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Transport, capture and exocytosis of single synaptic vesicles at active zones

Nature volume 406, pages 849–854 (2000)Cite this article

Abstract

To sustain high rates of transmitter release, synaptic terminals must rapidly re-supply vesicles to release sites and prime them for exocytosis. Here we describe imaging of single synaptic vesicles near the plasma membrane of live ribbon synaptic terminals. Vesicles were captured at small, discrete active zones near the terminal surface. An electric stimulus caused them to undergo rapid exocytosis, seen as the release of a fluorescent lipid from the vesicles into the plasma membrane. Next, vesicles held in reserve about 20 nm from the plasma membrane advanced to exocytic sites, and became release-ready 250 ms later. Apparently a specific structure holds vesicles at an active zone to bring v-SNAREs and t-SNAREs, the proteins that mediate vesicle fusion, within striking distance of each other, and then allows the triggered movement of such vesicles to the plasma membrane.

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Figure 1: Active zones on the terminal of a bipolar neuron.
Figure 2: Imaging single fusion events.
Figure 3: Size of the dye-containing organelle.
Figure 4: Kinetics of exocytosis.
Figure 5: Capture and escape of vesicles.
Figure 6: Stimulated approach of vesicles.

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References

  1. Liley, A. W. & North, K. A. K. An electrical investigation of effects of a repetitive stimulation on mammalian neuromuscular junction. J. Neurophysiol. 16, 509– 527 (1953).

    Article  CAS  Google Scholar 

  2. Zucker, R. S. Exocytosis: a molecular and physiological perspective. Neuron 17, 1049–1055 (1996).

    Article  CAS  Google Scholar 

  3. Neher, E. Vesicle pools and Ca2+ microdomains: new tools for understanding their roles in neurotransmitter release. Neuron 20, 389–399 (1998).

    Article  CAS  Google Scholar 

  4. Wu, L. G. & Borst, J. G. The reduced release probability of releasable vesicles during recovery from short-term synaptic depression. Neuron 23, 821–832 (1999).

    Article  CAS  Google Scholar 

  5. Schneggenburger, R., Meyer, A. C. & Neher, E. Released fraction and total size of a pool of immediately available transmitter quanta at a calyx synapse. Neuron 23, 399–409 (1999).

    Article  CAS  Google Scholar 

  6. Mennerick, S. & Matthews, G. Ultrafast exocytosis elicited by calcium current in synaptic terminals of retinal bipolar neurons. Neuron 17, 1241–1249 ( 1996).

    Article  CAS  Google Scholar 

  7. Sakaba, T., Tachibana, M., Matsui, K. & Minami, N. Two components of transmitter release in retinal bipolar cells: exocytosis and mobilization of synaptic vesicles. Neurosci. Res. 27, 357–370 (1997).

    Article  CAS  Google Scholar 

  8. von Gersdorff, H., Sakaba, T., Berglund, K. & Tachibana, M. Submillisecond kinetics of glutamate release from a sensory synapse. Neuron 21, 1177–1188 (1998).

    Article  CAS  Google Scholar 

  9. Neves, G. & Lagnado, L. The kinetics of exocytosis and endocytosis in the synaptic terminal of goldfish retinal bipolar cells. J. Physiol. (Lond.) 515, 181–202 (1999).

    Article  CAS  Google Scholar 

  10. Steyer, J. A., Horstmann, H. & Almers, W. Transport, docking and exocytosis of single secretory granules in live chromaffin cells. Nature 388, 474–478 (1997).

    Article  ADS  CAS  Google Scholar 

  11. Oheim, M., Loerke, D., Stuhmer, W. & Chow, R. H. The last few milliseconds in the life of a secretory granule. Docking, dynamics and fusion visualized by total internal reflection fluorescence microscopy (TIRFM). Eur. Biophys. J. 27, 83–98 (1998).

    Article  CAS  Google Scholar 

  12. Han, W., Ng, Y. K., Axelrod, D. & Levitan, E. S. Neuropeptide release by efficient recruitment of diffusing cytoplasmic secretory vesicles. Proc. Natl Acad. Sci. USA 96, 14577– 14582 (1999).

