Fast vesicle reloading and a large pool sustain high bandwidth transmission at a central synapse

What limits the rate at which sensory information can be transmitted across synaptic connections in the brain? High-frequency signalling is restricted to brief bursts at many central excitatory synapses, whereas graded ribbon-type synapses can sustain release and transmit information at high rates. Here we investigate transmission at the cerebellar mossy fibre terminal, which can fire at over 200 Hz for sustained periods in vivo, yet makes few synaptic contacts onto individual granule cells. We show that connections between mossy fibres and granule cells can sustain high-frequency signalling at physiological temperature. We use fluctuation analysis and pharmacological block of desensitization to identify the quantal determinants of short-term plasticity and combine these with a short-term plasticity model and cumulative excitatory postsynaptic current analysis to quantify the determinants of sustained high-frequency transmission. We show that release is maintained at each release site by rapid reloading of release-ready vesicles from an unusually large releasable pool of vesicles (approximately 300 per site). Our results establish that sustained vesicular release at high rates is not restricted to graded ribbon-type synapses and that mossy fibres are well suited for transmitting broad-bandwidth rate-coded information to the input layer of the cerebellar cortex.     

Intracellular calcium dependence of transmitter release rates at a fast central synapse

Calcium-triggered fusion of synaptic vesicles and neurotransmitter release are fundamental signalling steps in the central nervous system. It is generally assumed that fast transmitter release is triggered by elevations in intracellular calcium concentration ([Ca2+]i) to at least 100 microM near the sites of vesicle fusion. For synapses in the central nervous system, however, there are no experimental estimates of this local [Ca2+]i signal. Here we show, by using calcium ion uncaging in the large synaptic terminals of the calyx of Held, that step-like elevations to only 10 microM [Ca2+]i induce fast transmitter release, which depletes around 80% of a pool of available vesicles in less than 3 ms. Kinetic analysis of transmitter release rates after [Ca2+]i steps revealed the rate constants for calcium binding and vesicle fusion. These show that transient (around 0.5 ms) local elevations of [Ca2+]i to peak values as low as 25 microM can account for transmitter release during single presynaptic action potentials. The calcium sensors for vesicle fusion are far from saturation at normal release probability. This non-saturation, and the high intracellular calcium cooperativity in triggering vesicle fusion, make fast synaptic transmission very sensitive to modulation by changes in local [Ca2+]i.     

Dual-component NMDA receptor currents at a single central synapse

Present thinking about the way that the NMDA (N-methyl-D-aspartate) class of glutamate receptor operates at central synapses relies mainly on information obtained from single-channel and whole-cell recordings from cultured neurons stimulated by exogenous NMDA receptor agonists. The mechanisms that operate in the postsynaptic membrane of a normal neuron following release of the natural transmitter are far less clear. An important problem is that most normal neurons receive many excitatory synapses (10(3)-10(5) per cell) and these synapses are located on slender dendritic elements far away from the somatic recording site, making the study of discrete synaptic events difficult. Typically, when populations of synapses are activated, NMDA receptor-mediated synaptic potentials appear as slowly rising, long-lasting waves superimposed on faster, non-NMDA-receptor potentials. Although believed to be critical for NMDA receptor function, this slow time-course would not be predicted from single-channel kinetics and its origin remains puzzling. We have now analysed the events occurring at the level of a single excitatory synapse using a simple, small, neuron--the cerebellar granule cell--which has an unusually simple glutamatergic input. By applying high-resolution whole-cell recording techniques to these cells in situ, we were able to study the nature of elementary NMDA receptor-mediated synaptic currents. Contrary to expectations, the prominent currents are fast but are followed by slow ones. Both types of current are strongly voltage-dependent but differ subtly in this respect. Furthermore, the currents are absent unless glycine is provided.     

