Share and share alike: trading ofpresynaptic elements betweencentral synapsesKevin Staras

MRC Laboratory for Molecular Cell Biology and Cell Biology Unit, University College London, London WC1E 6BT, UK

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Central presynaptic terminals harbour synaptic vesicles(SVs) and synapse-specific proteins necessary for neuro-transmission. Classically, these elements were thought toreside more or less stably at individual mature synapses,giving rise to the idea that each terminal was essentiallyan independent functional unit. However, emergingevidence from fluorescence imaging studies in hippo-campal cultured neurons is now challenging this view,suggesting that neighbouring synapses along axonsshare vesicles, and also other synaptic elements, at highlevels. This raises the possibility that control of importand export might be an important regulatory target forthe maintenance of release sites, modulation of synapticefficacy and formation of new synaptic contacts. Here,temporal synaptic stability and the functional con-sequences for presynaptic operation will be considered.

IntroductionMost information transfer in the CNS relies on fast,chemical transmission at small synapses, specializedneuron–neuron junction points at which presynaptic (axo-nal) and postsynaptic (e.g. dendritic) structures lie closelyapposed. The presynaptic terminal is characterized by acluster of neurotransmitter-filled synaptic vesicles (SVs)arranged near a dedicated release site on the membrane,and transmission proceeds by the exocytic fusion of thesevesicles and release of neurotransmitter into the extra-cellular synaptic cleft [1,2]. Subsequently, fused vesiclemembrane is reclaimed from the presynaptic plasmamem-brane through an endocytic pathway, and vesicles arerestored to the synaptic cluster. Such efficient recyclingof vesicles, achieved through the intimate coupling ofexocytic and endocytic pathways, facilitates sustainedtransmission under physiological conditions [3].

In the classical interpretation of the vesicle cycle,therefore, synapses represent essentially independentunits, efficiently recycling SVs for local reuse [4,5]. How-ever, although experimental support for local endocytosisof vesicles is overwhelming, emerging evidence is nowquestioning the assumption that all SVs are locally reusedat the synapse at which they are recycled. Using fluor-escent markers, recent findings in mature hippocampalneurons show that functional SVs are not anchored rigidlyto a host synapse but instead show a considerable degree of

Corresponding author: Staras, K. (k.staras@ucl.ac.uk).Available online 27 April 2007.

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extrasynaptic mobility [6,7]. Vesicles can leave stablesynaptic release sites and transit along axons to non-nativesynapses, whereby they can participate in exocytosis.Moreover, synaptic sharing is not restricted to vesicles;other presynaptic elements also move readily betweensynapses [8–10]. This is consistent with findings from arange of studies revealing extensive lateral mobility ofpostsynaptic components [10–15]. In combination, theseresults raise important questions about the operationalindependence of adjacent release sites. They also suggestthat processes regulating synaptic sharing might beimportant in the setting up, maintenance and modulationof presynaptic performance. This review examines theevidence for sharing of synaptic elements, in particularSVs, at central synapses and considers the consequences ofthis phenomenon for presynaptic operation.

Extrasynaptic vesicle trafficCentral presynaptic terminals are typically discretestructures along axons and locally recycle synapticmachinery to support ongoing release. As such, they formsmall, functionally autonomous compartments. Nonethe-less, synapses often lie close to neighbours along the axon(Figure 1a). This offers the possibility that if components,such as SVs, were laterally mobile they might be function-ally relevant to multiple synapses. Lateral sharingbetween adjacent synapses would raise interesting ques-tions about the interdependence of individual terminalsand might have important functional implications.

For laterally mobile vesicles to be relevant topresynaptic function, a necessary demand is that term-inals can access extrasynaptic vesicle traffic. Recent workat an invertebrate peripheral synapse has uncovered anelegant example of such a capability [16]. At the Droso-phila neuromuscular junction, ongoing peptidergic trans-mission relies on replenishment of large dense-corevesicles from the soma. To enable directed refilling offunctional pools, activated synapses are able to recruitmobile vesicles as they pass continuously throughsynapses [16]. Although this example does not relate tointersynaptic sharing per se, it provides an importantillustration of how a terminal can exploit extrasynapticallyderived mobile vesicles in a regulated manner to meetpresynaptic demand. Until recently, specific studies ofvesicle exchange between independent presynaptic term-inals have been limited. One definitive examination of

d. doi:10.1016/j.tins.2007.04.005

Figure 1. Arrangement of hippocampal synapses in culture. (a) Typical

organization of neighbouring synapses. Individual release sites are often formed

en passant, so the presynaptic terminal simply represents a swelling on one side of

the axon. Thus, adjacent synapses are connected by short lengths of axon,

providing a local pathway for the sharing of synaptic elements (arrows). (b)

Organization of vesicle pools at the presynaptic terminal. Serial electron

micrographs of a synapse labelled with FM dye and subsequently

photoconverted so that recycling vesicles (�40% of the total pool) appear black,

whereas nonreleasable vesicles have clear lumens [7] (left). Three-dimensional

reconstruction of the same synapse reveals the intermixing of recycling and

nonreleasable vesicles within the total vesicle cluster (right). Scale bars: 200 nm.

