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Similar Intracellular Ca2+ Requirements for Inactivation and Facilitation of Voltage-Gated Ca2+ Channels in a Glutamatergic Mammalian Nerve Terminal
Get PDFVoltage-gated Ca (2+) channels (VGCCs) of the P/Q-type, which are expressed at a majority of mammalian nerve terminals, show two types of Ca (2+)-dependent feedback regulation-inactivation (CDI) and facilitation (CDF). Because of the nonlinear relationship between Ca (2+) influx and transmitter release, CDI and CDF are powerful regulators of synaptic strength. To what extent VGCCs inactivate or facilitate during spike trains depends on the dynamics of free Ca (2+) ([Ca2+](i)) and the Ca2+ sensitivity of CDI and CDF, which has not been determined in nerve terminals. In this report, we took advantage of the large size of a rat auditory glutamatergic synapse-the calyx of Held-and combined voltage-clamp recordings of presynaptic Ca (2+) currents (I-Ca(v)) with UV-light flash-induced Ca (2+) uncaging and presynaptic Ca (2+) imaging to study the Ca (2+) requirements for CDI and CDF. We find that nearly half of the presynaptic VGCCs inactivate during 100 ms voltage steps and require several seconds to recover. This inactivation is caused neither by depletion of Ca (2+) ions from the synaptic cleft nor by metabotropic feedback inhibition, because it is resistant to blockade of metabotropic and ionotropic glutamate receptors. Facilitation of I-Ca(V) induced by repetitive depolarizations or preconditioning voltage steps decays within tens of milliseconds. Since Ca (2+) buffers only weakly affect CDI and CDF, we conclude that the Ca (2+) sensors are closely associated with the channel. CDI and CDF can be induced by intracellular photo release of Ca (2+) resulting in (Ca2+](i); elevations in the low micromolar range, implying a surprisingly high affinity of the Ca (2+) sensors
A sequential two-step priming scheme reproduces diversity in synaptic strength and short-term plasticity
No full textGlutamatergic synapses display variable strength and diverse short-term plasticity (STP), even for a given type of connection. Using nonnegative tensor factorization and conventional state modeling, we demonstrate that a kinetic scheme consisting of two sequential and reversible steps of release–machinery assembly and a final step of synaptic vesicle (SV) fusion reproduces STP and its diversity among synapses. Analyzing transmission at the calyx of Held synapses reveals that differences in synaptic strength and STP are not primarily caused by variable fusion probability (p(fusion)) but are determined by the fraction of docked synaptic vesicles equipped with a mature release machinery. Our simulations show that traditional quantal analysis methods do not necessarily report p(fusion) of SVs with a mature release machinery but reflect both p(fusion) and the distribution between mature and immature priming states at rest. Thus, the approach holds promise for a better mechanistic dissection of the roles of presynaptic proteins in the sequence of SV docking, two-step priming, and fusion. It suggests a mechanism for activity-induced redistribution of synaptic efficacy
Presynaptic Ca2+ influx and vesicle exocytosis at the mouse endbulb of Held: A comparison of two auditory nerve terminals.
No full textNon-technical summary The release of neurotransmitter from presynaptic nerve endings is triggered by Ca2+ influx through voltage-gated Ca2+ channels (VGCCs) that open when an action potential (AP) invades the presynaptic terminal. The functional properties of VGCCs expressed in presynaptic terminals remain elusive because most terminals are too small to be accessible to electrophysiological recordings. We performed direct presynaptic recordings to characterize Ca2+ channels and transmitter release in a large mammalian presynaptic terminal, the endbulb of Held. Endbulb terminals are formed by the endings of auditory nerve fibres that contact bushy cells located in the anterior ventral cochlear nucleus. We find that endbulb terminals are endowed with >1000 readily releasable vesicles and express an average number of >6000 VGCCs. About half of the VGCCs open during a single AP. Thus, multiple Ca2+ channels control the release of a single transmitter vesicle at the endbulb of Held
Author response image 1.
