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Vol. 12, No. 1, 2003
Issue release date: January–February 2003
Free Access
Neurosignals 2003;12:13–30
(DOI:10.1159/000068912)

Roles of SNARE Proteins and Synaptotagmin I in Synaptic Transmission: Studies at the Drosophila Neuromuscular Synapse

Kidokoro Y.
Institute for Behavioral Sciences, Gunma University School of Medicine, Maebashi, Japan
email Corresponding Author

Abstract

The roles of SNARE proteins, i.e. neuronal Synaptobrevin (n-Syb), SNAP-25 and Syntaxin 1A (Syx 1A), and Synaptotagmin I (Syt I) in synaptic transmission have been studied in situ using mutant embryos or larvae that lack these molecules or have alterations in them. Because of the ease of genetic manipulation, the Drosophila neuromuscular synapse is widely used for these studies. The functional properties of synaptic transmission have been studied in mutant embryos using the patch-clamp technique, and in larvae by recording with microelectrodes. A major vesicular membrane protein, n-Syb, is indispensable for nerve-evoked synaptic transmission. Spontaneous synaptic currents (minis), however, are present even in embryos totally lacking n-Syb (n-syb). Furthermore, Ca2+-independent enhancement of mini frequency induced by hypertonic sucrose solutions (hypertonicity response) is totally absent in n-syb. Embryos that have defects in SNAP-25 (SNAP-25) have similar but milder phenotypes than n-syb. The phenotype in synaptic transmission was most severe in the synapse lacking Syx 1A. Neither nerve-evoked synaptic currents nor minis occur in embryos lacking Syx 1A (syx 1A). No hypertonicity response was observed in them. Syt I binds Ca2+ in vitro and probably serves as a Ca2+ sensor for nerve-evoked synaptic transmission, since nerve-evoked synaptic currents were greatly reduced in embryos lacking Syt I (syt I). Also, Syt I has a role in vesicle recycling. Interestingly, the Ca2+-independent hypertonicity response is also greatly reduced in syt I. Minis persist in mutant embryos lacking any of these proteins (n-Syb, SNAP-25 and Syt I), except Syx, suggesting that minis have a distinct fusion mechanism from that for fast and synchronized release. It appears that these SNARE proteins and Syt I are coordinated for fast vesicle fusion. Minis, on the other hand, do not require SNARE complex nor Syt I, but Syx is absolutely required for vesicle fusion. The SNARE complex and Syt I are indispensable for the hypertonicity response. None of these molecules seem to serve for selective docking of synaptic vesicles to the release site. For further studies on synaptic transmission, the Drosophila neuromuscular synapse will continue to be a useful model.


 goto top of outline Key Words

  • Synaptic transmission
  • SNARE complex
  • Synaptobrevin
  • Syntaxin
  • SNAP-25
  • Synaptotagmin

 goto top of outline Abstract

The roles of SNARE proteins, i.e. neuronal Synaptobrevin (n-Syb), SNAP-25 and Syntaxin 1A (Syx 1A), and Synaptotagmin I (Syt I) in synaptic transmission have been studied in situ using mutant embryos or larvae that lack these molecules or have alterations in them. Because of the ease of genetic manipulation, the Drosophila neuromuscular synapse is widely used for these studies. The functional properties of synaptic transmission have been studied in mutant embryos using the patch-clamp technique, and in larvae by recording with microelectrodes. A major vesicular membrane protein, n-Syb, is indispensable for nerve-evoked synaptic transmission. Spontaneous synaptic currents (minis), however, are present even in embryos totally lacking n-Syb (n-syb). Furthermore, Ca2+-independent enhancement of mini frequency induced by hypertonic sucrose solutions (hypertonicity response) is totally absent in n-syb. Embryos that have defects in SNAP-25 (SNAP-25) have similar but milder phenotypes than n-syb. The phenotype in synaptic transmission was most severe in the synapse lacking Syx 1A. Neither nerve-evoked synaptic currents nor minis occur in embryos lacking Syx 1A (syx 1A). No hypertonicity response was observed in them. Syt I binds Ca2+ in vitro and probably serves as a Ca2+ sensor for nerve-evoked synaptic transmission, since nerve-evoked synaptic currents were greatly reduced in embryos lacking Syt I (syt I). Also, Syt I has a role in vesicle recycling. Interestingly, the Ca2+-independent hypertonicity response is also greatly reduced in syt I. Minis persist in mutant embryos lacking any of these proteins (n-Syb, SNAP-25 and Syt I), except Syx, suggesting that minis have a distinct fusion mechanism from that for fast and synchronized release. It appears that these SNARE proteins and Syt I are coordinated for fast vesicle fusion. Minis, on the other hand, do not require SNARE complex nor Syt I, but Syx is absolutely required for vesicle fusion. The SNARE complex and Syt I are indispensable for the hypertonicity response. None of these molecules seem to serve for selective docking of synaptic vesicles to the release site. For further studies on synaptic transmission, the Drosophila neuromuscular synapse will continue to be a useful model.

