Nature 440, 935-939 (13 April 2006) | doi:10.1038/nature04592; Received 18 November 2005; Accepted 20 January 2006

STED microscopy reveals that synaptotagmin remains clustered after synaptic vesicle exocytosis

Katrin I. Willig1,3, Silvio O. Rizzoli2,3, Volker Westphal1, Reinhard Jahn2 and Stefan W. Hell1

Synaptic transmission is mediated by neurotransmitters that are stored in synaptic vesicles and released by exocytosis upon activation. The vesicle membrane is then retrieved by endocytosis, and synaptic vesicles are regenerated and re-filled with neurotransmitter1. Although many aspects of vesicle recycling are understood, the fate of the vesicles after fusion is still unclear. Do their components diffuse on the plasma membrane, or do they remain together? This question has been difficult to answer because synaptic vesicles are too small (approx40 nm in diameter) and too densely packed to be resolved by available fluorescence microscopes. Here we use stimulated emission depletion (STED)2 to reduce the focal spot area by about an order of magnitude below the diffraction limit, thereby resolving individual vesicles in the synapse. We show that synaptotagmin I, a protein resident in the vesicle membrane, remains clustered in isolated patches on the presynaptic membrane regardless of whether the nerve terminals are mildly active or intensely stimulated. This suggests that at least some vesicle constituents remain together during recycling. Our study also demonstrates that questions involving cellular structures with dimensions of a few tens of nanometres can be resolved with conventional far-field optics and visible light.

  1. Departments of NanoBiophotonics and
  2. Neurobiology, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany
  3. *These authors contributed equally to this work

Correspondence to: Reinhard Jahn2 Correspondence and requests for materials should be addressed to R.J. (Email: rjahn@gwdg.de).

Received 18 November 2005 | Accepted 20 January 2006


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