The neuropeptide pigment-dispersing factor (PDF) has been studied extensively in Drosophila, and its role in circadian time-keeping has been firmly established. The role of PDF outside of the clock circuit, however, is poorly understood. A recent study suggested that PDF may act on the ellipsoid body (EB) to link the clock and sleep/activity circuits. We performed whole brain optical imaging with the fluorescence resonance energy transfer (FRET)-based cAMP sensor Epac1-camps expressed under control of the pdfR promoter to address how the clock and sleep deprivation affect the physiology of these cells. Basal cAMP levels in EB were regulated both by PDF and synaptic inputs that are controlled by the circadian clock. Acute application of PDF to the brain caused a significant, and PDF-receptor-dependent, increase in cAMP in EB cells. Application of TTX to block circuit-mediated effects of PDF increased the morning response but not the response at night, implying the existence of a temporally regulated, PDF-stimulated input that blocks cAMP generation. ACh produced both direct (TTX-insensitive) and indirect (TTX-sensitive) increases in cAMP during the day but was totally TTX-insensitive at night, indicating that ACh-stimulated inputs to the EB are suppressed at night. Sleep deprivation did not affect the cAMP responses of these cells to either PDF or ACh. These results suggest a novel role for PDF as a modulator of activity outside of the clock circuit. By elucidating the mechanisms by which the neuropeptide PDF act on its target cells, our work contributes to our understating of how the central clock coordinates activity and sleep.
Keywords: Drosophila; cAMP imaging; circadian clock; ellipsoid body; neuropeptide PDF; sleep.