Pink, active zone; light blue, presynaptic; light brown, postsynaptic. The left-hand

panel of (b) is reproduced, with permission, from [7].

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this issue came from work at the frog neuromuscularjunction [17]. Here, elegant experiments revealed thatvesicles were laterally immobile in both resting and stimu-lated nerve terminals. Only in the presence of okadaic acid,a protein phosphatase inhibitor that disrupts the integrityof vesicle clusters, was significant lateral movement ofvesicles between presynaptic terminals observed [17].Thus, in this system, regulatory processes must normallyfunction to confine vesicles to individual synaptic boutons.But, what about the well-characterized synapses of hippo-campal neurons? Here, lateral trafficking of vesicles alongaxons has been well-established in developing neurons inrelation to nascent synapse formation [18–20]. This offersthe possibility that lateral movement of vesicles might alsopersist in mature neurons, with potential relevance forpresynaptic function. With the advent of new imagingmethodologies, functional fluorescent probes and correla-tive fluorescence and ultrastructural imaging techniques,recent work has now specifically addressed this issue. Thefindings are outlined below.

Synaptic vesicle sharing at central synapsesIn culture, hippocampal synapses are typically composedof approximately equal numbers of recycling and nonre-leasable vesicles [3] (Figure 1b) mixed within the synapticterminal [7,21–23]. Using FM-styryl dyes, specific mar-kers of recycling vesicles [24,25] that label individualstable release sites as discrete fluorescent punctum,considerable information on the SV cycle has been gener-ated [3]. Recent work examining time-lapse sequenceshas also revealed another dynamic feature of FM-labelledneurons: the constitutive retrograde and anterograde

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transiting of fluorescent packets between synapses[6,7]. The fact that these mobile elements are FM-positiveimplies that they must correspond to recycling vesicles,presumably arising from full-fusion events at sites ofendocytosis, before being mobilized. This is confirmedby fluorescence imaging studies capturing the buddingof mobile packets from stable, mature terminals [6,7](Figure 2a). Furthermore, ultrastructural evidence,derived from FM-dye photoconversion techniques, rev-eals that these packets are composed of clusters of recy-cling (photoconverted) and nonphotoconverted SVs [7](Figure 2b). This latter group might correspond to newlysynthesized vesicles that have aggregated with mobilerecycling vesicles or, perhaps more probably, arise fromthe nonreleasable pool of synapses. Indeed, vesicle com-position in mobile packets versus donor synapses is strik-ingly similar, implying that mobile vesicle clusters budfrom stable synapses with little or no functional selectionbias [7].

Fate of mobile vesicle clusters

Do mobile SVs transiently enter and leave a synaptic hostor do they integrate and persist? If so, can they affectsynaptic release properties? Fluorescence recovery afterphotobleaching (FRAP; Box 1) experiments have offeredinsight into these issues [7,26]. Synapses targeted forphotobleaching in FM-labelled cultures rapidly acquirenew fluorescent vesicles arising from unbleached synapses(Figure 2c). Ultrastructural analysis of this FRAP signalreveals that mobile vesicles mix readily into the hostvesicle cluster and recently incorporated vesicles representa significant fraction of the recycling pool (�20% over aperiod of 20 min; Figure 2d) [7]. Moreover, recentlyimported vesicles are capable of depolarization-evokedexocytosis, with normal release kinetics [7]. Thus, theaccumulation of mobile SV clusters at release sites hasconsiderable functional relevance for ongoing synapticoperation.

In addition to trafficking between release sites, analternative operational fate for mobile SV clusters hasbeen identified. FM-positive vesicles budding from stable,mature synapses can aggregate at new sites along theaxon, even if there is no postsynaptic apposition (so-called‘orphan’ synapses) [6]. These newly formed release sitesare capable of Ca2+-dependent exocytosis, with releasekinetics equivalent to the parent synapse. Moreover, thisrelease capability seems to be acquired rapidly, in theorder of minutes or less [6]. How do newly generatedsynapses maintain or acquire release-competency if theyare remote from the specialized presynaptic terminal?Demand for functional release machinery at nascent sitesmight be met in several ways. For example, the target N-ethylmaleimide-sensitive fusion attachment protein recep-tor (SNARE) proteins, 25-kDa synaptosomal-associatedprotein (SNAP-25) and syntaxin, are relatively uniformlydistributed along the axonal membrane [27], implying thattheymight be locally available at new sites. However, othersynaptically localized elements must be actively recruitedto nascent synapses. Whether these travel as part of themobile vesicle ensemble [6] or separately is unclear. None-theless, emerging evidence suggests that, similar to SVs,

Figure 2. Lateral vesicle mobility between stable synaptic release sites. (a) The budding of SV-containing fluorescent packets (yellow–orange arrowheads) from a stable,

mature, FM-dye-labelled synaptic release site (blue arrowhead). The timescales are minutes to seconds. Reproduced, with permission, from [6]. (b) Electron micrograph of a

mobile vesicle cluster along an axon composed of recycling (dark core) and nonreleasable (clear core) vesicles. Scale bar: 200 nm. Reproduced, with permission, from [7].

