Disturbances of soluble N-ethylmaleimide-sensitive factor attachment proteins in hippocampal synaptosomes contribute to cognitive impairment after repetitive formaldehyde inhalation in male rats

SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein) complex, a four-helical bundle composed of syntaxin1 and synaptosome-associated protein 25 (SNAP25) on the plasma membrane and synaptobrevin/VAMP2 (vesicle-associated membrane protein 2) on the vesicle membrane, plays a key role in synaptic exocytosis and facilitates neurotransmission. Disturbances of SNARE proteins were uncovered in some neurodegenerative diseases, neuroendocrine disturbances and even after environmental interventions. In the present study, we evaluated the effects of formaldehyde (FA) inhalation (13.5±1.5 ppm, twice 30-min each day for two rounds of 14 consecutive days) on learning and memory in Morris water maze and thereafter explored the SNARE protein levels in hippocampal synaptosomes. The formaldehyde-treated rats showed learning and memory impairment in escape latency and probe trials, without mobility disturbances in Morris water maze. Using western blotting assays, we detected the SNARE proteins in hippocampal synaptosomes and identified decrease of both SNAP25 and VAMP2 after formaldehyde treatment without significant changes of another SNARE protein, syntaxin 1, and synaptic vesicle marker, synaptophysin. Furthermore, the neuronal morphology and number detected in Nissl stain and western blotting assay of neurofilament-150 and synaptophysin were not affected after FA treatment. These results suggested that the specific decrease of SNAP25 and VAMP2 in hippocampal synaptosomes served as a potential contributing mechanism underlying learning and memory impairments after repetitive formaldehyde inhalation treatment.     

Vesicle-Associated Membrane Protein 4 and Syntaxin 6 Interactions at the Chlamydial Inclusion

The predominant players in membrane fusion events are the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) family of proteins. We hypothesize that SNARE proteins mediate fusion events at the chlamydial inclusion and are important for chlamydial lipid acquisition. We have previously demonstrated that trans-Golgi SNARE syntaxin 6 localizes to the chlamydial inclusion. To investigate the role of syntaxin 6 at the chlamydial inclusion, we examined the localization and function of another trans-Golgi SNARE and syntaxin 6-binding partner, vesicle-associated membrane protein 4 (VAMP4), at the chlamydial inclusion. In this study, we demonstrate that syntaxin 6 and VAMP4 colocalize to the chlamydial inclusion and interact at the chlamydial inclusion. Furthermore, in the absence of VAMP4, syntaxin 6 is not retained at the chlamydial inclusion. Small interfering RNA (siRNA) knockdown of VAMP4 inhibited chlamydial sphingomyelin acquisition, correlating with a log decrease in infectious progeny. VAMP4 retention at the inclusion was shown to be dependent on de novo chlamydial protein synthesis, but unlike syntaxin 6, VAMP4 recruitment is observed in a species-dependent manner. Notably, VAMP4 knockdown inhibits sphingomyelin trafficking only to inclusions in which it localizes. These data support the hypothesis that VAMP proteins play a central role in mediating eukaryotic vesicular interactions at the chlamydial inclusion and, thus, support chlamydial lipid acquisition and chlamydial development.     

SNARE和synaptotagmin对学习记忆作用的研究进展

可溶性NSF附着蛋白受体(SNARE)复合物是神经元囊泡胞吐过程重要的一部分,一般认为它参与调节突触前膜融合过程。SNARE复合体核心蛋白主要有3个,突触小体相关蛋白(SNAP-25),syntaxin和囊泡相关蛋白(VAMP)。突触结合蛋白(synaptotagmin,syt)是一个钙感受器蛋白,通过和SNARE结合起到调控膜融合过程的作用。SNARE复合物中的syntaxin,SNAP-25以及syt基因的缺失可造成小鼠认知以及学习记忆损伤。本文主要综述两者在突触前膜的作用过程,并分析其对学习记忆的影响与其调节突触前膜融合过程密切相关,以期了解syntaxin,SNAP-25及syt影响学习记忆的分子机制。     

Snapin: a SNARE-associated protein implicated in synaptic transmission

Synaptic vesicle docking and fusion are mediated by the assembly of a stable SNARE core complex of proteins, which include the synaptic vesicle membrane protein VAMP/synaptobrevin and the plasmalemmal proteins syntaxin and SNAP-25. We have now identified another SNAP-25-binding protein, called Snapin. Snapin was enriched in neurons and exclusively located on synaptic vesicle membranes. It associated with the SNARE complex through direct interaction with SNAP-25. Binding of recombinant Snapin-CT to SNAP-25 blocked the association of the SNARE complex with synaptotagmin. Introduction of Snapin-CT and peptides containing the SNAP-25 binding sequence into presynaptic superior cervical ganglion neurons in culture reversibly inhibited synaptic transmission. These results suggest that Snapin is an important component of the neurotransmitter release process through its modulation of the sequential interactions between the SNAREs and synaptotagmin.     