Get PDFNeurotransmitter is released at synapses by fusion of synaptic vesicles with the plasma membrane. To sustain synaptic transmission, compensatory retrieval of membranes and vesicular proteins is essential. We combined capacitance measurements and pH-imaging via pH-sensitive vesicular protein marker (anti-synaptotagmin2-cypHer5E), and compared the retrieval kinetics of membranes and vesicular proteins at the calyx of Held synapse. Membrane and Syt2 were retrieved with a similar time course when slow endocytosis was elicited. When fast endocytosis was elicited, Syt2 was still retrieved together with the membrane, but endocytosed organelle re-acidification was slowed down, which provides strong evidence for two distinct endocytotic pathways. Strikingly, CaM inhibitors or the inhibition of the Ca2+-calmodulin-Munc13-1 signaling pathway only impaired the uptake of Syt2 while leaving membrane retrieval intact, indicating different recycling mechanisms for membranes and vesicle proteins. Our data identify a novel mechanism of stimulus-and Ca2+-dependent regulation of coordinated endocytosis of synaptic membranes and vesicle proteins
Computational Study of the Origins of the Enhanced Li Storage Capacity and the Kinetic Behaviors of Li Ions on Reduced Graphene Oxide
No full text本研究運用第一原理計算來探討鋰離子電池中石墨烯負極材料的儲鋰物理機制，以及研究其與石墨相比下增加的儲鋰量成因。我們分別研究了不同濃度的官能基─包含修飾邊緣的官能基和存在於石墨烯平面上的官能基─以及不同鋰化程度下的石墨烯的儲鋰行為。此外，我們還進一步研究分析鋰離子在此石墨烯電極上的動力學行為模式。我們將這篇論文分成兩個部分：熱力學層面和動力學層面。 在第一部分的研究中，我們主要探討官能基在熱力學層面下對於儲鋰物理機制的影響。我們首先探討邊緣修飾之官能基的影響。我們的結果顯示，石墨烯以及以氫和羥基修飾邊緣的石墨烯奈米緞帶無法儲鋰。另一方面，羰基和羰基-環醚對的存在卻可以非常有效地增加鋰在石墨烯奈米緞帶上的吸附。此外，我們的研究發現這兩個官能基所修飾的石墨烯奈米緞帶在整個鋰化反應過程中最穩定的吸鋰位置並非是位於碳原子所構成的六方形中心，而是在邊緣修飾的官能基上。對於增加儲鋰量的能力，我們發現羰基要比羰基-環醚對來的好，顯示環醚的存在相對來說並無法有效增加儲鋰量。至於在石墨烯平面上的官能基─環氧基和羥基─它們在鋰化過程中扮演成核中心的角色。當鋰靠近這些官能基時，能夠形成氧化鋰和氫氧化鋰的小分子團簇，因而增加了石墨烯的儲鋰量。我們的結果發現，這些位於平面上的官能基對於增加儲鋰量的能力要比在邊緣修飾的官能基還要好。 在第二部分的研究中，我們探討官能基在動力學層面下對於鋰的擴散以及成核成長的物理機制造成的影響。我們的結果顯示，邊緣修飾的官能機種類和邊緣的形狀都會影響鋰的擴散行為。此外，當邊緣修飾的官能基是羰基和羰基-環醚對的時候，在石墨奈米緞帶邊緣附近的鋰能夠輕易地擴散到邊緣，最終吸附在官能基上。至於在石墨烯平面上的官能基對於鋰的動力學行為的影響是比較複雜的。當鋰遠離這些官能基時，鋰的擴散行為與在沒有官能基存在的石墨烯上差不多。然而，當鋰靠近環氧基/羥基時，它們會成核形成鋰-羰基對/氫氧化鋰分子。而每當有鋰靠近 這個已形成的核時，它將不斷擴大直到長成氧化四鋰/氫氧化三鋰分子團簇。從分子動態模擬的結果發現，這些分子團簇能夠持續吸附在石墨烯上，並沒有脫附而造成儲鋰量的流失。除此之外，我們更發現它們在石墨烯上的擴散能障比單顆鋰原子要來得小許多。由於它們在石墨烯上的擴散較為容易，因此有相當大的機率它們會互相碰撞。當碰撞發生時，我們的結果顯示它們會進一步形成更大的團簇分子，進而影響石墨烯鋰化過程的動力學行為。此外，我們發現當羥基存在於石墨烯表面時，其所催化形成的大分子團簇將有機會脫離石墨烯表面而造成儲鋰量的損失。從材料設計的觀點來看，在製造石墨烯負極材料時時應該盡量減少羥基的含量以避免儲鋰量的不可逆損失。We employed first-principles density functional theory calculations to investigate the lithiation mechanism of the reduced graphene oxides (RGO) and explore the origins of their enhanced storage capacity for the anode of Li-ion batteries. Here we have investigated the Li storage and kinetic behaviors of various types of functional groups located at the edge as well as those lying on the basal plane within different levels of lithiation and functionalization of RGO. In the first part of the thesis, we investigated the effect of functional groups on the lithaition behaviors of graphene narrow ribbons (GNRs) in terms of the thermodynamic viewpoints. Our result shows that lithiation is almost unlikely to happen in pristine graphene and GNRs terminated with OH and H groups. For functional groups terminating the edge, only ketone and K-E pair can effectively enhance Li adsorption on GNRs, and the most favorable sites for Li adsorption turn out to be these edged-oxidized groups rather than the hollow sites on the basal plane. Furthermore, as the ketone-terminated GNRs were fully lithiated, the Li/O atomic ratio was found to be mostly around 1.0~1.25 depending on the