    Article  ADS  CAS  Google Scholar 

  13. Lang, T. et al. Role of actin cortex in the subplasmalemmal transport of secretory granules in PC-12 cells. Biophys. J. 78, 2863–2877 (2000).

    Article  ADS  CAS  Google Scholar 

  14. Schmoranzer, J., Goulian, M., Axelrod, D. & Simon, S. M. Imaging constitutive exocytosis with total internal reflection fluorescence microscopy. J. Cell Biol. 149, 23–32 (2000).

    Article  CAS  Google Scholar 

  15. Toomre, D., Steyer, J. A., Keller, P., Almers, W. & Simons, K. Fusion of constitutive membrane traffic with the cell surface observed by evanescent wave microscopy. J. Cell Biol. 149, 33–40 (2000).

    Article  CAS  Google Scholar 

  16. Betz, W. J. & Bewick, G. S. Optical analysis of synaptic vesicle recycling at the frog neuromuscular junction. Science 255, 200–203 (1992).

    Article  ADS  CAS  Google Scholar 

  17. Ryan, T. A., Reuter, H. & Smith, S. J. Optical detection of a quantal presynaptic membrane turnover. Nature 388, 478– 482 (1997).

    Article  ADS  CAS  Google Scholar 

  18. Murthy, V. N. & Stevens, C. F. Synaptic vesicles retain their identity through the endocytic cycle. Nature 392, 497–501 (1998).

    Article  ADS  CAS  Google Scholar 

  19. von Gersdorff, H., Vardi, E., Matthews, G. & Sterling, P. Evidence that vesicles on the synaptic ribbon of retinal bipolar neurons can be rapidly released. Neuron 16, 1221– 1227 (1996).

    Article  CAS  Google Scholar 

  20. Lagnado, L., Gomis, A. & Job, C. Continuous vesicle cycling in the synaptic terminal of retinal bipolar cells. Neuron 17, 957–967 (1996).

    Article  CAS  Google Scholar 

  21. Rouze, N. C. & Schwartz, E. A. Continuous and transient vesicle cycling at a ribbon synapse. J. Neurosci. 18, 8614–8624 (1998).

    Article  CAS  Google Scholar 

  22. Takei, K., Mundigl, O., Daniell, L. & De Camilli, P. The synaptic vesicle cycle: a single vesicle budding step involving clathrin and dynamin. J. Cell Biol. 133, 1237– 1250 (1996).

    Article  CAS  Google Scholar 

  23. Schmidt, A., Hannah, M. J. & Huttner, W. B. Synaptic-like microvesicles of neuroendocrine cells originate from a novel compartment that is continuous with the plasma membrane and devoid of transferrin receptor. J. Cell Biol. 137 , 445–458 (1997).

    Article  CAS  Google Scholar 

  24. Zhang, B. et al. Synaptic vesicle size and number are regulated by a clathrin adaptor protein required for endocytosis. Neuron 21 , 1465–1475 (1998).

    Article  CAS  Google Scholar 

  25. von Gersdorff, H. & Matthews, G. Inhibition of endocytosis by elevated internal calcium in a synaptic terminal. Nature 370, 652–655 ( 1994).

    Article  ADS  CAS  Google Scholar 

  26. Kobayashi, K. & Tachibana, M. Ca2+ regulation in the presynaptic terminals of goldfish retinal bipolar cells. J. Physiol. (Lond.) 483, 79–94 ( 1995).

    Article  CAS  Google Scholar 

  27. Smith, C. B. & Betz, W. J. Simultaneous independent measurement of endocytosis and exocytosis. Nature 380, 531–534 (1996).

    Article  ADS  CAS  Google Scholar 

  28. Betz, W. J., Mao, F. & Smith, C. B. Imaging exocytosis and endocytosis. Curr. Opin. Neurobiol. 6, 365–371 ( 1996).

    Article  CAS  Google Scholar 

  29. Betz, W. J. & Bewick, G. S. Optical monitoring of transmitter release and synaptic vesicle recycling at the frog neuromuscular junction. J. Physiol. (Lond.) 460, 287– 309 (1993).

