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Analysis of synaptic transmission in Caenorhabditis elegans using an aldicarb-sensitivity assay

Nature Protocols volume 1, pages 1772–1777 (2006)Cite this article

Abstract

Caenorhabditis elegans has emerged as a powerful model system for studying the biology of the synapse. Here we describe a widely used assay for synaptic transmission at the C. elegans neuromuscular junction. This protocol monitors the sensitivity of C. elegans to the paralyzing affects of an acetylcholinesterase inhibitor, aldicarb. Briefly, adult worms are incubated in the presence of aldicarb and scored for the time-course of aldicarb-induced paralysis. Animals harboring mutations in genes that affect synaptic transmission generally exhibit a change in their sensitivity to aldicarb (either increased sensitivity for enhancements in synaptic transmission or decreased sensitivity for blockage in synaptic transmission). This technique provides a simple assay for the accurate comparative analysis of synaptic transmission in multiple C. elegans strains. The protocol described can be performed relatively quickly and is a practical alternative to other techniques used to study synaptic transmission. This protocol can also be modified to follow the paralytic effects with other pharmacological reagents. The assay can be performed in about 3-6 hours depending on the severity of synaptic transmission defects.

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Figure 1: Schematic diagram demonstrating the molecular mechanisms that underlie aldicarb-induced paralysis in the nematode C. elegans.
Figure 2: Results from aldicarb assays on aldicarb-resistant and aldicarb-hypersensitive mutants.

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Author information

Author notes

  1. Timothy R Mahoney and Shuo Luo: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Anatomy and Neurobiology, Washington University School of Medicine, St. Louis, 63110, Missouri, USA

    Timothy R Mahoney, Shuo Luo & Michael L Nonet

Authors
  1. Timothy R Mahoney
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  2. Shuo Luo
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Corresponding author

Correspondence to Michael L Nonet.

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Supplementary information

Supplementary Video 1

Wild-type C. elegans on 1 mM Aldicarb. Video was captured (approximately 7 frames per second) using a Retiga Exi camera controlled by OpenLab Software on a Zeiss stereomicroscope with a trans-illuminated stage. Paralyzed worms were prodded with a platinum wire as described in the protocol. Speed of movie is about 1× (or 7 frames per second). Time = 0 min. A wild-type worm moving at normal velocity. (MOV 417 kb)

Supplementary Video 2

Wild-type C. elegans on 1 mM Aldicarb. Time = 30 min. A wild-type worm moving sluggishly. Note the weak coiling phenotype towards the end of the movie, commonly seen in synaptic transmission mutants. Also, note the rigidity of the movements and the awkward twists of the body. (MOV 750 kb)

Supplementary Video 3

Wild-type C. elegans on 1 mM Aldicarb. Time = 90 min. A wild-type worm that is partially paralyzed. Note the animal appears hyper-contracted and is partially paralyzed. However, the head is still moving/foraging for food. Also, note the first gentle touch near the head of the animal elicits movement in the tail. Based on the observation of movements of the head and the ability to stimulate movement by tapping gently with a platinum wire, this animal is not yet considered paralyzed. (MOV 1007 kb)

Supplementary Video 4

Wild-type C. elegans on 1mM Aldicarb. Time = 90 min. A wild-type worm that is partially paralyzed. (MOV 522 kb)

Supplementary Video 5

Wild-type C. elegans on 1 mM Aldicarb. Time = 120 min. A wild-type worm that is almost completely paralyzed. Note the animal is hyper-contracted and virtually paralyzed. It is essentially unresponsive to touch, however the pharynx is still pumping and the tip of the head is showing some foraging movements. It is not uncommon to see a worm become more paralyzed after prodding with a platinum wire or to see animals release eggs as they become paralyzed. This video illustrates the subjective nature of defining an endpoint. It is likely that some experimenters would score this animal as paralyzed. However, if there is concern over consistency in scoring paralysis or if the experimenter is testing severely paralyzed mutants we suggest waiting for an even more paralyzed animal, such as seen in Supplementary Video 6. (MOV 789 kb)

Supplementary Video 6

Wild-type C. elegans on 1mM Aldicarb. Time = 120 min. A wild-type worm that is completely paralyzed. This animal is similarly hyper-contracted to the animal in Supplementary Video 5. Unlike the example in Supplementary Video 5, this animal is completely paralyzed and completely unresponsive to touch. Throughout the entire video the pharynx pumps only twice. This animal is a good example of a completely paralyzed animal. (MOV 949 kb)

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Mahoney, T., Luo, S. & Nonet, M. Analysis of synaptic transmission in Caenorhabditis elegans using an aldicarb-sensitivity assay. Nat Protoc 1, 1772–1777 (2006). https://doi.org/10.1038/nprot.2006.281

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