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Published March 5, 2018 | Published + Accepted Version
Journal Article Open

Dual-sided Voltage-sensitive Dye Imaging of Leech Ganglia


In this protocol, we introduce an effective method for voltage-sensitive dye (VSD) loading and imaging of leech ganglia as used in Tomina and Wagenaar (2017). Dissection and dye loading procedures are the most critical steps toward successful whole-ganglion VSD imaging. The former entails the removal of the sheath that covers neurons in the segmental ganglion of the leech, which is required for successful dye loading. The latter entails gently flowing a new generation VSD, VF2.1(OMe).H, onto both sides of the ganglion simultaneously using a pair of peristaltic pumps. We expect the described techniques to translate broadly to wide-field VSD imaging in other thin and relatively transparent nervous systems.

Additional Information

© 2018 Tomina and Wagenaar. This article is distributed under the terms of the Creative Commons Attribution License (CC BY 4.0). We thank Evan Miller for supplying the VF2.1(OMe).H dye; Annette Stowasser for her role in developing a prototype of the double-sided microscope and many helpful conversations; Angela Bruno for useful discussions regarding data analysis; Ng Cai Tong for reading and checking the manuscript. This work was supported by the Burroughs Welcome Fund through a Career Award at the Scientific Interface and by the National Institute of Neurological Disorders and Stroke through grant R01 NS094403 (both to DAW). YT was supported by JSPS Overseas Research Fellowships. This protocol was adapted from procedures published in Tomina and Wagenaar (2017). The authors of this work declare no conflicts of interest.

Attached Files

Published - Bio-protocol2751.pdf

Accepted Version - nihms948707.pdf


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October 18, 2023