(c) FRAP experiment revealing the temporal accumulation of mobile FM-dye-labelled vesicles (yellow arrowhead) at a bleached synapse (white box), arising from

neighbouring release sites (indicated by green arrows). In the ‘merge’ panel, green staining represents (enhanced green fluorescent protein) EGFP–GluR2, a postsynaptic

marker, indicating that FM-dye-labelled terminals are co-apposed to postsynaptic structures. Scale bar: 2 mm. The timescales are minutes to seconds. Reproduced, with

permission, from [7]. (d) Ultrastructural readout of FRAP signal. Dark vesicles represent newly incorporated FM-dye-positive vesicles that have recently entered a host

vesicle cluster (clear vesicles). Newly integrated vesicles are present near the active zone (pink), in the cluster core and at the edge of the cluster, implying that they become

spatially mixed into the host synapse. Scale bar: 100 nm. Redrawn, with permission, from [7]. (e) Three-dimensional reconstruction from serial sections of a CA3 axon from

adult hippocampus. Stable synapses with vesicle clusters and postsynaptic densities (red) are shown. Along axons, small numbers of vesicles are observed at intersynaptic

regions, providing indirect evidence for the presence of lateral vesicle exchange in adult brain. Scale bar: 500 nm. Reproduced, with permission, from [32] �1998, Society

for Neuroscience.

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many synaptic proteins are also laterally mobile [8–10](see below), indicating that necessary elements mighttransit continuously along axons and, therefore, be locallyavailable at nascent sites.

Box 1. Characterizing lateral mobility of synaptic elements

Time-lapse imaging methods used in combination with specific

fluorescent probes represent a powerful approach for examining

mobility of elements in living tissue [64]. Researchers often rely on

FRAP, in which fluorophores attached to a target element are

irreversibly photobleached within a defined region using focused

laser light [65]. Temporal recovery of a fluorescence signal at the

bleach site corresponds to movement of new fluorescent elements

from unbleached areas and provides a quantitative readout of both

the mobile fraction within the bleached region and its rate of

mobility. For examining intersynaptic sharing, researchers typically

photobleach an entire fluorescently labelled synapse [7–10,17]; in

this case, recovery at the bleached synapse corresponds to move-

ment of fluorescence from adjunct axons and neighbouring release

sites. An emerging variation on conventional FRAP uses recently

developed photoactivatable or photoswitchable fluorophores that, if

excited at specific wavelengths, undergo molecular changes that

alter their spectral properties. These include pa-GFP (photoactiva-

table green fluorescent protein), a variant of GFP, which increases

its fluorescence emission 100-fold if photoactivated [66], and Kaede

[67] and Dendra [68] green-emitting fluorophores, which can be

converted to red-emitting forms if photoswitched. If tagged to

intracellular elements of interest, these compounds can be used to

track the temporal spread of fluorescence from localized target

regions [10,13] and offer great promise for future work in this field.

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Sharing of vesicles: a developmental or mature

process?

Could vesicle sharing in mature neurons represent avestige of synaptic mobility associated with neuronal de-velopment? It is well established that vesicle clusters arehighly labile during neurite outgrowth and exhibit a Ca2+-dependent form of SV recycling associated with vesiclematuration and formation of nascent synapses [28,29].This immature form of vesicle exocytosis relies on differentmolecular machinery to release processes at maturesynapses, because it is insensitive to tetanus toxin [avesicle-associated membrane protein 2 (VAMP2) inactiva-tor] [30] and sensitive to brefeldin A (an inhibitor of trans-Golgi trafficking) [31]. Do vesicle fusion and retrieval ofmobile SV units use immature or mature release pro-cesses? Pharmacological experiments on orphan synapsessuggest the latter [6], indicating that they have maturerelease machinery and their mobility and release proper-ties are distinct from the processes underlying develop-mental synaptogenesis. Nonetheless, nominally maturecultured neurons are far fromperfectmodels of adult brain.Is extrasynaptic vesicle sharing evident in intact adulthippocampus? Serial reconstructions from adult hippo-campal slices show that CA3 axons, similar to culturedneurons, form many en passant synapses (�3 mm apart) sothat neighbouring terminals locally share an axon [32].Moreover, lone vesicles or small vesicle clusters are readilyobserved at intersynaptic regions [32] (Figure 2e).Whether

Figure 3. Remodelling synapses by modulating vesicle sharing. (a) The cartoon

illustrates the relationship between stable synaptic release sites and extrasynaptic

mobile vesicle pools located along the axon. In resting synapses, exchange

between pools (red arrows) is balanced so that synaptic vesicle clusters maintain

their size over time. (b) A possible mechanism for the remodelling of SV pools.

Biasing of vesicle exchange towards vesicle accumulation or disassembly would

lead to a rescaling of SV clusters, and this might have functional consequences for

synaptic strength (top; see main text). Directed recruitment of mobile vesicles to

nascent sites along axons, represents a potential mechanism to permit rapid

formation of new functional release sites (bottom). Closed circles, recycling

vesicles; open circles, nonreleasable vesicles; pink lines, active zone; light blue,

presynaptic; light brown, postsynaptic.

