Journal of Comparative Physiology A

, Volume 191, Issue 12, pp 1157–1171 | Cite as

A dye mixture (Neurobiotin and Alexa 488) reveals extensive dye-coupling among neurons in leeches; physiology confirms the connections

  • Ruey-Jane Fan
  • Antonia Marin-Burgin
  • Kathleen A. French
  • W. Otto Friesen
Original Paper
First Online:
Received:
Revised:
Accepted:

Abstract

Although the neuronal circuits that generate leech movements have been studied for over 30 years, the list of interneurons (INs) in these circuits remains incomplete. Previous studies showed that some motor neurons (MNs) are electrically coupled to swim-related INs, e.g., rectifying junctions connect IN 28 to MN DI-1 (dorsal inhibitor), so we searched for additional neurons in these behavioral circuits by co-injecting Neurobiotin and Alexa Fluor 488 into segmental MNs DI–1, VI–2, DE–3 and VE–4. The high molecular weight Alexa dye is confined to the injected cell, whereas the smaller Neurobiotin molecules diffuse through gap junctions to reveal electrical coupling. We found that MNs were each dye-coupled to approximately 25 neurons, about half of which are likely to be INs. We also found that (1) dye-coupling was reliably correlated with physiologically confirmed electrical connections, (2) dye-coupling is unidirectional between MNs that are linked by rectifying connections, and (3) there are novel electrical connections between excitatory and inhibitory MNs, e.g. between excitatory MN VE-4 and inhibitory MN DI-1. The INs found in this study provide a pool of novel candidate neurons for future studies of behavioral circuits, including those underlying swimming, crawling, shortening, and bending movements.

Keywords

Hirudo Circuits Gap junctions Diode Motoneurons 
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Notes

Acknowledgements

We thank our colleagues, Drs. DeForest Mellon and Masashi Kawasaki for helpful discussions. We also thank Dr. Periasamy and Ms. Chen for technical assistance with the confocal microscopy and Courtney Milianes for expert editorial assistance. This research was supported by the National Science Foundation (Grant IBN-0110607 to W.O.F.) and a Fundacion Antorchas grant to AMB. Experiments comply with publication No. 86-23, revised 1985, of the National Institute of Health and with US laws.

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Copyright information

© Springer-Verlag 2005

Authors and Affiliations

  • Ruey-Jane Fan
    • 1
  • Antonia Marin-Burgin
    • 2
  • Kathleen A. French
    • 2
  • W. Otto Friesen
    • 1
  1. 1.Department of BiologyUniversity of VirginiaCharlottesvilleUSA
  2. 2.Division of Biological Sciences, Neurobiology SectionUniversity of CaliforniaSan Diego La JollaUSA

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