Neuropeptide modulation in circadian neural circuits

PROJECT SUMMARY/ ABSTRACT The biological circadian clock generates a series of time markers across the 24 h day, by which different aspects of physiology and behavior (e.g., sleep, hormone release, temperature dynamics) may be aligned to local time for optimal efficiency. The molecular mechanisms of the PER-dependent circadian clock (which is a cell autonomous timekeeping system), and how the clock is set to local time, are both well-studied. Less is known about how the molecular clock couples within pacemaker cells to cell physiology, and how the clock uses intercellular communication to generate the multiple phase (markers) that are used by other clock cells, and by non-clock bearing downstream cells and circuits. This laboratory studies circadian neurophysiology in the model system Drosophila and the overall goal of this project is to understand how intercellular communication among pacemakers is used to generate different phases of circadian output. The fly’s daily rhythmic locomotor behavior is controlled by ~150 dedicated circadian pacemaker neurons whose internal molecular clocks are highly synchronized. Neuropeptide signaling in circadian neural circuits of both insects and mammals supports molecular synchrony: in Drosophila, the neuropeptide PDF (via the PDF receptor, a G Protein-Coupled Receptor (GPCR)) serves this function. Importantly, PDF modulation supports normal pacemaker circuitry in a second critical way: it inhibits the neuronal activity of identified pacemaker groups to specific non-morning phases. Prior work generated a model whereby the cell-intrinsic molecular clock in all pacemakers drives a common morning phase of enhanced neuronal activity. Yet in vivo, real-time measurements of calcium dynamics reveal that different pacemaker groups in the Drosophila brain are not synchronously active in the morning. Instead the different groups exhibit a stereotyped sequence of activity periods, at different times of the day and night. Hence this diversity of active periods (circadian phases) is generated primarily by cell interactions (especially neuropeptide modulation). Together these activity periods represent the poly-phasic outputs of the pacemaker system. Now, to test and extend the model, this research program will evaluate the hypothesis that two distinct forms of the PDF receptor may differentially contribute to molecular clock synchrony versus to inhibition of pacemaker neuronal activity. Experiments are also designed to identify the signaling pathways used by the receptor isoforms. Finally, work will extend studies based on the recent identification of a membrane protein that interacts with PDF receptor: an evolutionarily conserved magnesium ion transporter which co-regulates locomotor behavior along with PDF receptor. Together, this work performed in Drosophila will also inform an understanding of circadian output mechanisms in the mammalian brain, because the fundamental mechanisms of GPCR modulation are highly conserved. More generally, it will be relevant to fundamental mechanisms of neural circuit modulation by neuropeptides.