Copyright © 2003 S. Karger AG, Basel


 goto top of outline References
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  3. Südhof TC: The synaptic vesicle cycle: A cascade of protein-protein interactions. Nature 1995;375:645–653.
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  7. Broadie K, Bate M: Activity-dependent development of the neuromuscular synapse during Drosophila embryogenesis. Neuron 1993;11:607–619.
  8. Kidokoro Y, Nishikawa K: Miniature endplate currents at the newly formed neuromuscular junction in Drosophila embryos and larvae. Neurosci Res 1994;19:143–154.
  9. Nishikawa K, Kidokoro Y: Junctional and extrajunctional glutamate receptor channels in Drosophila embryos and larvae. J Neurosci 1996;15:7905–7915.
  10. Simon SM, Llinás RR: Compartmentalization of the submembrane calcium activity during calcium influx and its significance in transmitter release. Biophys J 1985;48:485–498.
  11. Adler EM, Augustine GJ, Duffy SN, Charlton MP: Alien intracellular calcium chelators attenuate neurotransmitter release at the squid synapse. J Neurosci 1991;11:1496–1507.
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 goto top of outline Author Contacts

Yoshi Kidokoro
Institute for Behavioral Sciences, Gunma University School of Medicine
3-39-22 Showa-machi
Maebashi 371-8511 (Japan)
Tel. +81 27 220 8040, Fax +81 27 220 8046, E-Mail kidokoro@med.gunma-u.ac.jp


 goto top of outline Article Information

Received: August 27, 2002
Accepted after revision: October 16, 2002
Number of Print Pages : 18
Number of Figures : 7, Number of Tables : 0, Number of References : 83


 goto top of outline Publication Details

Neurosignals
Founded 1992 as ‘Biological Signals’ by S.F. Pang (1992–2001) continued as ‘Biological Signals and Receptors’ (1999–2001)

Vol. 12, No. 1, Year 2003 (Cover Date: January-February 2003)

Journal Editor: Nancy Y. Ip, Hong Kong
ISSN: 1424–862X (print), 1424–8638 (Online)

For additional information: http://www.karger.com/journals/nsg


Copyright / Drug Dosage / Disclaimer

Copyright: All rights reserved. No part of this publication may be translated into other languages, reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying, recording, microcopying, or by any information storage and retrieval system, without permission in writing from the publisher or, in the case of photocopying, direct payment of a specified fee to the Copyright Clearance Center.
Drug Dosage: The authors and the publisher have exerted every effort to ensure that drug selection and dosage set forth in this text are in accord with current recommendations and practice at the time of publication. However, in view of ongoing research, changes in goverment regulations, and the constant flow of information relating to drug therapy and drug reactions, the reader is urged to check the package insert for each drug for any changes in indications and dosage and for added warnings and precautions. This is particularly important when the recommended agent is a new and/or infrequently employed drug.
Disclaimer: The statements, opinions and data contained in this publication are solely those of the individual authors and contributors and not of the publishers and the editor(s). The appearance of advertisements or/and product references in the publication is not a warranty, endorsement, or approval of the products or services advertised or of their effectiveness, quality or safety. The publisher and the editor(s) disclaim responsibility for any injury to persons or property resulting from any ideas, methods, instructions or products referred to in the content or advertisements.

Abstract

The roles of SNARE proteins, i.e. neuronal Synaptobrevin (n-Syb), SNAP-25 and Syntaxin 1A (Syx 1A), and Synaptotagmin I (Syt I) in synaptic transmission have been studied in situ using mutant embryos or larvae that lack these molecules or have alterations in them. Because of the ease of genetic manipulation, the Drosophila neuromuscular synapse is widely used for these studies. The functional properties of synaptic transmission have been studied in mutant embryos using the patch-clamp technique, and in larvae by recording with microelectrodes. A major vesicular membrane protein, n-Syb, is indispensable for nerve-evoked synaptic transmission. Spontaneous synaptic currents (minis), however, are present even in embryos totally lacking n-Syb (n-syb). Furthermore, Ca2+-independent enhancement of mini frequency induced by hypertonic sucrose solutions (hypertonicity response) is totally absent in n-syb. Embryos that have defects in SNAP-25 (SNAP-25) have similar but milder phenotypes than n-syb. The phenotype in synaptic transmission was most severe in the synapse lacking Syx 1A. Neither nerve-evoked synaptic currents nor minis occur in embryos lacking Syx 1A (syx 1A). No hypertonicity response was observed in them. Syt I binds Ca2+ in vitro and probably serves as a Ca2+ sensor for nerve-evoked synaptic transmission, since nerve-evoked synaptic currents were greatly reduced in embryos lacking Syt I (syt I). Also, Syt I has a role in vesicle recycling. Interestingly, the Ca2+-independent hypertonicity response is also greatly reduced in syt I. Minis persist in mutant embryos lacking any of these proteins (n-Syb, SNAP-25 and Syt I), except Syx, suggesting that minis have a distinct fusion mechanism from that for fast and synchronized release. It appears that these SNARE proteins and Syt I are coordinated for fast vesicle fusion. Minis, on the other hand, do not require SNARE complex nor Syt I, but Syx is absolutely required for vesicle fusion. The SNARE complex and Syt I are indispensable for the hypertonicity response. None of these molecules seem to serve for selective docking of synaptic vesicles to the release site. For further studies on synaptic transmission, the Drosophila neuromuscular synapse will continue to be a useful model.