concentration of ketone and types of edge, while that for the K-E pair was found to be 0.5 irrespective of the K-E pair concentration and the orientations of graphene edge. This indicates that these edge-oxidized functional groups can indeed enhance Li storage capacity of GNRs. As for the functional groups located on the basal plane, they can serve as the nucleation centers for Li clustering, thereby enhancing the Li storage capacity of GNRs. The Li/O atomic ratio was found to be 4 for the epoxy and 3 (2 for armchair GNR and graphene) for the hydroxyl group. These results indicate that the epoxy and hydroxyl groups on the basal plane tend to be more effective in enhancing the Li storage capacity than the edge functional groups. In the second part of the thesis, we studied the effect of functional groups on the kinetic and dynamic behaviors of the lithaition process in RGO. Our results show that as Li atom is not located in the vicinity of the edge functional groups, the migration energy barriers of Li on the basal plane are mostly comparable to that on pristine graphene. As a Li atom was located near the edge ketone /K-E pair, it was found to diffuse easily toward the edge sites and then adsorb onto the edge functional groups without any sizable energy barrier. As for the functional groups on the basal plane, their effect on the kinetics and dynamics of the lithiation process is much more complicated. The migration energy barrier for Li away from the epoxy/hydroxyl group is nearly identical to that on pristine graphene. However, as Li is in the vicinity of the epoxy/hydroxyl group, Li can diffuse readily towards the functional group and form Li-O pair/Li(OH) cluster on the basal plane without any sizeable energy barrier. Furthermore, these small clusters can grow bigger through the interaction with other Li atoms at the cost of a very low energy barrier. Very interestingly, theses Li clusters can undergo diffusion with a much smaller energy barrier than that for Li diffusion on pristine graphene. Accordingly, these LinO/Lin(OH) clusters are very likely to diffuse towards the neighboring sites of other nanoclusters and then coalesce into an even bigger one as evidenced in our ab initio MD simulations. On the other hand, our MD simulations also show that some of these nanoclusters containing OH groups may tend to desorb from the RGO surface, which can thus lead to the irreversible loss of the Li storage capacity for RGO
Number and relative abundance of synaptic vesicles in functionally distinct priming states determine synaptic strength and short‐term plasticity
No full textAbstract Heterogeneity in synaptic strength and short‐term plasticity (STP) was characterized in post‐hearing rat calyx of Held synapses at near‐physiological external [Ca 2+ ] under control conditions and after experimentally induced synaptic potentiation. Kinetic modelling was combined with non‐negative tensor factorization (NTF) to separate changes in synaptic vesicle (SV) priming kinetics from those in SV fusion probability ( p fu sion ). Heterogeneous synaptic strength and STP under control conditions can be fully accounted for by assuming a uniform p fusion among calyx synapses yet profound synapse‐to‐synapse variation in the resting equilibrium of SVs in functionally distinct priming states. Although synaptic potentiation induced by either elevated resting [Ca 2+ ] i , elevated external [Ca 2+ ] or stimulation of the diacylglycerol (DAG) signalling pathway leads to seemingly similar changes, that is, stronger synapses with less facilitation and more