The millimeter-long nematode Caenorhabditis elegans offers several unique advantages over other model organisms, including 60% homology with human genes, a short life cycle, 95 body wall muscle cells that are similar to vertebrate muscles, and 302 mapped neurons1-3. Its short lifespan and high genetic conservation enable the identification of genes and mechanisms regulating neuromuscular health and decline with age4, impacting a broad range of human diseases from muscular dystrophies5 to neurodegenerative disorders6-8.

However, assay culture conditions typically do not reflect the C. elegans natural habitat, in which they burrow in three dimensions (i.e., in soil, rotten fruit, and fluid drops)16-20, and so it is important to develop alternative assays that can better capture this environment7. Previous attempts have used agar plates or thrashing assays that score animal swimming locomotion8,9,10.

Although these methods have revealed some aspects of C. elegans neurological control of locomotion, many aspects remain to be explored and these include the role of non-muscle cells in generating 3D movement11-12, the ability of animals to maneuver and change direction in this environment16-17, and the contribution of chemotaxis in burrowing prowess8,9,10. We have recently developed a new method for evaluation of C. elegans burrowing performance that can address these limitations, and we describe it here in detail for whole-organism assessment16-20.

A Novel 3D Gel Based Burrowing Assay

We have adapted an existing agar-based nematode burrowing assay to use Pluronic F-127, a biocompatible thermoreversible hydrogel. Its optical transparency23-27 and favorable temperature range24,25 make it an excellent choice as the medium for a gel-based burrowing assay that does not cause harm to nematodes16,18,20-21.

We show that this gel-based burrowing assay can be easily used to assess the burrowing performance of both wild-type and mutant C. elegans, and that this assay can be adjusted to modulate the animal's burrowing performance by changing the gel height or concentration. Furthermore, we show that the gel-based assay can be used to evaluate different strains in parallel and that the method allows for easy recovery of animals for follow-up analysis. These findings expand our application of the gel-based burrowing assay to a variety of areas, enabling sensitive dynamic range assessments that have previously not been possible.

Author:

Kerry C.

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