Automated Analysis of <em>C. elegans</em> Swim Behavior Using CeleST Software

Abstract

Dissecting the neuronal and neuromuscular circuits that regulate behavior remains a major challenge in biology. The nematode Caenorhabditis elegans has proven to be an invaluable model organism in helping to tackle this challenge, from inspiring technological approaches, building the human brain connectome, to actually shedding light on the specific molecular drivers of basic functional patterns. The bulk of the behavioral studies in C. elegans have been performed on solid substrates. In liquid, animals exhibit behavioral patterns that include movement at a range of speeds in 3D, as well as partial body movements, such as a posterior curl without anterior shape change, which introduce new challenges for quantitation. The steps of a simple procedure, and use of a software that enables high-resolution analysis of C. elegans swim behavior, are presented here. The software, named CeleST, uses a specialized computer program that tracks multiple animals simultaneously and provides novel measures of C. elegans locomotion in liquid (swimming). The measures are mostly grounded in animal posture and based on mathematics used in computer vision and pattern recognition, without computational requirements for threshold cut-offs. The software tool can be used to both assess overall swimming prowess in hundreds of animals from combined small batch trials and to reveal novel phenotypes even in wellcharacterized genetic mutants. The preparation of specimens for analysis with CeleST is simple and low-tech, enabling wide adaptation by the scientific community. Use of the computational approach described here should therefore contribute to the greater understanding of behavior and behavioral circuits in the C. elegans model.

Healthspan Maintenance and Prevention of Parkinson’s-like Phenotypes with Hydroxytyrosol and Oleuropein Aglycone in C. elegans

“…The study of locomotion was realized with a swim assay according to Restif et al [61] and Ibáñez-Ventoso et al [124]. Wells with a depth of 0.5 mm and Ø 10 mm were created with two self-adhesive marking films for microscope slides and filled with M9 buffer.…”

“…Each video was converted into single frames and processed with Photoshop (version 19.1.7; Adobe Inc., San José, CA, USA) to meet the required settings. Thereafter, pictures were analysed with the CeleST software (version 3.1; distributed by GitHub Pages, https://github.com/DCS-LCSR/CeleST, accessed on 08 April 2020) as described by Restif et al [61] and Ibáñez-Ventoso, et al [124]. The thrashing rate, the body wave number and the activity index were evaluated as representative parameters of motor activity.…”

Multi-environment phenotyping ofC. elegansfor robust evaluation of physical performance

“…Our data support the notion that the burrowing assay enhances the dynamic range in which we can evaluate drug interventions that counter dys-1 deficits. [26,35,36], crawling on NGM plates [22,27,[44][45][46], and burrowing [24,31] have been documented, but none have investigated individuals in these different environments in the context of assessing Fig. 6.…”

“…CeleST swim assay C. elegans Swim Test (CeleST) assays were conducted as previously described [35,36]. Animals were picked and placed into a 50 µL aliquot of M9 Buffer on a specialized glass slide with two, 10-mm pre-printed rings on the surface (Thermo Fisher Scientific), as previously used by the original developers of the CeleST assay.…”

“…The second physical environment we tested is liquid, in which swim motion is assayed using the C. elegans Swim Test (CeleST), analyzed with software that evaluates 8 measures related to body morphology and activity [35,36]. The four morphological measures include (i) body wave number -a measure of waviness of the body posture, (ii) asymmetry -the degree to which the animal bends more toward one side or the other, (iii) stretch -a measure of how deep or flat the body bends are, and (iv) curling -percentage of time that an animal spends overlapping with itself.…”