Presynaptic glycine receptors enhance transmitter release at a mammalian central synapse

Glycine and GABAA (gamma-aminobutyric acid A) receptors are inhibitory neurotransmitter-gated Cl- channels localized in postsynaptic membranes. In some cases, GABAA receptors are also found presynaptically, but they retain their inhibitory effect as their activation reduces excitatory transmitter release. Here we report evidence for presynaptic ionotropic glycine receptors, using pre- and postsynaptic recordings of a calyceal synapse in the medial nucleus of the trapezoid body (MNTB). Unlike the classical action of glycine, presynaptic glycine receptors triggered a weakly depolarizing Cl- current in the nerve terminal. The depolarization enhanced transmitter release by activating Ca2+ channels and increasing resting intraterminal Ca2+ concentrations. Repetitive activation of glycinergic synapses on MNTB neurons also enhanced glutamatergic synaptic currents, indicating that presynaptic glycine receptors are activated by glycine spillover. These results reveal a novel site of action of the transmitter glycine, and indicate that under certain conditions presynaptic Cl- channels may increase transmitter release.     

Developmental regulation of NMDA receptor-mediated synaptic currents at a central synapse.

The central nervous system has extraordinary plasticity in early life. This is thought to involve N-methyl-D-aspartate (NMDA) receptors which, along with the non-NMDA receptors, mediate fast excitatory synaptic transmission. Although NMDA receptors may be transiently enhanced early in life, it has not been possible to demonstrate directly a functional change in the NMDA receptor-mediated synaptic response because of the voltage-dependence of the NMDA conductance and the overlapping inhibitory synaptic conductances. Here I report that the duration of evoked NMDA-receptor-mediated excitatory postsynaptic currents (e.p.s.cs) in the superior colliculus is several times longer at early developmental stages compared to that measured in older animals. In contrast, the amplitude of NMDA-receptor-mediated miniature e.p.s.cs does not change during development. The kinetic response of excised membrane patches to a brief activation of NMDA receptors is similar to that of the NMDA e.p.s.c, which suggests that the time course of the NMDA e.p.s.c. in the superior colliculus reflects slow NMDA channel properties as in the hippocampus. Therefore, these data indicate that the molecular properties of NMDA receptors are developmentally regulated and thus may be controlling the ability of synapses to change in early life.     

Two modes of fusion pore opening revealed by cell-attached recordings at a synapse

Fusion of a vesicle with the cell membrane opens a pore that releases transmitter to the extracellular space. The pore can either dilate fully so that the vesicle collapses completely, or close rapidly to generate 'kiss-and-run' fusion. The size of the pore determines the release rate. At synapses, the size of the fusion pore is unclear, 'kiss-and-run' remains controversial, and the ability of 'kiss-and-run' fusion to generate rapid synaptic currents is questionable. Here, by recording fusion pore kinetics during single vesicle fusion, we found both full collapse and 'kiss-and-run' fusion at calyx-type synapses. For full collapse, the initial fusion pore conductance (G(p)) was usually >375 pS and increased rapidly at > or =299 pS ms(-1). 'Kiss-and-run' fusion was seen as a brief capacitance flicker (<2 s) with G(p) >288 pS for most flickers, but within 15-288 pS for the remaining flickers. Large G(p) (>288 pS) might discharge transmitter rapidly and thereby cause rapid synaptic currents, whereas small G(p) might generate slow and small synaptic currents. These results show that 'kiss-and-run' fusion occurs at synapses and that it can generate rapid postsynaptic currents, and suggest that various fusion pore sizes help to control the kinetics and amplitude of synaptic currents.     

Direct modulation of synaptic vesicle priming by GABA(B) receptor activation at a glutamatergic synapse

Second messenger cascades involving G proteins and calcium are known to modulate neurotransmitter release. A prominent effect of such a cascade is the downmodulation of presynaptic calcium influx, which markedly reduces evoked neurotransmitter release. Here we show that G-protein-mediated signalling, such as through GABA (gamma-amino butyric acid) subtype B (GABA(B)) receptors, retards the recruitment of synaptic vesicles during sustained activity and after short-term depression. This retardation occurs through a lowering of cyclic AMP, which blocks the stimulatory effect of increased calcium concentration on vesicle recruitment. In this signalling pathway, cAMP (functioning through the cAMP-dependent guanine nucleotide exchange factor) and calcium/calmodulin cooperate to enhance vesicle priming. The differential modulation of the two forms of synaptic plasticity, presynaptic inhibition and calcium-dependent recovery from synaptic depression, is expected to have interesting consequences for the dynamic behaviour of neural networks.     