Role of snare proteins in CFTR and ENaC trafficking

The apical membrane ion channels, CFTR and ENaC, undergo regulated trafficking as a means of controlling their plasma membrane density. This provides a mechanism for regulating the Cl and Na conductance properties of epithelial apical membranes, and thus the transepithelial ion transport rates. Physical and functional interactions between these channels and SNARE proteins, in particular syntaxin 1A (S1A), provide a mechanism for linking the known vesicle fusion machinery with this process. In this paper we summarize evidence indicating that the interaction of S1A with CFTR and ENaC reduces channel currents in a syntaxin-isoform-specific manner. The acute cAMP-regulated CFTR trafficking event, which is reported by an increase in membrane capacitance in response to cAMP, is also inhibited by exogenous S1A expression. We tagged both channels with flag epitopes on their extracellular surfaces to monitor their plasma membrane expression as a function of S1A co-expression. The data indicate that the reduction in current caused by S1A is associated with a marked decrease in the amount of CFTR or ENaC detected at the cell surface. These findings suggest that S1A inhibits ion channel insertion into the plasma membrane, either by disrupting the stoichiometry of SNARE protein associations that mediate channel trafficking, or by physically associating with the channels to prevent their insertion. These data link the SNARE machinery to the regulation of apical membrane ion channel density, and suggest that phosphorylation-dependent interactions of these channels with SNARE proteins may acutely regulate this process.     

Regulation of syntaxin1A–munc18 complex for SNARE pairing in HEK293 cells

The formation and dissolution of SNARE protein complexes is essential for Ca²⁺-triggered fusion of neurotransmitter-filled vesicles at the presynaptic membrane. Among the pre-synaptic SNARE proteins, the activation of the Q-SNARE syntaxin1A is a critical event for SNARE complex formation. Activation requires syntaxin1A to transit from a munc18-bound non-interacting state to one competent for SNARE binding. The molecular mechanisms that regulate this transition remain unclear. The propensity of syntaxin1A to promote voltage-dependent steady-state inactivation of N-type Ca²⁺ channels and accelerate their entry into inactivation was used in a heterologous cell expression system to elucidate regulation of syntaxin1A protein–protein interactions. We report that coexpression of munc18 eliminated the promoting effect of syntaxin1A on inactivation. This effect of munc18 was completely disrupted by coexpression of munc13-1, but not munc13-2 or munc13-3. Also, since expression of munc13-1 with syntaxin1A resulted in an inactivation phenotype identical to that of munc18 with syntaxin1A, the action of munc13-1 on the munc18–syntaxin1A complex was functionally unique and did not result from competitive binding interactions. Furthermore, munc13 expressed with syntaxin1A and munc18 promoted redistribution of a cytosolic SNAP25 mutant to the membrane, a result indicative of syntaxin1A–SNAP25 SNARE pairing. These data demonstrate an important role of munc13 to control the protein–protein interactions of syntaxin1A in vivo, and support munc13 as critical to dissociating syntaxin1A–munc18 complexes and making syntaxin1A available for SNARE interactions.     

Immunohistochemical Localization of SNARE Proteins in Dental Pulp and Periodontal Ligament of the Rat Incisor