pronounced depression, the underlying mechanisms are different. Specifically, synaptic potentiation induced by the DAG mimetic and Munc13/PKC activator phorbol 12,13‐dibutyrate (PDBu) only moderately enhances p fusion but strongly increases the abundance of fusion‐competent maturely primed SVs, demonstrating that the dynamic equilibrium of differentially primed SVs critically determines synaptic strength and STP. Activation of the DAG pathway not only stimulates priming at resting [Ca 2+ ] i but further promotes SV pool replenishment at elevated [Ca 2+ ] i following pool‐depleting stimulus trains. A two‐step priming and fusion scheme which recapitulates the sequential build‐up of the molecular SV fusion machinery is capable of reproducing experimentally induced changes in synaptic strength and STP in numerical simulations with a small number of plausible model parameter changes. image Key points A relatively simple two‐step synaptic vesicle (SV) priming and fusion scheme is capable of reproducing experimentally induced changes in synaptic strength and short‐term plasticity with a small number of plausible parameter changes. The combination of non‐negative tensor factorization (NTF)‐decomposition analysis and state modelling allows one to separate experimentally induced changes in SV priming kinetics from those in SV fusion probability. A relatively low sensitivity of the SV priming equilibrium to changes in resting [Ca 2+ ] i suggests that the amplitude of the ‘effective’ action potential (AP)‐induced Ca 2+ transient is quite large, likely representing contributions of global and local Ca 2+ signals. Enhanced synaptic strength and stronger depression after stimulation of the diacylglycerol (DAG) signalling pathway is primarily caused by enhanced SV priming, leading to increased abundance of maturely primed SVs at rest with comparably small changes in SV fusion probability. Application of DAG mimetics enhances the Ca 2+ ‐dependent acceleration of SV priming causing a faster recovery of synaptic strength after pool‐depleting stimuli
Going Beyond Counting First Authors in Author Co-citation Analysis
Get PDFThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Complexin has a dual synaptic function as checkpoint protein in vesicle priming and as a promoter of vesicle fusion
Get PDFThe presynaptic SNARE-complex regulator complexin (Cplx) enhances the fusogenicity of primed synaptic vesicles (SVs). Consequently, Cplx deletion impairs action potential-evoked transmitter release. Conversely, though, Cplx loss enhances spontaneous and delayed asynchronous release at certain synapse types. Using electrophysiology and kinetic modeling, we show that such seemingly contradictory transmitter release phenotypes seen upon Cplx deletion can be explained by an additional of Cplx in the control of SV priming, where its ablation facilitates the generation of a "faulty" SV fusion apparatus. Supporting this notion, a sequential two-step priming scheme, featuring reduced vesicle fusogenicity and increased transition rates into the faulty primed state, reproduces all aberrations of transmitter release modes and short-term synaptic plasticity seen upon Cplx loss. Accordingly, we propose a dual presynaptic function for the SNARE-complex interactor Cplx, one as a "checkpoint" protein that guarantees the proper assembly of the fusion machinery during vesicle priming, and one in boosting vesicle fusogenicity
Variations on the Author
Get PDF“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
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