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Three modes of synaptic vesicular recycling revealed by single-vesicle imaging

Abstract

Synapses recycle their spent vesicles in order to keep up with on-going neurotransmitter release. To investigate vesicle recycling in the small synapses of hippocampal neurons, we have used an optical recording method that permits us to resolve single-vesicle events. Here we show that an exocytic event can terminate with three modes of vesicle retrieval: a fast (400–860 ms) ‘kiss-and-run’ mode that has a selective fusion pore; a slow (8–21 s) ‘compensatory’ mode; and a ‘stranded’ mode of recycling, in which a vesicle is left on the cell surface until a nerve impulse triggers its retrieval. We have also observed that, in response to a nerve impulse, synapses with low release probability primarily use the kiss-and-run mode, whereas high release probability terminals predominantly use the compensatory mode of vesicle retrieval.

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Figure 1: Imaging synaptic vesicle recycling by sensing vesicular pH.
Figure 2: Detection of single-vesicle release.
Figure 3: Evoked single-vesicle retrieval.
Figure 4: Kiss-and-run, compensatory and stranded events.
Figure 5: Reduced access of pH buffers to kiss-and-run vesicles.
Figure 6: Balance of recycling modes depends on release probability.

References

  1. Ryan, T. A. & Smith, S. J. Vesicle pool mobilization during action potential firing at hippocampal synapses. Neuron 14, 983–989 (1995)

    CAS  Article  Google Scholar 

  2. Harata, N. et al. Limited numbers of recycling vesicles in small CNS nerve terminals: implications for neural signaling and vesicular cycling. Trends Neurosci. 24, 637–643 (2001)

    CAS  Article  Google Scholar 

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

    CAS  Article  Google Scholar 

  4. von Gersdorff, H. & Matthews, G. Dynamics of synaptic vesicle fusion and membrane retrieval in synaptic terminals. Nature 367, 735–739 (1994)

    ADS  CAS  Article  Google Scholar 

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

    ADS  CAS  Article  Google Scholar 

  6. Wu, L. G. & Betz, W. J. Nerve activity but not intracellular calcium determines the time course of endocytosis at the frog neuromuscular junction. Neuron 17, 769–779 (1996)

    CAS  Article  Google Scholar 

  7. Ryan, T. A., Smith, S. J. & Reuter, H. The timing of synaptic vesicle endocytosis. Proc. Natl Acad. Sci. USA 93, 5567–5571 (1996)

    ADS  CAS  Article  Google Scholar 

  8. Klingauf, J., Kavalali, E. T. & Tsien, R. W. Kinetics and regulation of fast endocytosis at hippocampal synapses. Nature 394, 581–585 (1998)

    ADS  CAS  Article  Google Scholar 

  9. Pyle, J. L., Kavalali, E. T., Piedras-Renteria, E. S. & Tsien, R. W. Rapid reuse of readily releasable pool vesicles at hippocampal synapses. Neuron 28, 221–231 (2000)

    CAS  Article  Google Scholar 

  10. Stevens, C. F. & Williams, J. H. ‘Kiss and run’ exocytosis at hippocampal synapses. Proc. Natl Acad. Sci. USA 97, 12828–12833 (2000)

    ADS  CAS  Article  Google Scholar 

  11. Sankaranarayanan, S. & Ryan, T. A. Real-time measurements of vesicle-SNARE recycling in synapses of the central nervous system. Nature Cell Biol. 2, 197–204 (2000)

    CAS  Article  Google Scholar 

  12. Neves, G., Gomis, A. & Lagnado, L. Calcium influx selects the fast mode of endocytosis in the synaptic terminal of retinal bipolar cells. Proc. Natl Acad. Sci. USA 98, 15282–15287 (2001)

    ADS  CAS  Article  Google Scholar 

  13. Sankaranarayanan, S. & Ryan, T. A. Calcium accelerates endocytosis of vSNAREs at hippocampal synapses. Nature Neurosci. 4, 129–136 (2001)

    CAS  Article  Google Scholar 

  14. Heidelberger, R. ATP is required at an early step in compensatory endocytosis in synaptic terminals. J. Neurosci. 21, 6467–6474 (2001)

    CAS  Article  Google Scholar 

  15. Sun, J. Y., Wu, X. S. & Wu, L. G. Single and multiple vesicle fusion induce different rates of endocytosis at a central synapse. Nature 417, 555–559 (2002)

    ADS  CAS  Article  Google Scholar 

  16. Zenisek, D., Steyer, J. A., Feldman, M. E. & Almers, W. A membrane marker leaves synaptic vesicles in milliseconds after exocytosis in retinal bipolar cells. Neuron 35, 1085–1097 (2002)

    CAS  Article  Google Scholar 

  17. Heidelberger, R., Zhou, Z. Y. & Matthews, G. Multiple components of membrane retrieval in synaptic terminals revealed by changes in hydrostatic pressure. J. Neurophysiol. 88, 2509–2517 (2002)

    Article  Google Scholar 

  18. Miesenbock, G., De Angelis, D. A. & Rothman, J. E. Visualizing secretion and synaptic transmission with pH-sensitive green fluorescent proteins. Nature 394, 192–195 (1998)

