In metazoan organisms, circadian (∼24 h) rhythms are regulated by pacemaker neurons organized in a master–slave hierarchy. Although it is widely accepted that master pacemakers and slave oscillators generate rhythms via an identical negative feedback loop of transcription factor CLOCK (CLK) and repressor PERIOD (PER), their different roles imply heterogeneity in their molecular clockworks. Indeed, in Drosophila, defective binding between CLK and PER disrupts molecular rhythms in the master pacemakers, small ventral lateral neurons (sLNvs), but not in the slave oscillator, posterior dorsal neuron 1s (DN1ps). Here, we develop a systematic and expandable approach that unbiasedly searches the source of the heterogeneity in molecular clockworks from time-series data. In combination with in vivo experiments, we find that sLNvs exhibit higher synthesis and turnover of PER and lower CLK levels than DN1ps. Importantly, light shift analysis reveals that due to such a distinct molecular clockwork, sLNvs can obtain paradoxical characteristics as the master pacemaker, generating strong rhythms that are also flexibly adjustable to environmental changes. Our results identify the different characteristics of molecular clockworks of pacemaker neurons that underlie hierarchical multi-oscillator structure to ensure the rhythmic fitness of the organism.

Metabolism influences locomotor behaviors, but the understanding of neural curcuit control for that is limited. Under standard light-dark cycles, Drosophila exhibits bimodal morning (M) and evening (E) locomotor activities that are controlled by clock neurons. Here, we showed that a high-nutrient diet progressively extended M activity but not E activity. Drosophila tachykinin (DTk) and Tachykinin-like receptor at 86C (TkR86C)-mediated signaling was required for the extension of M activity. DTk neurons were anatomically and functionally connected to the posterior dorsal neuron 1s (DN1ps) in the clock neuronal network. The activation of DTk neurons reduced intracellular Ca2+ levels in DN1ps suggesting an inhibitory connection. The contacts between DN1ps and DTk neurons increased gradually over time in flies fed a high-sucrose diet, consistent with the locomotor behavior. DN1ps have been implicated in integrating environmental sensory inputs (e.g., light and temperature) to control daily locomotor behavior. This study revealed that DN1ps also coordinated nutrient information through DTk signaling to shape daily locomotor behavior.

Sleep is a fundamental behavior in an animal’s life influenced by many internal and external factors, such as aging and diet. Critically, poor sleep quality places people at risk of serious medical conditions. Because aging impairs quality of sleep, measures to improve sleep quality for elderly people are needed. Given that diet can influence many aspects of sleep, we investigated whether a high-sucrose diet (HSD) affected aging-induced sleep fragmentation using the fruit fly, Drosophila melanogaster . Drosophila is a valuable model for studying sleep due to its genetic tractability and many similarities with mammalian sleep. Total sleep duration, sleep bout numbers (SBN), and average sleep bout length (ABL) were compared between young and old flies on a normal sucrose diet (NSD) or HSD. On the NSD, old flies slept slightly more and showed increased SBN and reduced ABL, indicating increased sleep fragmentation. Short-term maintenance of flies in HSD (up to 8 days), but not long-term maintenance (up to 35 days), suppressed aging-induced sleep fragmentation. Our study provides meaningful strategies for preventing the deterioration of sleep quality in the elderly.