Distribution of three soluble N‐ethylmaleimide‐sensitive fusion protein attachment protein receptor (SNARE) proteins, syntaxin‐1, synaptosomal‐associated protein of 25 kDa (SNAP‐25), and vesicle‐associated membrane protein‐2 (VAMP‐2), was examined in dental pulp and periodontal ligament of the rat incisor. In the trigeminal ganglion, syntaxin‐1 and SNAP‐25 immunoreactivity was predominately detected in medium‐ to large‐sized neurons. Most syntaxin‐1 immunoreactive neurons expressed SNAP‐25. In contrast, VAMP‐2 was localized in small‐ to medium‐sized neurons and in slender‐shaped cells surrounding SNAP‐25‐immunopositive neurons. When the inferior alveolar nerve, one of the mandibular nerve branches innervating the dental pulp and periodontal ligament, was ligated, SNARE proteins accumulated at the site proximal to the ligation. In the incisor dental pulp, all nerve fibers displayed immunoreactivity for syntaxin‐1, SNAP‐25, and VAMP‐2. In the periodontal ligament of the incisor, almost all nerve fibers displayed both syntaxin‐1 and SNAP‐25 immunoreactivity, but lacked VAMP‐2 immunoreactivity. SNAP‐25 protein expression was localized around the vesicle membranes at the axon terminal of the periodontal mechanoreceptors. These present data suggest that these three SNARE proteins are synthesized at the trigeminal ganglion, transported centrally and peripherally, and expressed in sensory endings where apparent synapses are not present. Because those proteins participate in docking and exocytosis of synapse vesicles in the central nervous system, they might also contribute to vesicle exocytosis at receptive fields where apparent synapses are not present. Anat Rec, 2010. © 2010 Wiley‐Liss, Inc.     

血栓烷A_2类似物对大鼠肺动脉血管平滑肌细胞钾离子通道表达的影响

目的:观察血栓烷A2(TXA2)类似物U46619对实验大鼠肺动脉血管平滑肌细胞(PASMCs)K+通道Kv1.2、Kv1.5、Kv2.1蛋白质和mRNA表达的影响。方法:采用酶法分离、培养Wistar大鼠PASMCs,通过Western-blot和RT-PCR方法分别从蛋白质水平和mRNA水平分析U46619对Kv1.2、Kv1.5、Kv2.1表达的抑制作用。结果:100nmol/LU46619对Kv1.2表达有明显抑制作用。结论:TXA2类似物U46619可能通过抑制Kv1.2表达而参与大鼠肺动脉血管的收缩。     

慢性低氧对大鼠肺组织电压门控钾通道表达的影响

目的:探讨慢性低氧对大鼠肺组织电压门控钾通道亚型Kv1.5、Kv2.1、Kv9.3表达的影响。方法:将12只Wistar大鼠随机分为对照组和慢性低氧组,每组6只。采用RT-PCR和Westernblot对大鼠肺组织匀浆中Kv1.5、Kv2.1、Kv9.3mRNA和Kv1.5蛋白质的表达进行观察。结果:慢性低氧组大鼠肺组织匀浆中Kv1.5、Kv2.1、Kv9.3mRNA和Kv1.5蛋白质表达明显低于对照组(P<0.01)。结论:慢性低氧可以从转录、翻译两个水平抑制大鼠肺组织中Kv1.5、Kv2.1Kv9.3的表达,Kv1.5、Kv2.1、Kv9.3表达的减少必将导致Kv数目的减少和电流的降低。Kv1.5、Kv2.1、Kv9.3可能是氧敏感钾通道亚型。     

Kv1.2、Kv1.5、Kv2.1钾通道在缺氧性肺血管收缩中的作用

目的和方法:雄性Wistar大鼠随机分为两组:常氧对照组和低氧组。用酶消化的方法获得单个大鼠肺内动脉平滑肌细胞(PASMC)。采用全细胞膜片钳技术,记录PASMC静息膜电位(Em)和电压门控性钾通道电流(IKv),通过细胞内灌流Kv1.2/Kv1.5/Kv2.1抗体混合液(1∶125),探讨Kv1.2、Kv1.5、Kv2.1钾通道在缺氧性肺血管收缩(HPV)中的作用。结果:①低氧组膜电位明显去极化,由(-51.8±0.8) mV 去极到(-47.2± 0.7) mV,P<0.01,IKv与常氧组相比显著降低,在测试电压-30 mV时, IKv由(6.16±0.58) pA/pF 降为 (3.31±0.37) pA/pF (P<0.01)。②细胞内灌流Kv1.2/Kv1.5/Kv2.1抗体混合液可显著抑制常氧对照组PASMC 的IKv,使Em去极化,然而细胞内灌流Kir2.1/Kir2.3/Kir4.1(1∶125)抗体混合液对常氧对照组PASMC 的IKv和Em无显著影响。③细胞内灌流Kv1.2/Kv1.5/Kv2.1抗体混合液和Kir2.1/Kir2.3/Kir4.1抗体混合液对低氧组PASMC的IKv和Em均无显著影响。结论:Kv1.2、Kv1.5、Kv2.1可能是氧敏感型通道,并介导了低氧性肺血管收缩。     

The septin CDCrel-1 binds syntaxin and inhibits exocytosis.