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these vesicles aremobile, or indeed functional, is unknown,but their appearance is strikingly similar to the arrange-ment of trafficking vesicles in culture, offering indirectevidence that lateral vesicle sharing might be a conservedfeature in adult brain.

Molecular mechanisms underlying vesicle sharing

What machinery relays vesicles along axons and underliesSV capture and release at synapses? Vesicle trafficking isthought to be an active process involving microtubule andactin cytoskeletons [33]. In support of this, reported mobi-lity rates for SV clusters are 0.1–2.2 mm/s [6,7], broadlyconsistent with transport on microtubule-based motors[34]. Actin is also implicated because jasplakinolide, anactin stabilizer, inhibits SV cluster trafficking betweensynapses and prevents incorporation of new vesicles atrelease sites [7]. Thus, actin is probably functioning coop-eratively withmicrotubule-based transport mechanisms totransfer vesicle cargo along axons and perhaps also oper-ates at synapses to convey mobile vesicles between stableclusters and axonal transport machinery. The cytomatrixprotein synapsin is another candidate regulator, because itis known to be involved in control of vesicle cluster integ-rity and mobility at synapses [35,36]; disruption of synap-sin–vesicle crosslinks, using the phosphatase inhibitorokadaic acid, triggers lateral movement of vesicles awayfrom clusters and increases vesicle mobility �30-foldwithin the terminal [37]. Thus, synapsin phosphorylationmight be an important factor in regulating SV clustermaintenance and facilitating lateral exchange of vesiclesbetween release sites [36]. Recent work has also implicatedthe neurotrophic factor brain-derived neurotrophic factor(BDNF) in the regulation of vesicle clustering at synapsesthrough its action on the cadherin–b-catenin complex [38].BDNF signalling phosphorylates b-catenin through theactivation of neurotrophin trkB receptors, disruptingvesicle tethering at release sites. Acute application ofBDNF can increase the number of mobile SV clustersand promote the formation of new synapses [38]. Notably,BDNF is also required for presynaptic expression of long-term potentiation (LTP) at CA3–CA1 synapses, throughmodulation of release probability [39,40] and, specifically,changes in the size of the functional vesicle pool [40]. Thesemultiple roles suggest that BDNF could be a pivotalelement, both in regulation of lateral vesicle sharingand, perhaps, in presynaptic remodelling underlying lo-ng-term plasticity.

Vesicle sharing as a substrate for presynapticremodellingFunctional vesicle clusters moving continuously betweensynapses essentially form an extrasynaptic vesicle pool(Figure 3a), operating as both a reserve and a sink forlocal presynaptic terminals. What function could thismobile pool serve? Under basal conditions, vesicle gainand loss seem to be balanced so that a stable synapticcluster approximately maintains its size over time [7].However, disrupting the equilibrium of vesicle exchangecould potentially have important implications for synapticrelease properties. The basis for this hypothesis is thefinding that release probability, one of the fundamental

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parameters defining synaptic strength, is positivelycorrelated to the size of the recycling pool [41]. Thus,differential regulation of vesicle exchange at individualsynapses could theoretically participate in the settingand maintenance of synaptic weight through control ofpool sizes.

Synapse-specific recruitment of extrasynaptically-derived vesicle traffic already has an experimental basisin recent, elegant work in vivo at the Drosophila neuro-muscular junction [16]. Here, depleted peptidergic dense-core vesicle pools at synapses are refilled by accessing anexcess retrograde vesicle flux passing continuouslythrough terminals. Specific mechanisms operating withinindividual boutons control the amount of vesicle capturefrom this transiting pool in an activity-dependent manner.Accordingly, a distally and slowly generated resource canbe rapidly accumulated at the synapses that need them[16]. Could similar regulation of vesicle capture operatingat central synapses contribute to synapse-specific forms oflong-term plasticity? There is now compelling evidence tosuggest that both long-term depression (LTD) and LTP canhave a presynaptic locus of expression consistent with adecrease [42] and increase [39,40,43–45], respectively, inthe probability of vesicle release. However, the underlyingmechanisms have not been clearly established. Theoreti-cally, local regulation of vesicle exchange, leading to a

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dynamic rescaling of pool sizes and associated change insynaptic strength, could contribute to plasticity-inducedalteration in presynaptic function [7,46] (Figure 3b). Forexample, retrograde signals arising from postsynapticinduction of long-term plasticity might trigger either lossor recruitment of vesicles, leading to a reduction orincrease, respectively, in the presynaptic recycling poolsize. In support of this, burst-patterned stimulation ofimmature neurons leads to the removal of recyclingvesicles from boutons in anN-methyl-D-aspartate (NMDA)receptor-dependent manner, and this synaptic disassem-bly is associated with a decrease in presynaptic function[19]. Augmentation of a functional subset of the recyclingpool has recently been correlated to a presynapticallyexpressed form of LTP [40], although so far a link betweenoverall increases in the total recycling pool size and LTPhas not been established. Thus, a specific role for vesicle-pool rescaling in forms of synapse-specific long-termplasticity is, at present, unclear. Nonetheless, in homeo-static forms of plasticity, increases in functional pool sizesseem to be an important mechanism for achieving com-pensatory increases in release probability in response tolong-term silencing of neuronal activity [47,48].

