| Lethargus is a sleep-like developmental state that occurs between larval stages in the nematode Caenorhabditis elegans around the time when it molts. Lethargus is characterized by reversible behavioral quiescence, a stereotypical posture, and homeostasis. The complex behavioral patterns during lethargus can be dissected to quantifiable elements, which exhibit rich temporal dynamics and are actively regulated by the nervous system. In order to study this state in detail, we developed a collection of methods to measure and perturb the behavior of the worm both within and outside of lethargus. We observed that lethargus behavior is composed of alternating bouts of quiescence and motion which last for a few seconds to a few minutes, and quiescence duration depends partially on the duration of the previous bout of motion. During lethargus, the worm regularly adopts an otherwise atypically straight posture, often with only a single bend; we show that such a posture is mechanically required to generate the axial rotations or "flips" that it performs in order to loosen the cuticle in preparation for the molt. Using the posture-detecting software we developed, we also show that awake worms exhibit consolidated epochs of directed motion and quiet wakefulness. During lethargus, however, sustained forward motion is suppressed. Finally, we studied the homeostasis of lethargus quiescence in detail by applying both weak and strong perturbations in motion bout duration via aversive stimuli; we found that weak perturbations evoke local homeostasis in the form of increased quiescence in the minute immediately following stimulus, while strong perturbations evoke global homeostasis which manifests as an increased level of baseline quiescence in worms which receive stimulus regularly throughout lethargus. These two responses require separate molecular pathways to function. Similarly, we find that worms exposed to extreme amounts of stimulus will compensate either locally (by ignoring ongoing stimulus after about 2--3 minutes) or globally (by completely ignoring stimuli that cycle on and off more rapidly than that). By developing techniques to accurately measure the behavioral output of the nervous system, it is possible to observe subtle phenotypic effects on long-timescale phenomena produced by the many genetic, cellular, and environmental manipulations possible in C. elegans, towards an eventual understanding of the function of a simple nervous system. |
