Temperature cues are integrated in a flexible circadian neuropeptidergic feedback circuit to remodel sleep-wake patterns in flies.

Organisms detect temperature signals through peripheral neurons, which relay them to central circadian networks to drive adaptive behaviors. Despite recent advances in Drosophila research, how circadian circuits integrate temperature cues with circadian signals to regulate sleep/wake patterns remains unclear. In this study, we used the FlyWire brain electron microscopy connectome to map neuronal connections, identifying lateral posterior neurons LPNs as key nodes for integrating temperature information into the circadian network. LPNs receive input from both circadian and temperature-sensing neurons, promoting sleep behavior. Through connectome analysis, genetic manipulation, and behavioral assays, we demonstrated that LPNs, downstream of thermo-sensitive anterior cells (ACs), suppress activity-promoting lateral dorsal neurons LNds via the AstC pathway, inducing sleep Disrupting LPN-LNd communication through either AstCR1 RNAi in LNds or in an AstCR1 mutant significantly impairs the heat-induced reduction in the evening activity peak. Conversely, optogenetic calcium imaging and behavioral assays revealed that cold-activated LNds subsequently stimulate LPNs through NPF-NPFR signaling, establishing a negative feedback loop. This feedback mechanism limits LNd activation to appropriate levels, thereby fine-tuning the evening peak increase at lower temperatures. In conclusion, our study constructed a comprehensive connectome centered on LPNs and identified a novel peptidergic circadian feedback circuit that coordinates temperature and circadian signals, offering new insights into the regulation of sleep patterns in Drosophila.