Sleep is an ancient behavioral state required for the maintenance of an animal's physiological health. Neuropeptide signaling has been shown to regulate sleep. During larval stage transitions, C. elegans larvae undergo a state of behavioral quiescence called lethargus, which exhibits sleep-like properties (Raizen et al, 2008). We find that behavioral quiescence is regulated by
nlp-22 and -23, which encode for FRPamide neuropeptides. Microarray analyses have shown that
nlp-22 is upregulated prior to lethargus and
nlp-23 is upregulated during lethargus. Over expression in young adulthood of
nlp-22 or
nlp-23 causes an inhibition of pharyngeal pumping, locomotion, and egg-laying. Animals over-expressing
nlp-22 or -23 are less responsive to olfactory and photic stimuli yet normally responsive to strong mechanical stimuli, demonstrating an elevated arousal threshold, a property of sleep. Interestingly,
nlp-22 (OE) worms woken by strong mechanical stimuli display a curious behavior of nearly continuous backwards movements for several minutes.
nlp-22 induced quiescence requires an intact signal sequence, suggesting that secretion of the peptide is required.
nlp-22 induced quiescence requires
egl-4, a gene required for quiescent behavior during lethargus (Raizen et al, 2008), but does not require
ceh-17, a gene required for the normal function of the ALA lethargus-regulating neuron (Van Buskirk and Sternberg, 2007). This suggests that
nlp-22 functions upstream of
egl-4 but downstream of or in parallel with ALA signaling during the regulation of lethargus.
Using a transgenic RNAi approach, we show that
nlp-22 is required for the proper induction of lethargus and for completion of the molts. We hypothesize that the molting-defective phenotype is a consequence of impaired lethargus regulation since, in separate experiments in our lab, we found that worms deprived mechanically or genetically of lethargus show varying degrees of molting defects and larval lethality (Unpublished). Our preliminary results indicate that
nlp-22(RNAi) worms enter lethargus but show a lower arousal threshold, suggesting impaired quality of sleep-like behavior.
nlp-22 fluorescent reporters containing the
nlp-22 3'UTR are expressed in a small set of neurons, most prominently in the pair of RIA interneurons. We are currently testing the hypothesis that
nlp-22 function is required in the nervous system and specifically in the RIA neurons. We will report additional experiments testing the neuroanatomical locus of
nlp-22 action and on our efforts to identify the
nlp-22 receptor.