    ADS  CAS  Article  Google Scholar 

  19. Murthy, V. N., Sejnowski, T. J. & Stevens, C. F. Heterogeneous release properties of visualized individual hippocampal synapses. Neuron 18, 599–612 (1997)

    CAS  Article  Google Scholar 

  20. Huang, E. P. & Stevens, C. F. Estimating the distribution of synaptic reliabilities. J. Neurophysiol. 78, 2870–2880 (1997)

    CAS  Article  Google Scholar 

  21. Goda, Y. & Stevens, C. F. Two components of transmitter release at a central synapse. Proc. Natl Acad. Sci. USA 91, 12942–12946 (1994)

    ADS  CAS  Article  Google Scholar 

  22. Smith, C. & Neher, E. Multiple forms of endocytosis in bovine adrenal chromaffin cells. J. Cell Biol. 139, 885–894 (1997)

    CAS  Article  Google Scholar 

  23. Taraska, J. W., Perrais, D., Ohara-Imaizumi, M., Nagamatsu, S. & Almers, W. Secretory granules are recaptured largely intact after stimulated exocytosis in cultured endocrine cells. Proc. Natl Acad. Sci. USA 100, 2070–2075 (2003)

    ADS  CAS  Article  Google Scholar 

  24. Beutner, D., Voets, T., Neher, E. & Moser, T. Calcium dependence of exocytosis and endocytosis at the cochlear inner hair cell afferent synapse. Neuron 29, 681–690 (2001)

    CAS  Article  Google Scholar 

  25. Engisch, K. L. & Nowycky, M. C. Compensatory and excess retrieval: two types of endocytosis following single step depolarizations in bovine adrenal chromaffin cells. J. Physiol. 506, 591–608 (1998)

    CAS  Article  Google Scholar 

  26. Hsu, S. F. & Jackson, M. B. Rapid exocytosis and endocytosis in nerve terminals of the rat posterior pituitary. J. Physiol. 494, 539–553 (1996)

    CAS  Article  Google Scholar 

  27. Neale, E. A., Bowers, L. M., Jia, M., Bateman, K. E. & Williamson, L. C. Botulinum neurotoxin A blocks synaptic vesicle exocytosis but not endocytosis at the nerve terminal. J. Cell Biol. 147, 1249–1260 (1999)

    CAS  Article  Google Scholar 

  28. Thomas, P., Lee, A. K., Wong, J. G. & Almers, W. A triggered mechanism retrieves membrane in seconds after Ca(2 + )-stimulated exocytosis in single pituitary cells. J. Cell Biol. 124, 667–675 (1994)

    CAS  Article  Google Scholar 

  29. Albillos, A. et al. The exocytotic event in chromaffin cells revealed by patch amperometry. Nature 389, 509–512 (1997)

    ADS  CAS  Article  Google Scholar 

  30. Klyachko, V. A. & Jackson, M. B. Capacitance steps and fusion pores of small and large-dense-core vesicles in nerve terminals. Nature 418, 89–92 (2002)

    ADS  CAS  Article  Google Scholar 

  31. Ales, E. et al. High calcium concentrations shift the mode of exocytosis to the kiss- and-run mechanism. Nature Cell Biol. 1, 40–44 (1999)

    CAS  Article  Google Scholar 

  32. Cousin, M. A. & Robinson, P. J. Two mechanisms of synaptic vesicle recycling in rat brain nerve terminals. J. Neurochem. 75, 1645–1653 (2000)

    CAS  Article  Google Scholar 

  33. Murthy, V. N. & Stevens, C. F. Reversal of synaptic vesicle docking at central synapses. Nature Neurosci. 2, 503–507 (1999)

    CAS  Article  Google Scholar 

  34. Sankaranarayanan, S., De Angelis, D., Rothman, J. E. & Ryan, T. A. The use of pHluorins for optical measurements of presynaptic activity. Biophys. J. 79, 2199–2208 (2000)

    CAS  Article  Google Scholar 

  35. Wu, M. M. et al. Mechanisms of pH regulation in the regulated secretory pathway. J. Biol. Chem. 276, 33027–33035 (2001)

    CAS  Article  Google Scholar 

  36. Wu, M. M. et al. Organelle pH studies using targeted avidin and fluorescein-biotin. Chem. Biol. 7, 197–209 (2000)

    CAS  Article  Google Scholar 

  37. Nishi, T. & Forgac, M. The vacuolar (H + )-ATPases—nature's most versatile proton pumps. Nature Rev. Mol. Cell Biol. 3, 94–103 (2002)

    CAS  Article  Google Scholar 

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Acknowledgements

We thank R. Jacobs, M. Pilla and J. Sullivan for technical support; D. De Angelis for the super-ecliptic phluorin construct; Roger Tsien for suggesting the use of photobleaching; and N. Spitzer for a critical reading of this manuscript.

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Correspondence to Charles F. Stevens.

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Gandhi, S., Stevens, C. Three modes of synaptic vesicular recycling revealed by single-vesicle imaging. Nature 423, 607–613 (2003). https://doi.org/10.1038/nature01677

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