    Article  CAS  Google Scholar 

  30. Gray, E. G. & Pease, H. L. On understanding the organisation of the retinal receptor synapses. Brain Res. 35, 1–15 (1971).

    Article  CAS  Google Scholar 

  31. Raviola, E. & Gilula, N. B. Intramembrane organization of specialized contacts in the outer plexiform layer of the retina. A freeze-fracture study in monkeys and rabbits. J. Cell Biol. 65, 192–222 (1975).

    Article  CAS  Google Scholar 

  32. Lenzi, D., Runyeon, J. W., Crum, J., Ellisman, M. H. & Roberts, W. M. Synaptic vesicle populations in saccular hair cells reconstructed by electron tomography. J. Neurosci. 19, 119–132 (1999).

    Article  CAS  Google Scholar 

  33. Issa, N. P. & Hudspeth, A. J. Clustering of Ca2+ channels and Ca(2+)-activated K+ channels at fluorescently labeled presynaptic active zones of hair cells. Proc. Natl Acad. Sci. USA 91, 7578–7582 (1994).

    Article  ADS  CAS  Google Scholar 

  34. Sutton, R. B., Fasshauer, D., Jahn, R. & Brunger, A. T. Crystal structure of a SNARE complex involved in synaptic exocytosis at 2.4 A resolution. Nature 395, 347–353 ( 1998).

    Article  ADS  CAS  Google Scholar 

  35. Lowe, M., Nakamura, N. & Warren, G. Golgi division and membrane traffic. Trends Cell Biol. 8, 40–44 ( 1998).

    Article  CAS  Google Scholar 

  36. Heidelberger, R. & Matthews, G. Calcium influx and calcium current in single synaptic terminals of goldfish retinal bipolar neurons. J. Physiol. (Lond.) 447, 235– 256 (1992).

    Article  CAS  Google Scholar 

  37. Stout, A. L. & Axelrod, D. Evanescent field excitation of fluorescence by epi-illumination microscopy. Appl. Opt. 28, 5237–5242 (1989).

    Article  ADS  CAS  Google Scholar 

  38. Steyer, J. A. & Almers, W. Tracking single secretory granules in live chromaffin cells by evanescent-field fluorescence microscopy. Biophys. J. 76, 2262–2271 (1999).

    Article  ADS  CAS  Google Scholar 

  39. Axelrod, D., Hellen, E. H. & Fulbright, R. M. in Topics in Fluorescence Spectroscopy, Volume 3: Biomedical Applications (ed. Lakowicz, J. R.) 289– 343 (Plenum, New York, 1992).

    Google Scholar 

  40. Parsons, T. D., Coorssen, J. R., Horstmann, H. & Almers, W. Docked granules, the exocytic burst, and the need for ATP hydrolysis in endocrine cells. Neuron 15, 1085– 1096 (1995).

    Article  CAS  Google Scholar 

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Acknowledgements

This work was spported by a grant from the NIH (W.A.) and by the Max Planck Society. D.Z. was supported by an NIH Training Program in Neuronal Signaling and J.A.S. by a fellowship from the Max Planck Society. We thank M. Feldman for his comments on the manuscript.

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Author notes

  1. D. Zenisek and J. A. Steyer: These authors contributed equally to this work

Authors and Affiliations

  1. Vollum Institute, Oregon Health Sciences University, 3181 SW Sam Jackson Park Road, Portland, 97201, Oregon, USA

    D. Zenisek & W. Almers

  2. Department of Molecular and Cell Biology, University of California, Berkeley, 94720, California , USA

    J. A. Steyer

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  3. W. Almers
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Zenisek, D., Steyer, J. & Almers, W. Transport, capture and exocytosis of single synaptic vesicles at active zones. Nature 406, 849–854 (2000). https://doi.org/10.1038/35022500

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