Synaptic vesicles (SVs) are small, membrane-bound organelles that are found in the synaptic terminal of neurons, and which are crucial in neurotransmission. After a rise in internal [Ca(2+)] during neuronal stimulation, SVs fuse with the plasma membrane releasing their neurotransmitter content, which then signals neighboring neurons. SVs are subsequently recycled and refilled with neurotransmitter for further rounds of release. Recently, tremendous progress has been made in elucidating the molecular composition of SVs, as well as putative protein-protein interactions. However, what is lacking is an empirical description of SV structure at the supramolecular level-which is necessary to enable us to fully understand the processes of membrane fusion, retrieval, and recycling. Using small-angle x-ray scattering, we have directly investigated the size and structure of purified SVs. From this information, we deduced detailed size and density parameters for the protein layers responsible for SV function, as well as information about the lipid bilayer. To achieve a convincing model fit, a laterally anisotropic structure for the protein shell is needed, as a rotationally symmetric density profile does not explain the data. Not only does our model confirm many of the preexisting ideas concerning SV structure, but also for the first time, to our knowledge, it indicates structural refinements, such as the presence of protein microdomains.

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We discuss different spherically symmetric and anisotropic form factor models and test them against high resolution synchrotron based small-angle x-ray scattering (SAXS) data from synaptic vesicles (SVs), isolated from rat brain. Anisotropy of the model form factors is found to be a key ingredient for the description of the native synaptic vesicle structure. We describe changes in structural parameters due to protease digestion of SVs, and present SAXS data of SVs recorded under different pH conditions.

Abstract. The size polydispersity distribution of synaptic vesicles (SVs) is characterized under quasiphysiological conditions by dynamic light scattering (DLS). Highly purified fractions of SVs obtained from rat brain still contain a small amount of larger contaminant structures, which can be quantified by DLS and further reduced by asymmetric-flow field-flow (AFFF) fractionation. The intensity autocorrelation functions g 2(τ ) recorded from these samples are analyzed by a constrained regularization method as well as by an alternative direct modeling approach. The results are in quantitative agreement with the polydispersity obtained from cryogenic electron microscopy of vitrified SVs. Next, different vesicle fusion assays based on samples composed of SVs and small unilamellar proteoliposomes with the fusion proteins syntaxin 1 and SNAP-25A are characterized by DLS. The size increase of the proteoliposomes due to SNARE-dependent fusion with SVs is quantified by DLS under quasi-physiological conditions.

Synaptic vesicles (SVs) are small, membrane-bound organelles that are found in the synaptic terminal of neurons. Although tremendous progress has been made in understanding the protein machinery that drives fusion of SVs with the presynaptic membrane, little progress has been made in understanding changes in the membrane structure that accompany this process. We used lipid monolayers of defined composition to mimic biological membranes, which were probed by x-ray reflectivity and grazing incidence x-ray diffraction. These techniques allowed us to successfully monitor structural changes in the membranes at molecular level, both in response to injection of SVs in the subphase below the monolayer, as well as to physiological cues involved in neurotransmitter release, such as increases in the concentration of the membrane lipid PIP(2), or addition of physiological levels of Ca(2+). Such structural changes may well modulate vesicle fusion in vivo.

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Solid supported lipid bilayers are highly useful model systems for cell membranes. However, the close proximity of the membrane and the solid support may adversely impact protein structure and mobility. One approach to screen the membrane from the underlying support and to better mimic the cellular cytoskeleton and the extracellular matrix is to fabricate a water rich polymer