 goto top of outline Author Contacts

Yoshi Kidokoro
Institute for Behavioral Sciences, Gunma University School of Medicine
3-39-22 Showa-machi
Maebashi 371-8511 (Japan)
Tel. +81 27 220 8040, Fax +81 27 220 8046, E-Mail kidokoro@med.gunma-u.ac.jp


 goto top of outline Article Information

Received: August 27, 2002
Accepted after revision: October 16, 2002
Number of Print Pages : 18
Number of Figures : 7, Number of Tables : 0, Number of References : 83


 goto top of outline Publication Details

Neurosignals
Founded 1992 as ‘Biological Signals’ by S.F. Pang (1992–2001) continued as ‘Biological Signals and Receptors’ (1999–2001)

Vol. 12, No. 1, Year 2003 (Cover Date: January-February 2003)

Journal Editor: Nancy Y. Ip, Hong Kong
ISSN: 1424–862X (print), 1424–8638 (Online)

For additional information: http://www.karger.com/journals/nsg


Copyright / Drug Dosage

Copyright: All rights reserved. No part of this publication may be translated into other languages, reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying, recording, microcopying, or by any information storage and retrieval system, without permission in writing from the publisher or, in the case of photocopying, direct payment of a specified fee to the Copyright Clearance Center.
Drug Dosage: The authors and the publisher have exerted every effort to ensure that drug selection and dosage set forth in this text are in accord with current recommendations and practice at the time of publication. However, in view of ongoing research, changes in goverment regulations, and the constant flow of information relating to drug therapy and drug reactions, the reader is urged to check the package insert for each drug for any changes in indications and dosage and for added warnings and precautions. This is particularly important when the recommended agent is a new and/or infrequently employed drug.
Disclaimer: The statements, opinions and data contained in this publication are solely those of the individual authors and contributors and not of the publishers and the editor(s). The appearance of advertisements or/and product references in the publication is not a warranty, endorsement, or approval of the products or services advertised or of their effectiveness, quality or safety. The publisher and the editor(s) disclaim responsibility for any injury to persons or property resulting from any ideas, methods, instructions or products referred to in the content or advertisements.

References

  1. Shi S-H, Hayashi Y, Esteban JA, Malinow R: Subunit-specific rules governing AMPA receptor trafficking to synapses in hippocampal pyramidal neurons. Cell 2001;105:331–343.
  2. Turrigiano GG: AMPA receptors unbound: Membrane cycling and synaptic plasticity. Neuron 2000;26:5–8.
  3. Südhof TC: The synaptic vesicle cycle: A cascade of protein-protein interactions. Nature 1995;375:645–653.
  4. Sutton RB, Fassbauer D, Jahn R, Brunger AT: Crystal structure of a SNARE complex involved in synaptic exocytosis at 2.4 Å resolution. Nature 1998;395:347–353.
  5. Broadie K, Bate M: Innervation directs receptor synthesis and localization in Drosophila embryo synaptogenesis. Nature 1993;361:350–353.
  6. Broadie K, Bate M: Development of the embryonic neuromuscular synapse of Drosophila melanogaster. J Neurosci 1993;13:144–166.
  7. Broadie K, Bate M: Activity-dependent development of the neuromuscular synapse during Drosophila embryogenesis. Neuron 1993;11:607–619.
  8. Kidokoro Y, Nishikawa K: Miniature endplate currents at the newly formed neuromuscular junction in Drosophila embryos and larvae. Neurosci Res 1994;19:143–154.
  9. Nishikawa K, Kidokoro Y: Junctional and extrajunctional glutamate receptor channels in Drosophila embryos and larvae. J Neurosci 1996;15:7905–7915.
  10. Simon SM, Llinás RR: Compartmentalization of the submembrane calcium activity during calcium influx and its significance in transmitter release. Biophys J 1985;48:485–498.
  11. Adler EM, Augustine GJ, Duffy SN, Charlton MP: Alien intracellular calcium chelators attenuate neurotransmitter release at the squid synapse. J Neurosci 1991;11:1496–1507.
  12. Roberts WM, Jacobs RA, Hudspeth AJ: Colocalization of ion channels involved in frequency selectivity and synaptic transmission at presynaptic active zones of hair cells. J Neurosci 1990;10:3664–3684.
  13. Miledi R, Thies R: Tetanic and post-tetanic rise in frequency of miniature endplate potentials in low-calcium solutions. J Physiol 1971;212:246–257.
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