Dynamic recruitment of vesiclesmight also participate informs of activity-dependent synaptogenesis. For example,growth of new dendritic spines is thought to be important informs of long-term synaptic plasticity [49–52] and is alsolikely to be accompanied by changes to the existing, or theformation of new, release sites [53–55]. How might newpresynaptic terminals be formed? Conventional models ofde novo synaptogenesis, extensively studied in developingneurons, implicate a multistage mechanism requiring thetrafficking and deposition of active-zone elements and, sub-sequently,maturation and clustering ofSVsat the samesite[19,29,56,57]. Directed recruitment of rapidly release-com-petent vesicles from local pools could offer an alternativemechanism for rapid synaptogenesis in mature neurons(Figure 3b) [6]. One potential benefit of such a mechanismwould be that new release sites could be constructed orremodelled rapidly, in minutes or less. Also, it should bereadily reversible, with the elimination of release sitesarising from disassembly and mobilization of synaptic com-ponents. Further research will be necessary to establish thepromise of this mechanism and its potential role in activity-dependent synaptogenesis.

Sharing of other presynaptic elementsIn addition to SVs, nonvesicular presynaptic elements arealso highly labile. This mobility can be broadly divided intotwo types. First, localized redistribution, which is typicallyactivity-dependent and transient in nature (over a time-scale of seconds tominutes), has been reported for synapsin[36], N-cadherin [58], synaptobrevin [8,59], Rab3 [60],actin [54,61] and clathrin [59,62]. The tight correlationto neuronal stimulation suggests that this relates specifi-cally to exocytic and endocytic processes associated withSV cycling. Thus, dispersion of synaptic elements is oftenlocal to the synaptic terminal, although there are excep-tions: activity-driven surface movement of the vesicleSNARE protein synaptobrevin can extend as far as neigh-bouring synapses following its deposition on the plasma

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membrane following vesicle fusion [8]. Second, constitutivesharing of molecular elements between release sites, akinto lateral exchange of SVs; for example, the cytomatrixprotein synapsin is continuously lost from, redistributedamong and reincorporated into synapses over a timescaleof minutes to hours [10]. In this example, loss and incorp-oration rates are both accelerated by stimulation.

A major potential weakness of these types of studies isthat tracking of synaptic proteins relies onoverexpressionoffluorescent fusion constructs. Thus, spatial and temporaldynamics of protein mobility might be complicated byexpression levels, mistargeting and potential interactionswith native proteins, in addition to the fact that mobilitymight be influenced by the attached fluorescent moiety.Emerging knock-in technology, which relies on endogenousexpression of mutated genes, offers a new approach thatcircumvents several (althoughnotall) of these problemsandits promise is suggested by recent work demonstratingconstitutive intersynaptic exchange of the active-zoneelementMunc13–1 [9]. A further technical problem in char-acterizing mobility dynamics of expressed proteins is tounequivocally discriminate between sharing of establishedproteins at synapses and the trafficking of nascent cargofrom somatic sources. Photoswitchable or photoactivablefluorophores (Box 1) that enable target regions (e.g. thosecontaining resident synaptic proteins) to be selectively‘switched’ offer an interesting approach to addressing thisproblem, as suggested by recent studies [10,13].

A synthesis of the findings outlined above suggests that,similar to lateral SV mobility, sharing of molecularelements is inconsistent with the view of presynapticterminals as independent operational units. What func-tional role might sharing of synaptic constituents serve?One possibility is that it provides a substrate for effectingpresynaptic change. Strong evidence now indicates thatthe molecular composition of synapses might contribute tosynaptic performance [63]. Thus, directed recruitment ofspecific proteins to synapses from locally trafficking poolscould bring about functional modulation of presynapticproperties. Another possibility is that local sharing ofresources might provide a rapid means to allocate proteinsto newly forming synapses before distantly located trans-lation and transcription machinery can respond to thedemand.

Examples of lateral sharing in presynaptic componentsaremore thanmatched by a burgeoning literature reportingmobility of proteinsatpostsynaptic sites.Key components ofthe postsynaptic density, including Ca2+/calmodulin-de-pendent protein kinase II (CaMKII) [12,14], PSD-95[11,13,14], NR1 subunit of the NMDA receptor [14], GluR1(glutamate receptor) subunit of a-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) [14,15] and Pro-SAP2 (proline-rich synapse-associated protein 2) [10], showconsiderable lateral diffusion into and out of synapses.Moreover, strong evidence suggests that potentiatedsynapses redistribute important postsynaptic elements asthe basis of forms of synaptic plasticity [14]. Thus, thereseems to be some consensus, both presynaptically and post-synaptically, regarding the dynamic nature of synapticconstituents. Understanding the mechanisms that mightregulate and coordinate these processes across both

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synaptic compartments represents an important challengefor the future.

