| Summary: | Sleep is an evolutionarily conserved biological process in essentially all animals with a nervous system, but the core function of sleep remains unknown. One of the most proven strategies for understanding complex phenomena is to perturb simple systems that model the complex phenomenon of interest. With this in mind, this dissertation uses lethargus, a 2-3 hour period of developmentally-timed sleep, in the nematode Caenorhabditis elegans to explore how quiescence is homeostatically regulated and how disturbances during this period affect the organism.
Here, I report that two behaviorally and genetically distinct mechanisms regulate behavioral quiescence during lethargus. I show that weak stimuli evoke homeostasis in response to extended motion by extending the subsequent bout of quiescence. I find that neuropeptides are required for this homeostatic response. Specifically, I find that the neuropeptide-Y receptor homolog NPR-1 in the RMG neuron class and inhibitory FLP-18 peptides are necessary for this "weak" homeostatic response. In contrast, strong stimuli repeated throughout lethargus result in a distinct homeostatic response: an elevation of the overall time spent in quiescence during lethargus. This "strong" homeostatic response does not require NPR-1, but instead requires the function of DAF-16/FOXO insulin-signaling transcription factor in neurons. Conversely, the "weak" homeostatic response does not require DAF-16/FOXO.
I further show that lethargus is a specifically vulnerable period during C. elegans development. I report that nonlethal deprivation of quiescence during lethargus beyond the regime that evokes homeostasis results in proteotoxic stress, that if left unmitigated, can cause long-lasting deficits in the animal. These deficits are anatomically, functionally, and genetically distinct. Specifically, I describe an automated protocol for depriving C. elegans of developmentally-timed sleep in a severe yet nonlethal manner. I then characterize three lasting effects of nonlethal deprivation of quiescence: a deficiency in pharyngeal pumping, a reduction in brood size as a result of germ cell apoptosis, and an excess of twitching in the vulval muscles of young adults. Importantly, our protocol for depriving quiescence when applied outside of lethargus does not result in deficits. I show that while both the pumping and fecundity defects are mitigated by DAF-16/FOXO, deprivation evokes distinct proteotoxic stress responses in the germline and pharynx. The mitochondrial UPRMT mitigates the pumping defect while the endoplasmic reticulum UPRER mitigates the fecundity and vulval muscle defects.
Taken together, this work identifies a number of new insights about a primitive form of sleep. This thesis shows that neuropeptides modulate quiescence during lethargus, but that strong perturbations evoke a distinct behavioral and genetic response to increase quiescence. This thesis also shows that stronger perturbations beyond the regime of homeostasis result in the activation of distinct stress responses in distinct tissues and circuits of the animal. Importantly, these same perturbations inside and outside of lethargus evoke stress responses only during lethargus. Therefore, I show that lethargus is a vulnerable period of C. elegans development.
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