Institute of Neuroscience Faculty

Information is exchanged between neurons at synapses, which are essentially specialized sites of cell-cell adhesion . A mature synapse is defined as an accumulation of synaptic vesicles within the axon, in close apposition to a dendritic membrane studded with receptors (see figure)which are held in place by a submembranous scaffold (Sheng and Kim, 2002) . The formation of such an intercellular structure requires spatially and temporally controlled changes in morphology and molecular content at sites of contacts. Recent advances in subcellular fluorescence microscopy have revealed that this process involves the rapid recruitment and stabilization of both pre- and postsynaptic elements. These studies have shown that major components of the synaptic vesicle and active zone machinery travel in clusters together with other presynaptic proteins, such as calcium channels, and are rapidly recruited to new sites of contact (Ahmari et al., 2000; Zhai et al., 2001; Washbourne et al., 2002) .

On the postsynaptic side, receptor subunits and components of the scaffold or post-synaptic density (PSD) are recruited separately and with distinct time courses within minutes to hours after initial contact (Friedman et al., 2000; Bresler et al., 2001; Washbourne et al., 2002; Bresler et al., 2004)

Despite these advances the basic mechanisms by which synapse formation is induced at discrete locations and by which the molecular machinery is recruited to sites of contact remain elusive. We are currently using both mammalian primary neuronal cultures and zebrafish embryos to investigate molecules that are involved in the mechanisms of synapse formation. Techniques currently employed are live confocal imaging of fluorescently-tagged synaptic components, electron microscopy, biochemistry and molecular biology.

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Representative Publications

• Washbourne, P., Schiavo, G. and Montecucco, C. (1995). Vesicle-associated membrane protein-2 (synaptobrevin-2) forms a complex with synaptophysin. Biochem J 305 ( Pt 3), 721-4.

• Washbourne, P., Pellizzari, R., Baldini, G., Wilson, M. C. and Montecucco, C. (1997). Botulinum neurotoxin types A and E require the SNARE motif in SNAP-25 for proteolysis. FEBS Lett 418, 1-5.

• Washbourne, P., Pellizzari, R., Rossetto, O., Bortoletto, N., Tugnoli, V., De Grandis, D., Eleopra, R. and Montecucco, C. (1998). On the action of botulinum neurotoxins A and E at cholinergic terminals. J Physiol Paris 92, 135-9.

• Washbourne, P., Bortoletto, N., Graham, M. E., Wilson, M. C., Burgoyne, R. D. and Montecucco, C. (1999). Botulinum neurotoxin E-insensitive mutants of SNAP-25 fail to bind VAMP but support exocytosis. J Neurochem 73, 2424-33.

• Graham, M. E., Washbourne, P., Wilson, M. C. and Burgoyne, R. D. (2001). SNAP-25 with mutations in the zero layer supports normal membrane fusion kinetics. J Cell Sci 114, 4397-405.

• Washbourne, P., Cansino, V., Mathews, J. R., Graham, M., Burgoyne, R. D. and Wilson, M. C. (2001). Cysteine residues of SNAP-25 are required for SNARE disassembly and exocytosis, but not for membrane targeting. Biochem J 357, 625-34.

• Washbourne, P., Thompson, P.M., Carta, M., Costa, E.M., Mathews, J.R., Lopez-Bendito, G., Molnar, Z., Becher, M.W., Valenzuela, C.F., Partridge, L.D. and Wilson, M.C. (2002) “Genetic ablation of the t-SNARE SNAP-25 distinguishes mechanisms of neuroexocytosis” Nature Neuroscience 5: 19-26.
• Washbourne, P., Bennett, J. and McAllister, A.K.(2002) “Rapid recruitment of NMDA receptor transport packets to nascent synapses” Nature Neuroscience 5: 751-759

• Fu, Z., Washbourne, P., Ortinski, P. and Vicini, S. (2003). Functional excitatory synapses in HEK293 cells expressing neuroligin and glutamate receptors. J Neurophysiol 90, 3950-7.

• Washbourne P. (2004) Greasing transmission; palmitoylation at the synapse. Neuron, 44(6): 901-2, Dec. 16, 2004
• Washbourne P, Dityatev A, Scheiffele P, Biederer T, Weiner JA, Christopherson KS, El-Husseini A. (2004) Cell adhesion molecules in synapse formation. J Neurosci. 24(42): 9244-9, Oct. 20, 2004
• Washbourne, P., Liu, X-B., Jones, E.G. and McAllister, A.K. (2004) “Exo/Endocytic cycling of NMDA receptors during trafficking in neurons before synapse formation” J. Neuroscience, 24 (38) 8253-64, Sept 22, 2004
• Pietri T, Easley-Neal C, Wilson C, Washbourne P. (2008) Six cadm/synCAM genes are expressed in the nervous system of developing zebrafish. Devlopmental Dynamics. 237(1):233-46.

• Barrow, S.L.; Constable, J.R.L.; Clark, E.; El-Sabeawy, F.; McAllister, A.K. and Washbourne, P. (2009) Neuroligin1: a cell adhesion molecule that recruits PSD-95 and NMDARs by distinct mechanisms during synaptogenesis. Neural Dev. 18;4:17.  PMCID: PMC2694798.

• Pietri, T., Manalo, E., Ryan, J., Saint-Amant, L. and Washbourne P. (2009). Glutamate Drives the Touch Response through a Rostral Loop in the Spinal Cord of Zebrafish Embryos. Developmental Neurobiology 69, 780-795.

• Hoy, J., Constable, J.R.L., Vicini, S., Fu, Z. and Washbourne, P. (2009). SynCAM1 recruits NMDA receptors via protein 4.1B. Molecular and Cellular Neuroscience 42, 466-483.

• Davey, C., Tallafuss, A. and Washbourne, P. (2010) Differential expression of neuroligin genes in the nervous system of zebrafish. Developmental Dynamics 239, 703-714.

• Reviews and Book Chapters

• Pellizzari, R., Rossetto, O., Washbourne, P., Tonello, F., Nicotera, P. L. and Montecucco, C. (1998). In vitro biological activity and toxicity of tetanus and botulinum neurotoxins. Toxicol Lett 102-103, 191-7.

• Washbourne, P. “b-Leptinotarsin-h” in “Guidebook to Protein Toxins and their Use in Cell Biology”, editors Rappuoli R. and Montecucco, C., publishers Sambrook & Tooze, 182-183, 1998.

• Graham, M., Washbourne, P., Wilson, M.C. and Burgoyne, R.D. Molecular Analysis of SNAP-25 Function in Exocytosis”. Annals N.Y. Acad. Sci., 971, 210-221, 2002.

• Washbourne, P. McAllister, A.K. (2002). Techniques for gene transfer into neurons. Curr Opin Neurobio. 12, 566-73.

• Washbourne, P. (2004). Greasing transmission: palmitoylation at the synapse. Neuron 44, 901-2.

• Washbourne, P., Dityatev, A., Scheiffele, P., Biederer, T., Weiner, J. A., Christopherson, K. S. and El-Husseini, A. (2004). Cell adhesion molecules in synapse formation. J Neurosci 24, 9244-9.

• Washbourne, P. “Postsynaptic Transport Packets”(2006) Chapter in “Synapse Formation”, Editors: Dityatev, A. and El-Husseini, A., Springer Verlag.

• Tallafuss, A., Constable, J.R.L. and Washbourne, P. Organization of central synapses by adhesion molecules. Eur. J. Neuroscience, in press

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