Concluding remarksIndividual components of central synapses are highlydynamic; synapses share vesicles and synaptic machinerywith neighbouring release sites over timescales of minutesto hours. These processes seem to be distinct frommechanisms related to developmental synaptogenesis,suggesting that they are relevant to mature synaptic oper-ation. Thus, presynaptic terminals are not defined bysharply drawn boundaries but are in equilibrium with amobile resource pool used by multiple release sites. Whatare the functional consequences of this? In broad terms,sharing of elements would seem to undermine the inde-pendent operation of release sites because each synapsecompetes for shared and highly mobile resources. None-theless, despite the instability of their constituents,synapses maintain structural and functional integrity,suggesting that precise regulatory mechanisms must func-tion to preserve enduring synaptic organization. Delineat-ing these regulatory mechanisms is important not onlybecause they contribute to the maintenance of synapses,but also because they could potentially be used by neuronsto modulate synaptic properties. For example, regulationof the specific balance of vesicle capture and loss at stablerelease sites could contribute to the scaling of vesicleclusters and, therefore, synaptic strength or provide amechanism for rapid synaptogenesis. A similar argumentcan be presented for sharing of molecular synaptic con-stituents. In combination, these findings offer a new,dynamic view for presynaptic terminals, with the proper-ties of a release site in part defined by the balance ofsynaptic resources moving into and out of the synapse.This view is consistent with emerging evidence indicatingthat many postsynaptic elements are also highly mobilebetween synaptic structures and that sharing can bemodu-lated by synaptic use. Although the phenomenon of pre-synaptic vesicle and protein sharing is now established, thefunctional importance of this process still needs to beclarified. Thus, characterization of molecular pathwaysthat control lateral sharing and the conditions under whichthe balance of influx and efflux at synapses might bemodulated are important issues to address. Emergenttechnology, including new developments in fluorescentprobes and high-resolution imaging methodologies, shouldoffer additional technical firepower to shed light on thesequestions.

AcknowledgementsI am indebted to Yukiko Goda for her outstanding support, and to her,Kevin Darcy, Tiago Branco and Ruth Rea for useful and critical commentson this manuscript.

References1 Murthy, V.N. and De Camilli, P. (2003) Cell biology of the presynaptic

terminal. Annu. Rev. Neurosci. 26, 701–7282 Sudhof, T.C. (2004) The synaptic vesicle cycle. Annu. Rev. Neurosci. 27,

509–5473 Fernandez-Alfonso, T. and Ryan, T.A. (2006) The efficiency of the

synaptic vesicle cycle at central nervous system synapses. TrendsCell Biol. 16, 413–420

www.sciencedirect.com

4 Ceccarelli, B. et al. (1973) Turnover of transmitter and synaptic vesiclesat the frog neuromuscular junction. J. Cell Biol. 57, 499–524

5 Heuser, J.E. and Reese, T.S. (1973) Evidence for recycling of synapticvesicle membrane during transmitter release at the frog neuromuscularjunction. J. Cell Biol. 57, 315–344

6 Krueger, S.R. et al. (2003) The presynaptic release apparatus isfunctional in the absence of dendritic contact and highly mobilewithin isolated axons. Neuron 40, 945–957

7 Darcy, K.J. et al. (2006) Constitutive sharing of recycling synapticvesicles between presynaptic boutons. Nat. Neurosci. 9, 315–321

8 Li, Z. and Murthy, V.N. (2001) Visualizing postendocytic traffic ofsynaptic vesicles at hippocampal synapses. Neuron 31, 593–605

9 Kalla, S. et al. (2006) Molecular dynamics of a presynaptic active zoneprotein studied in Munc13-1-enhanced yellow fluorescent proteinknock-in mutant mice. J. Neurosci. 26, 13054–13066

10 Tsuriel, S. et al. (2006) Local sharing as a predominant determinant ofsynaptic matrix molecular dynamics. PLoS Biol. 4, e271

11 Okabe, S. et al. (2001) Rapid redistribution of the postsynaptic densityprotein PSD-Zip45 (Homer 1c) and its differential regulation by NMDAreceptors and calcium channels. J. Neurosci. 21, 9561–9571

12 Okamoto, K. et al. (2004) Rapid and persistent modulation of actindynamics regulates postsynaptic reorganization underlyingbidirectional plasticity. Nat. Neurosci. 7, 1104–1112

13 Gray, N.W. et al. (2006) Rapid redistribution of synaptic PSD-95 in theneocortex in vivo. PLoS Biol. 4, e370

14 Sharma, K. et al. (2006) Postsynaptic protein mobility in dendriticspines: long-term regulation by synaptic NMDA receptor activation.Mol. Cell. Neurosci. 31, 702–712

15 Bats, C. et al. (2007) The interaction between Stargazin and PSD-95regulates AMPA receptor surface trafficking. Neuron 53, 719–734