Septins are GTPases required for the completion of cytokinesis in diverse organisms, yet their roles in cytokinesis or other cellular processes remain unknown. Here we describe studies of a newly identified septin, CDCrel-1, which is predominantly expressed in the nervous system. This protein was associated with membrane fractions, and a significant fraction of the protein copurified and coprecipitated with synaptic vesicles. In detergent extracts, CDCrel-1 and another septin, Nedd5, immunoprecipitated with the SNARE protein syntaxin by directly binding to syntaxin via the SNARE interaction domain. Transfection of HIT-T15 cells with wild-type CDCrel-1 inhibited secretion, whereas GTPase dominant-negative mutants enhanced secretion. These data suggest that septins may regulate vesicle dynamics through interactions with syntaxin.     

Neurotrophin B receptor kinase increases Kv subfamily member 1.3 (Kv1.3) ion channel half-life and surface expression

Kv subfamily member 1.3 (Kv1.3), a member of the Shaker family of potassium channels, has been found to play diverse roles in immunity, metabolism, insulin resistance, sensory discrimination, and axonal targeting in addition to its traditional role in the stabilization of the resting potential. We demonstrate that the neurotrophin B receptor (TrkB) causes an upregulation of Kv1.3 ion channel protein expression in the absence of the preferred ligand for the receptor (brain-derived neurotrophic factor; BDNF) and oppositely downregulates levels of Kv subfamily member 1.5. Although the effect occurs in the absence of the ligand, Kv1.3 upregulation by TrkB is dependent upon the catalytic domain of the TrkB kinase as well as tyrosine (Y) residues in the N and C terminus of the Kv1.3 channel. Using pulse-chase experiments we find that TrkB alters the half-life residence of the channel by approximately 2x and allows it to sustain activity as reflected in an increased current magnitude without alteration of kinetic properties. TrkB and Kv1.3 co-immunoprecipitate from tissue preparations of the mouse olfactory bulb and olfactory cortex, and by immunocytochemical approaches, are found to be co-localized in the glomerular, mitral cell, and internal plexiform layers of the olfactory bulb. These data suggest that Kv1.3 is not only modulated by direct phosphorylation in the presence of BDNF-activated TrkB kinase, but also may be fine tuned via regulation of surface expression while in the proximity of neurotrophic factor receptors. Given the variability of TrkB expression during development, regeneration, or neuronal activation, modulation of surface expression and turnover of Kv channels could significantly impact neuronal excitability, distinct from that of tyrosine kinase phosphorylation.     

脂质体转染Kv1.5反义寡核苷酸（AsOND）对人气道平滑肌Kv活性的影响

目的： 研究人气道平滑肌细胞（HASMCs）转染Kv1.5反义寡核苷酸（AsOND）后，电压依赖延迟整流钾通道（Kv）的活性变化，探讨其基因亚型Kv1.5在调节Kv活性中的作用。方法： 采用脂质体转染、逆转录聚合酶链反应（RT-PCR）、Western blotting和全细胞膜片钳技术，观察转染Kv1.5 AsOND后，HASMCs Kv1.5 mRNA及蛋白表达的变化，以及转染对HASMCs Kv活性的影响。 结果： 脂质体转染Kv1.5 AsOND后，HASM细胞Kv1.5 mRNA和蛋白质的表达均下降；Kv电流值受到显著抑制；使细胞膜电位（E_m）趋向于去极化方向。 结论： 转染Kv1.5 AsOND可导致HASMCs Kv功能降低，Kv1.5基因亚型在调节Kv活性中可能起重要作用。     

Involvement of vesicle-associated membrane protein 7 in human gastric epithelial cell vacuolation induced by Helicobacter pylori-produced VacA

