, the synapse returns to its baseline state. The information bounds are functions of both the asynchronous and synchronous release parameters. If synchronous release facilitates more than asynchronous release, the mutual information rate increases. In contrast, short-term facilitation degrades information transmission when the synchronous release probability is intrinsically high. As synaptic release is energetically expensive, we exploit the information bounds to determine the energy–information trade-off in facilitating synapses. We show that unlike information rate, the energy-normalized information rate is robust with respect to variations in the strength of facilitation.

Many central synapses contain a single presynaptic active zone and a single postsynaptic density. Vesicular release statistics at such "simple synapses" indicate that they contain a small complement of docking sites where vesicles repetitively dock and fuse. In this work, we investigate functional and morphological aspects of docking sites at simple synapses made between cerebellar parallel fibers and molecular layer interneurons. Using immunogold labeling of SDS-treated freeze-fracture replicas, we find that Cav2.1 channels form several clusters per active zone with about nine channels per cluster. The mean value and range of intersynaptic variation are similar for Cav2.1 cluster numbers and for functional estimates of docking-site numbers obtained from the maximum numbers of released vesicles per action potential. Both numbers grow in relation with synaptic size and decrease by a similar extent with age between 2 wk and 4 wk postnatal. Thus, the mean docking-site numbers were 3.15 at 2 wk (range: 1-10) and 2.03 at 4 wk (range: 1-4), whereas the mean numbers of Cav2.1 clusters were 2.84 at 2 wk (range: 1-8) and 2.37 at 4 wk (range: 1-5). These changes were accompanied by decreases of miniature current amplitude (from 93 pA to 56 pA), active-zone surface area (from 0.0427 μm² to 0.0234 μm²), and initial success rate (from 0.609 to 0.353), indicating a tightening of synaptic transmission with development. Altogether, these results suggest a close correspondence between the number of functionally defined vesicular docking sites and that of clusters of voltage-gated calcium channels. [ABSTRACT FROM AUTHOR]

The number and availability of vesicle release sites at the synaptic active zone (AZ) are critical factors governing neurotransmitter release; yet, these fundamental synaptic parameters have remained undetermined. Moreover, how neural activity regulates the spatiotemporal properties of the release sites within individual central synapses is unknown. Here, we combined a nanoscale imaging approach with advanced image analysis to detect individual vesicle fusion events with -27 nm localization precision at single hippocampal synapses under physiological conditions. Our results revealed the presence of multiple distinct release sites within individual hippocampal synapses. Release sites were distributed throughout the AZ and underwent repeated reuse. Furthermore, the spatiotemporal properties of the release sites were activity dependent with a reduction in reuse frequency and a shift in location toward the AZ periphery during high-frequency stimulation. These findings have revealed fundamental spatiotemporal properties of individual release sites in small central synapses and their activity-dependent modulation. [ABSTRACT FROM AUTHOR]

Classic theories link cognitive abilities to synaptic properties and human-specific biophysical features of synapses might contribute to the unparalleled performance of the human cerebral cortex. Paired recordings and multiple probability fluctuation analysis revealed similar quantal sizes, but 4-times more functional release sites in human pyramidal cell to fast-spiking interneuron connections compared to rats. These connections were mediated on average by three synaptic contacts in both species. Each presynaptic active zone (AZ) contains 6.2 release sites in human, but only 1.6 in rats. Electron microscopy (EM) and EM tomography showed that an AZ harbors 4 docked vesicles in human, but only a single one in rats. Consequently, a Katz’s functional release site occupies ~0.012 μm2 in the human presynaptic AZ and ~0.025 μm2 in the rat. Our results reveal a robust difference in the biophysical properties of a well-defined synaptic connection of the cortical microcircuit of human and rodents.