16 Shakiryanova, D. et al. (2006) Activity-dependent synaptic capture oftransiting peptidergic vesicles. Nat. Neurosci. 9, 896–900

17 Henkel, A.W. et al. (1996) Synaptic vesicle movements monitored byfluorescence recovery after photobleaching in nerve terminals stainedwith FM1-43. J. Neurosci. 16, 3960–3967

18 Ahmari, S.E. et al. (2000) Assembly of presynaptic active zones fromcytoplasmic transport packets. Nat. Neurosci. 3, 445–451

19 Hopf, F.W. et al. (2002) Stability and plasticity of developing synapsesin hippocampal neuronal cultures. J. Neurosci. 22, 775–781

20 Sabo, S.L. et al. (2006) Formation of presynaptic terminals atpredefined sites along axons. J. Neurosci. 26, 10813–10825

21 Kraszewski, K. et al. (1996) Mobility of synaptic vesicles in nerveendings monitored by recovery from photobleaching of synapticvesicle-associated fluorescence. J. Neurosci. 16, 5905–5913

22 Harata, N. et al. (2001) Visualizing recycling synaptic vesicles inhippocampal neurons by FM 1-43 photoconversion. Proc. Natl. Acad.Sci. U. S. A. 98, 12748–12753

23 Micheva, K.D. and Smith, S.J. (2005) Strong effects of subphysiologicaltemperature on the function and plasticity of mammalian presynapticterminals. J. Neurosci. 25, 7481–7488

24 Betz, W.J. and Bewick, G.S. (1992) Optical analysis of synaptic vesiclerecycling at the frog neuromuscular junction. Science 255, 200–203

25 Ryan, T.A. et al. (1993) The kinetics of synaptic vesicle recyclingmeasured at single presynaptic boutons. Neuron 11, 713–724

26 Darcy, K.J. et al. (2006) An ultrastructural readout of fluorescencerecovery after photobleaching using correlative light and electronmicroscopy. Nat. Prot, DOI: 10.1038/nprot.2006.146

27 Garcia, E.P. et al. (1995) rbSec1A and B colocalize with syntaxin 1 andSNAP-25 throughout the axon, but are not in a stable complex withsyntaxin. J. Cell Biol. 129, 105–120

28 Matteoli, M. et al. (1992) Exo-endocytotic recycling of synaptic vesiclesin developing processes of cultured hippocampal neurons. J. Cell Biol.117, 849–861

29 Matteoli, M. et al. (2004) Vesicle turnover in developing neurons: howto build a presynaptic terminal. Trends Cell Biol. 14, 133–140

30 Verderio, C. et al. (1999) Tetanus toxin blocks the exocytosis of synapticvesicles clustered at synapses but not of synaptic vesicles in isolatedaxons. J. Neurosci. 19, 6723–6732

31 Zakharenko, S. et al. (1999) Neurotransmitter secretion along growingnerve processes: comparison with synaptic vesicle exocytosis. J. CellBiol. 144, 507–518

32 Shepherd, G.M. and Harris, K.M. (1998) Three-dimensional structureand composition of CA3!CA1 axons in rat hippocampal slices:

298 Review TRENDS in Neurosciences Vol.30 No.6

implications for presynaptic connectivity and compartmentalization. J.Neurosci. 18, 8300–8310

33 Goode, B.L. et al. (2000) Functional cooperation between themicrotubule and actin cytoskeletons. Curr. Opin. Cell Biol. 12, 63–71

34 Brown, A. (2003) Axonal transport ofmembranous andnonmembranouscargoes: a unified perspective. J. Cell Biol. 160, 817–821

35 Chi, P. et al. (2001) Synapsin dispersion and reclustering duringsynaptic activity. Nat. Neurosci. 4, 1187–1193

36 Chi, P. et al. (2003) Synaptic vesicle mobilization is regulated by distinctsynapsin I phosphorylation pathways at different frequencies. Neuron38, 69–78

37 Shtrahman, M. et al. (2005) Probing vesicle dynamics in singlehippocampal synapses. Biophys. J. 89, 3615–3627

38 Bamji, S.X. et al. (2006) BDNF mobilizes synaptic vesicles andenhances synapse formation by disrupting cadherin-b–catenininteractions. J. Cell Biol. 174, 289–299

39 Zakharenko, S.S. et al. (2003) Presynaptic BDNF required for apresynaptic but not postsynaptic component of LTP at hippocampalCA1-CA3 synapses. Neuron 39, 975–990

40 Tyler, W.J. et al. (2006) BDNF increases release probability and thesize of a rapidly recycling vesicle pool within rat hippocampalexcitatory synapses. J. Physiol. 574, 787–803

41 Murthy, V.N. et al. (1997) Heterogeneous release properties ofvisualized individual hippocampal synapses. Neuron 18, 599–612

42 Zakharenko, S.S. et al. (2002) Altered presynaptic vesicle release andcycling during mGluR-dependent LTD. Neuron 35, 1099–1110

43 Zakharenko, S.S. et al. (2001) Visualization of changes in presynapticfunction during long-term synaptic plasticity. Nat. Neurosci. 4, 711–717