Helicobacter pylori-produced cytotoxin VacA induces intracellular vacuolation. The VacA-induced vacuole is assumed to represent the pathological status of intracellular trafficking. The fusion mechanism of the endosomes requires the formation of a tight complex between the Q-SNAREs and the R-SNAREs. We recently reported that syntaxin 7, a family member of the Q-SNARE protein, is involved in VacA-induced vacuole formation. In order to further elucidate the molecular mechanism, we identified the participation of vesicle-associated membrane protein 7 (VAMP7) as a partner of syntaxin 7. Immunocytochemistry revealed endogenous VAMP7 to be localized to the vacuoles induced by VacA. A Northern blotting study demonstrated that VacA intoxication increased VAMP7 mRNA in a time-dependent manner. VAMP7 was coimmunoprecipitated with syntaxin 7, and the amounts of endogenous VAMP7 and syntaxin 7 bound to syntaxin 7 and VAMP7, respectively, increased in response to VacA. The down-regulation of VAMP7 using small interfering RNA inhibited VacA-induced vacuolation, and the transient transfection of dominant-negative mutant VAMP7, the N-terminal domain of VAMP7, also inhibited the vacuolation. We therefore conclude that R-SNARE VAMP7 plays an important role in VacA-induced vacuolation as a partner of Q-SNARE syntaxin 7.     

Aberrant localization of fusion receptors involved in regulated exocytosis in salivary glands of Sjögren's syndrome patients is linked to ectopic mucin secretion

Sj?gren's syndrome (SS) is a chronic inflammatory autoimmune disease that mainly affects tear and salivary glands, whereby SS-patients frequently complain of eye and mouth dryness. Salivary acinar cells of SS-patients display alterations in their cell polarity; which may affect the correct localization and function of proteins involved in regulated exocytosis. Here we determined whether the expression and localization of SNARE proteins (membrane fusion receptors) involved in regulated secretion, such as VAMP8, syntaxin 3 (STX3), STX4 and SNAP-23 were altered in salivary glands (SG) from SS-patients. Additionally, we investigated SNARE proteins function, by evaluating their ability to form SNARE complexes under basal conditions. In SG from SS-patients and control subjects mRNA and proteins levels of SNARE complex components were determined by real-time PCR and Western blotting, respectively. SNARE protein distribution and mucin exocytosis were determined by indirect immunofluorescence. In SS-patients, the expression levels of mRNA and protein for VAMP8, STX4 and STX3 were altered. STX4, STX3, SNAP-23 and VAMP8 relocated from the apical to the basal region of acinar cells. Increased formation of SNARE complexes in a manner independent of external stimuli for secretion was detected. Mucins were detected in the extracellular matrix (ECM). Presence of mucins in the ECM, together with the observed alterations in SNARE protein localization is indicative of ectopic exocytosis. In the context of SS, such aberrantly localized mucins are likely to favor a pro-inflammatory response, which may represent an important initial step in the pathogenesis of this disease.     

Interactions between the C-terminus of Kv1.5 and Kvβ regulate pyridine nucleotide-dependent changes in channel gating

Voltage-gated potassium (Kv) channels are tetrameric assemblies of transmembrane Kv proteins with cytosolic N- and C-termini. The N-terminal domain of Kv1 proteins binds to β-subunits, but the role of the C-terminus is less clear. Therefore, we studied the role of the C-terminus in regulating Kv1.5 channel and its interactions with Kvβ-subunits. When expressed in COS-7 cells, deletion of the C-terminal domain of Kv1.5 did not affect channel gating or kinetics. Coexpression of Kv1.5 with Kvβ3 increased current inactivation, whereas Kvβ2 caused a hyperpolarizing shift in the voltage dependence of current activation. Inclusion of NADPH in the patch pipette solution accelerated the inactivation of Kv1.5-Kvβ3 currents. In contrast, NADP⁺ decreased the rate and the extent of Kvβ3-induced inactivation and reversed the hyperpolarizing shift in the voltage dependence of activation induced by Kvβ2. Currents generated by Kv1.5ΔC+Kvβ3 or Kv1.5ΔC+Kvβ2 complexes did not respond to changes in intracellular pyridine nucleotide concentration, indicating that the C-terminus is required for pyridine nucleotide-dependent interactions between Kvβ and Kv1.5. A glutathione-S-transferase (GST) fusion protein containing the C-terminal peptide of Kv1.5 did not bind to apoKvβ2, but displayed higher affinity for Kvβ2:NADPH than Kvβ2:NADP⁺. The GST fusion protein also precipitated Kvβ proteins from mouse brain lysates. Pull-down experiments, structural analysis and electrophysiological data indicated that a specific region of the C-terminus (Arg543-Val583) is required for Kvβ binding. These results suggest that the C-terminal domain of Kv1.5 interacts with β-subunits and that this interaction is essential for the differential regulation of Kv currents by oxidized and reduced nucleotides.     