Possible role of acetylcholinesterase in regulation of postsynaptic receptor efficacy at a central inhibitory synapse of Aplysia

Most of the effects of acetylcholinesterase (AChE) on synaptic transmission are considered to be related to its acetylcholine (ACh) hydrolysing properties. This is clearly apparent from changes which occur in the characteristics of the miniature endplate potential and of the endplate potential at neuromuscular junctions when AChE is inhibited1-4 and during the development of enzymatic AChE activity at maturing synapses5. However, we report here that after inhibiting AChE in a cholinergic synapse in Aplysia, we found an increase not only in postsynaptic responses to presynaptic stimulation and to ionophoretic application of ACh on postsynaptic receptors, but also to ionophoretic application of carbachol. This could not be explained by the inhibition of the ACh hydrolysing function of the enzyme, as carbachol is not hydrolysed by AChE. A possible explanation of these observations is that inhibition of the enzyme affects a property of the ACh receptor (AChR) itself.     

循环加、卸载速率对砂岩变形和渗透特性的影响

通过煤岩热流固耦合试验系统(THM-2)对砂岩进行循环加、卸载试验,研究加、卸载速率对其变形和渗透特性的影响.结果表明:初始循环时,岩石的轴向变形量△ε1较大,随着循环试验的进行,△ε1趋于稳定,受卸载速率v2的影响较小.加载变形模量和卸载变形模量均逐渐上升,随着循环次数的增加,上升速度逐渐变缓;同一循环内,卸载变形模量大于加载变形模量,且随着加、卸载的进行,差值逐渐减小.加载阶段和卸载阶段渗透率变化量的差值随着循环次数的增加逐渐减小;从第5次循环开始,渗透率曲线呈“∞”形,渗透率演化规律可以用轴向应变的变化特点表征,轴向应变的变化量△εi1受到卸载速率vi2和应力加载上限σimax的综合作用,二者对应变在卸载初期起到积极的促进作用,三者之间的相互关系可用幂函数表达.     

Currents through the fusion pore that forms during exocytosis of a secretory vesicle

Exocytosis, or the fusion of cytoplasmic vesicles with the cell membrane, occurs in nearly all eukaryotic cells, but its mechanism is not understood. Morphological and electrophysiological studies have suggested that membrane fusion begins with the formation of a 'fusion pore', a narrow channel across the closely adjacent membranes of vesicle and cell that forms the first connection of the vesicle lumen with the cell exterior and later dilates to allow release of vesicle contents. We used the patch clamp technique to study exocytosis of single giant secretory vesicles in mast cells of beige mice. The first opening of the fusion pore was found to generate a brief current transient, whose size and direction indicated an initial pore conductance of about 230 pS and a lumen-positive vesicle membrane potential. In time-resolved a.c. admittance measurements, the pore conductance was found to increase to much larger values within milliseconds, as if the pore dilated soon after opening. We conclude that the earliest fusion event may be the formation of a structure similar to an ion channel. Its conductance is of the same order of magnitude as that of a single gap junction channel, the only other known channel that spans two membranes.     

Integrins mediate functional pre- and postsynaptic maturation at a hippocampal synapse

Coordinated signalling between presynaptic terminals and their postsynaptic targets is essential for the development and function of central synapses. In addition to diffusible molecules, this bidirectional flow of information could involve direct interactions through cell-adhesion molecules. Here, we show that one class of cell-adhesion molecule, the integrins, are required for the functional maturation of hippocampal synapses in vitro. At immature synapses, a high probability of glutamate release (Pr) was correlated with the expression of postsynaptic NMDA (N-methyl-D-aspartate) receptors containing the NR2B subunit. The activity-dependent reduction in Pr and a switch in the subunit composition of synaptic NMDA receptors was prevented by chronic blockade with peptides containing the integrin-binding site Arg-Gly-Asp (RGD), or by a functional antibody against the beta3 integrin subunit. Active synapses, monitored by the uptake of antibodies against the intraluminal domain of synaptotagmin I, also had beta3 subunit immunoreactivity. Our results provide evidence that integrin-mediated signalling is essential for the orchestrated maturation of central excitatory synapses.     