44 Emptage, N.J. et al. (2003) Optical quantal analysis reveals apresynaptic component of LTP at hippocampal Schaffer-associationalsynapses. Neuron 38, 797–804

45 Stanton, P.K. et al. (2005) Imaging LTP of presynaptic release of FM1-43 from the rapidly recycling vesicle pool of Schaffer collateral–CA1synapses in rat hippocampal slices. Eur. J. Neurosci. 22, 2451–2461

46 Krueger, S. and Fitzsimonds, R.M. (2006) Remodeling the plasticitydebate: the presynaptic locus revisited. Physiology (Bethesda) 21, 346–351

47 Murthy, V.N. et al. (2001) Inactivity produces increases inneurotransmitter release and synapse size. Neuron 32, 673–682

48 Thiagarajan, T.C. et al. (2005) Adaptation to synaptic inactivity inhippocampal neurons. Neuron 47, 725–737

49 Engert, F. and Bonhoeffer, T. (1999) Dendritic spine changesassociated with hippocampal long-term synaptic plasticity. Nature399, 66–70

Free journals for dev

The WHO and six medical journal publishers hav

Research Initiative, which enables nearly 70 of the

to biomedical literature

The science publishers, Blackwell, Elsevier, Harc

International Health and Science, Springer-Verlag a

and the British Medical Journal in 2001. Initially, m

free or at significantly reduced prices to universit

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journals are now available. Currently more than 7

Gro Harlem Brundtland, the former director-general o

the biggest step ever taken towards reducing the

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50 Maletic-Savatic, M. et al. (1999) Rapid dendritic morphogenesis in CA1hippocampal dendrites induced by synaptic activity. Science 283,1923–1927

51 Nagerl, U.V. et al. (2004) Bidirectional activity-dependentmorphological plasticity in hippocampal neurons. Neuron 44, 759–767

52 Park, M. et al. (2006) Plasticity-induced growth of dendritic spines byexocytic trafficking from recycling endosomes. Neuron 52, 817–830

53 Antonova, I. et al. (2001) Rapid increase in clusters of presynapticproteins at onset of long-lasting potentiation. Science 294, 1547–1550

54 Colicos, M.A. et al. (2001) Remodeling of synaptic actin induced byphotoconductive stimulation. Cell 107, 605–616

55 Nikonenko, I. et al. (2003) Presynaptic remodeling contributes toactivity-dependent synaptogenesis. J. Neurosci. 23, 8498–8505

56 Zhai, R.G. et al. (2001) Assembling the presynaptic active zone: acharacterization of an active zone precursor vesicle. Neuron 29, 131–143

57 Garner, C.C. et al. (2002) Molecular mechanisms of CNSsynaptogenesis. Trends Neurosci. 25, 243–251

58 Tanaka, H. et al. (2000) Molecular modification of N-cadherin inresponse to synaptic activity. Neuron 25, 93–107

59 Granseth, B. et al. (2006) Clathrin-mediated endocytosis is thedominant mechanism of vesicle retrieval at hippocampal synapses.Neuron 51, 773–786

60 Star, E.N. et al. (2005) Real-time imaging of Rab3a and Rab5a revealsdifferential roles in presynaptic function. J. Physiol. 569, 103–117

61 Sankaranarayanan, S. et al. (2003) Actin has a molecular scaffolding,not propulsive, role in presynaptic function. Nat. Neurosci. 6, 127–135

62 Mueller, V.J. et al. (2004) Monitoring clathrin-mediated endocytosisduring synaptic activity. J. Neurosci. 24, 2004–2012

63 Atwood, H.L. and Karunanithi, S. (2002) Diversification of synapticstrength: presynaptic elements. Nat. Rev. Neurosci. 3, 497–516

64 Lippincott-Schwartz, J. and Patterson, G.H. (2003) Development anduse of fluorescent protein markers in living cells. Science 300, 87–91

65 Reits, E.A. and Neefjes, J.J. (2001) From fixed to FRAP: measuringprotein mobility and activity in living cells. Nat. Cell Biol. 3, E145–E147

66 Patterson, G.H. and Lippincott-Schwartz, J. (2002) A photoactivatableGFP for selective photolabeling of proteins and cells. Science 297,1873–1877

67 Ando, R. et al. (2002) An optical marker based on the UV-inducedgreen-to-red photoconversion of a fluorescent protein. Proc. Natl. Acad.Sci. U. S. A. 99, 12651–12656

68 Gurskaya, N.G. et al. (2006) Engineering of a monomeric green-to-redphotoactivatable fluorescent protein induced by blue light. Nat.Biotechnol. 24, 461–465

eloping countries

e launched the Health InterNetwork Access to

world’s poorest countries to gain free access

through the internet.

ourt Worldwide STM group, Wolters Kluwer

nd John Wiley, were approached by the WHO

ore than 1500 journals were made available for

ies, medical schools, and research and public

ditional publishers joined, and more than 2000

0 publishers are participating in the program.

f the WHO, said that this initiative was ‘‘perhaps

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es’’.

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