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Front. Neuroanat. | doi: 10.3389/fnana.2018.00095

Verifying, Challenging, and Discovering New Synapses among Fully EM-Reconstructed Neurons in the Leech Ganglion

  • 1University of California, San Diego, United States
  • 2Division of Biological Sciences, University of California, San Diego, United States
  • 3National Center for Microscopy and Imaging Research, University of California at San Diego, United States
  • 4Department of Neuroscience, University of California, San Diego, United States

Neural circuits underpin the production of animal behavior, largely based upon the precise pattern of synaptic connectivity among the neurons involved. For large numbers of neurons, determining such “connectomes” by direct physiological means is difficult, as physiological accessibility is ultimately required to verify and characterize the function of synapses. We collected a volume of images spanning an entire ganglion of the juvenile leech nervous system via serial blockface electron microscopy (SBEM). We validated this approach by reconstructing a well-characterized circuit of motor neurons involved in the swimming behavior of the leech by locating the synapses among them. We confirm that there are multiple synaptic contacts between connected pairs of neurons in the leech, and that these synapses are widely distributed across the region of neuropil in which the neurons’ arbors overlap. We verified the anatomical existence of connections that had been described physiologically among longitudinal muscle motor neurons. We also found that some physiological connections were not present anatomically. We then drew upon the SBEM dataset to design additional physiological experiments. We reconstructed an uncharacterized neuron and one of its presynaptic partners identified from the SBEM dataset. We subsequently interrogated this cell pair via intracellular electrophysiology in an adult ganglion and found that the anatomically-discovered synapse was also functional physiologically. Our findings demonstrate the value of combining a connectomics approach with electrophysiology in the leech nervous system

Keywords: serial block face scanning electron microscopy, Invertebrate neurobiology, leech, EM reconstruction, synapse

Received: 16 Jul 2018; Accepted: 18 Oct 2018.

Edited by:

Yoshiyuki Kubota, National Institute for Physiological Sciences (NIPS), Japan

Reviewed by:

Marcel Oberlaender, CAESAR Zentrum für fortgeschrittene europäische Wissenschaft und Forschung, Germany
Antonia Marin-Burgin, CONICET Instituto de Investigación en Biomedicina de Buenos Aires (IBioBA), Argentina  

Copyright: © 2018 Pipkin, Bushong, Ellisman and Kristan. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

* Correspondence: Dr. Jason E. Pipkin, University of California, San Diego, San Diego, United States, jason.e.pipkin@gmail.com