Track-A-Worm 2.0: A Software Suite for Quantifying Properties of C. elegans Locomotion, Bending, Sleep, and Action Potentials

Comparative analyses of locomotor behavior and cellular electrical properties between wild-type and mutant Caenorhabditis elegans are crucial for exploring the gene basis of behaviors and the underlying cellular mechanisms. Although many tools have been developed by research labs and companies, their application is often hindered by implementation difficulties or lack of features specifically suited for C. elegans . Our system addresses these challenges with three key components: WormTracker , SleepTracker , and Action Potential (AP) Analyzer . WormTracker accurately quantifies a comprehensive set of locomotor and body bending metrics, incorporates user-identified dorsal and ventral orientation based on microscopic observation, continuously tracks the animal using a motorized stage, and seamlessly integrates external devices, such as a light source for optogenetic stimulation. SleepTracker detects and quantifies sleep-like behavior in freely moving animals. AP Analyzer assesses the resting membrane potential, afterhyperpolarization level, and various AP properties, including threshold, amplitude, mid-peak width, rise and decay times, and maximum and minimum slopes. Importantly, it addresses the challenge of AP threshold quantification posed by the absence of a preupstroke inflection point. This system is potentially a valuable tool for many C. elegans research labs due to its powerful functionality and ease of implementation.