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Synaptotagmin I is necessary for compensatory synaptic vesicle endocytosis in vivo

Nature volume 426, pages 559–563 (2003)Cite this article

Abstract

Neurotransmission requires a balance of synaptic vesicle exocytosis and endocytosis1. Synaptotagmin I (Syt I) is widely regarded as the primary calcium sensor for synaptic vesicle exocytosis2,3,4,5,6. Previous biochemical data suggest that Syt I may also function during synaptic vesicle endocytosis7,8,9,10,11,12,13,14,15,16; however, ultrastructural analyses at synapses with impaired Syt I function have provided an indirect and conflicting view of the role of Syt I during synaptic vesicle endocytosis3,8,9,10,14. Until now it has not been possible experimentally to separate the exocytic and endocytic functions of Syt I in vivo. Here, we test directly the role of Syt I during endocytosis in vivo. We use quantitative live imaging of a pH-sensitive green fluorescent protein fused to a synaptic vesicle protein (synapto-pHluorin) to measure the kinetics of endocytosis in sytI-null Drosophila. We then combine live imaging of the synapto-pHluorins with photoinactivation of Syt I, through fluorescein-assisted light inactivation, after normal Syt I-mediated vesicle exocytosis. By inactivating Syt I only during endocytosis, we demonstrate that Syt I is necessary for the endocytosis of synaptic vesicles that have undergone exocytosis using a functional Syt I protein.

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Figure 1: Endocytosis is impaired at sytI-null synapses.
Figure 2: Photoinactivation of Syt I4C impairs endocytosis after normal vesicle fusion.
Figure 3: n-Syb-pH remains on the synaptic plasma membrane after FlAsH-FALI of Syt I4C.
Figure 4: Photoinactivation of Syt I4C impairs stimulus-dependent FM4-64 loading at the NMJ.

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Acknowledgements

We thank M. Ramaswami and I. Robinson for Drosophila stocks, and D. DeAngelis, J. Rothman and R. Kelly for the superecliptic pHluorin GFP construct. We also thank E. Heckscher for comments on an earlier version of the manuscript.

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  1. Department of Biochemistry and Biophysics, Program in Neuroscience, University of California, San Francisco, 513 Parnassus Avenue, San Francisco, California, 94143, USA

    Kira E. Poskanzer, Kurt W. Marek, Sean T. Sweeney & Graeme W. Davis

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  1. Kira E. Poskanzer
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Correspondence to Graeme W. Davis.

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Poskanzer, K., Marek, K., Sweeney, S. et al. Synaptotagmin I is necessary for compensatory synaptic vesicle endocytosis in vivo. Nature 426, 559–563 (2003). https://doi.org/10.1038/nature02184

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