Automatically tracking feeding behavior in populations of foraging C. elegans

Abstract

Caenorhabditis elegans feeds on bacteria and other small microorganisms which it ingests using its pharynx, a neuromuscular pump. Currently, measuring feeding behavior requires tracking a single animal, indirectly estimating food intake from population-level metrics, or using restrained animals. To enable large throughput feeding measurements of unrestrained, crawling worms on agarose plates at a single worm resolution, we developed an imaging protocol and a complementary image analysis tool called PharaGlow. We image up to 50 unrestrained crawling worms simultaneously and extract locomotion and feeding behaviors. We demonstrate the tool’s robustness and high-throughput capabilities by measuring feeding in different use-case scenarios, such as through development, with genetic and chemical perturbations that result in faster and slower pumping, and in the presence or absence of food. Finally, we demonstrate that our tool is capable of long-term imaging by showing behavioral dynamics of mating animals and worms with different genetic backgrounds. The low-resolution fluorescence microscopes required are readily available in C. elegans laboratories, and in combination with our python-based analysis workflow makes this methodology easily accessible. PharaGlow therefore enables the observation and analysis of the temporal dynamics of feeding and locomotory behaviors with high-throughput and precision in a user-friendly system.

Mitochondrial energy state controls AMPK-mediated foraging behavior in C. elegans

“…Intriguingly, we found that AMPK re-expressed in serotonergic or dopaminergic neurons is sufficient to restore the global search behavior. While previous studies have reported the role of serotonergic and dopaminergic neurons in modulating food behavioral responses (43,44,(48)(49)(50)(51)(52), we have shown that AMPK activation in these neurons is also required. Overall, we have shown that PMF dissipation reduces ATP production (Fig.…”

Adapting and optimizing GCaMP8f for use inCaenorhabditis elegans

“…Using animals expressing either GCaMP7f or GCaMP8f in the pharyngeal muscle, we imaged groups of worms on a plate seeded with food. Compared to our previous work (Bonnard et al . 2022), we could detect animals labeled with GCaMP8f at even lower magnifications (0.5x instead of 1x), enabling a field of view of 1.5 cm (Fig.…”

“…While pumping can be detected without fluorescent labels, this requires imaging at higher magnifications and often laborious post-processing. We previously found that expressing a non-calcium dependent fluorophore enables detecting pumping at lower magnification and in freely moving animals (Bonnard et al 2022). Using animals expressing either GCaMP7f or GCaMP8f in the pharyngeal muscle, we imaged groups of worms on a plate seeded with food.…”

“…Images were analyzed using Pharaglow (Bonnard et al 2022). The peaks of the resulting intensity traces were found using the built-in peak detection and an average pumping rate per tracklet was calculated as N pumps /duration.…”

The Bacterial Gq Signal Transduction Inhibitor Fr900359 Impairs Soil-associated and Plant Pathogenic Nematodes

“…Animals were imaged at 15 fps for 5 min unless otherwise indicated. Animals were tracked using the tracking package trackpy (Allan et al, 2019) with a custom detection script in Python based on the pharaglow package (Bonnard et al, 2022). The animal speed was calculated from the resulting center-of-mass coordinates as follows: The trajectories (x, y, t) were subsampled from 14 fps to 2.8 fps and the speed was calculated as with .…”