Protein-protein interactions in neurotransmitter release

The arrival of a nerve impulse at a nerve terminal leads to the opening of voltage-gated Ca(2+) channels and a rapid influx of Ca(2+). The increase in Ca(2+) concentration at the active zone from the basal level of 100-200 mM triggers the fusion of docked synaptic vesicles, resulting in neurotransmitter release. A large number of proteins have been identified at nerve terminals and a cascade of protein-protein interactions has been suggested to be involved in the cycling of synaptic vesicle states. Functional studies in last half decade on synaptic-terminal proteins, including Ca(2+) channels, have revealed that the SNARE core complex, consisting of synaptobrevin VAMP, a synaptic vesicle-associated protein, syntaxin and SNAP-25, synaptic membrane-associated proteins, acts as the membrane fusion machinery and that proteins interacting with the SNARE complex play essential roles in synaptic vesicle exocytosis by regulating assembly and disassembly of the SNARE complex.     

Syntaxin 1A Co-Associates with Native Rat Brain and Cloned Large Conductance, Calcium-Activated Potassium Channels in situ

Large conductance, calcium-activated potassium channels (BK_Ca channels) are regulated by several distinct mechanisms, including phosphorylation/dephosphorylation events and protein-protein interactions. In this study, we have examined the interaction between BK_Ca channels and syntaxin 1A, a soluble N -ethylmaleimide-sensitive factor attachment protein receptor (SNARE) that is reported to modulate the activity and/or localization of different classes of ion channels. Using a reciprocal co-immunoprecipitation strategy, we observed that native BK_Ca channels in rat hippocampus co-associate with syntaxin 1A, but not the closely related homologue syntaxin 3. This BK_Ca channel-syntaxin 1A interaction could be further demonstrated in a non-neuronal cell line (human embryonic kidney (HEK) 293 cells) following co-expression of rat syntaxin 1A and BK_Ca channels cloned from either mouse brain or bovine aorta. However, co-expression of these same channels with syntaxin 3 did not lead to a detectable protein-protein interaction. Immunofluorescent co-staining of HEK 293 cells expressing BK_Ca channels and syntaxin 1A demonstrated overlapping distribution of these two proteins in situ . Functionally, co-expression of BK_Ca channels with syntaxin 1A, but not syntaxin 3, was observed to enhance channel gating and kinetics at low concentrations (1–4 μ m ) of free cytosolic calcium, but not at higher concentrations (≤ 10 μ m ), as judged by macroscopic current recordings in excised membrane patches. Interactions between BK_Ca channels and neighbouring membrane proteins may thus play important roles in regulating the activity and/or distribution of these channels within specialized cellular compartments.     

Single molecule measurements of mechanical interactions within ternary SNARE complexes and dynamics of their disassembly: SNAP25 vs. SNAP23

Regulated exocytosis is a crucial event for intercellular communication between neurons and astrocytes within the CNS. The soluble N -ethylmaleimide-sensitive fusion protein attachment protein receptor (SNARE) complex, composed of synaptobrevin 2, syntaxin and synaptosome-associated protein of 25 kDa or 23 kDa (SNAP25 or SNAP23), is essential in this process. It was reported that SNAP25 and SNAP23 have distinct roles in exocytotic release, where SNAP25, but not SNAP23, supports an exocytotic burst. It is not clear, however, whether this is due to the intrinsic properties of the ternary SNARE complex, containing either SNAP25 or SNAP23, or perhaps due to the differential association of these proteins with ancillary proteins to the complex. Here, using force spectroscopy, we show from single molecule investigations of the SNARE complex, that SNAP23A created a local interaction at the ionic layer by cuffing syntaxin 1A and synaptobrevin 2, similar to the action of SNAP25B; thus either of the ternary complexes would allow positioning of vesicles at a maximal distance of ∼13 nm from the plasma membrane. However, the stability of the ternary SNARE complex containing SNAP23A is less than half of that for the complex containing SNAP25B. Thus, differences in the stability of the two different ternary complexes could underlie some of the SNAP25/23 differential ability to control the exocytotic burst.