Mechanical characterization of Mg-B_4C nanocomposite fabricated at different strain rates

Magnesium has wide application in industry. The main purpose of this investigation was to improve the properties of magnesium by reinforcing it using B_4C nanoparticles. The reinforced nanocomposites were fabricated using a powder compaction technique for 0, 1.5 vol%, 3 vol%,5 vol%, and 10 vol% of B_4C. Powder compaction was conducted using a split Hopkinson bar(SHB), drop hammer(DH), and Instron to reach different compaction loading rates. The compressive stress–strain curves of the samples were captured from quasi-static and dynamic tests carried out using an Instron and split Hopkinson pressure bar, respectively. Results revealed that, to achieve the highest improvement in ultimate strength, the contents of B_4C were 1.5 vol%, 3 vol%, and 3 vol% for Instron, DH, and SHB, respectively. These results also indicated that the effect of compaction type on the quasi-static strength of the samples was not as significant, although its effect on the dynamic strength of the samples was remarkable. The improvement in ultimate strength obtained from the quasi-static stress–strain curves of the samples(compared to pure Mg) varied from9.9% for DH to 24% for SHB. The dynamic strength of the samples was improved(with respect to pure Mg) by 73%, 116%, and 141% for the specimens compacted by Instron, DH, and SHB, respectively. The improvement in strength was believed to be due to strengthening mechanisms,friction, adiabatic heating, and shock waves.     

不同应变速率下Mg?B4C纳米复合材料的力学性能表征

Magnesium has wide application in industry. The main purpose of this investigation was to improve the properties of magnesium by reinforcing it using B₄C nanoparticles. The reinforced nanocomposites were fabricated using a powder compaction technique for 0, 1.5vol%, 3vol%, 5vol%, and 10vol% of B₄C. Powder compaction was conducted using a split Hopkinson bar (SHB), drop hammer (DH), and Instron to reach different compaction loading rates. The compressive stress–strain curves of the samples were captured from quasi-static and dynamic tests carried out using an Instron and split Hopkinson pressure bar, respectively. Results revealed that, to achieve the highest improvement in ultimate strength, the contents of B₄C were 1.5vol%, 3vol%, and 3vol% for Instron, DH, and SHB, respectively. These results also indicated that the effect of compaction type on the quasi-static strength of the samples was not as significant, although its effect on the dynamic strength of the samples was remarkable. The improvement in ultimate strength obtained from the quasi-static stress–strain curves of the samples (compared to pure Mg) varied from 9.9% for DH to 24% for SHB. The dynamic strength of the samples was improved (with respect to pure Mg) by 73%, 116%, and 141% for the specimens compacted by Instron, DH, and SHB, respectively. The improvement in strength was believed to be due to strengthening mechanisms, friction, adiabatic heating, and shock waves.     

带有N控制策略的M_(λ1 ,λ2 )~[X])/G/1排队模型

在一般批量到达排队模型的基础上,通过运用补充变量的方法构造向量马氏过程,考虑了具有不同到达率且带N控制策略的批量到达排队系统 在此模型中,顾客的到达不是依据固定的到达率来进行,而是与服务员的状态(空闲、忙期)有关 在给出了系统队长分布和顾客在离去时刻点系统队长分布的同时,也给出了忙期内到达顾客的条件等待时间、闲期分布及其均值等排队指标     

带有N控制策略的M（λ1，λ2）^[X]／G／1排队模型

在一般批量到达排队模型的基础上，通过运用补充变量的方法构造向量马氏过程，考虑了具有不同到达率且带N控制策略的批量到达排队系统。在此模型中，顾客的到达不是依据固定的到达率来进行，而是与服务员的状态(空闲、忙期)有关。在给出了系统队长分布和顾客在离去时刻点系统队长分布的同时，也给出了忙期内到达顾客的条件等待时间、闲期分布及其均值等排队指标。