, 2009), cell autonomous activation of PDF-R solely in PDF-negative pacemaker neurons with a membrane-tethered GW786034 PDF construct promotes strong rhythmicity in Pdf null flies, which would otherwise be poorly rhythmic ( Choi et al., 2012). Flies deficient in PDF or PDF-R display severe deficits in circadian rhythms and alterations in PER molecular rhythms during constant dark (DD) conditions. Among the four small LNvs, rhythms are maintained but become desynchronized (Lear et al., 2005; Lin et al., 2004). Among PDF target pacemaker groups like the LNd, the amplitude and period of the PER rhythm decrease but cells remain synchronized (Lin et al., 2004; Yoshii et al., 2009). Thus PDF neuropeptide acts over

many daily cycles to

promote the amplitude and pace of PER cycling—it has access to the molecular clockworks in diverse pacemakers and affects them differently. Recent observations have begun to shed light on the signaling pathways by which PDF affects synchronization and how these may differ according to cell type. Because PDF modulation system profoundly affects the circadian molecular oscillator within individual pacemaker neurons, the molecular learn more details of the signaling pathway downstream of PDF-R gains in significance. Among the identified neurons in the pacemaker network, the PDF-expressing subset are termed M cells based on their abilities to influence “Morning” activity levels; several non-PDF pacemakers are termed E cells based on their abilities to influence “Evening” time activity levels (Grima et al., 2004; Stoleru et al., 2004; Yoshii et al., 2004; reviewed by Helfrich-Förster, 2009). Duvall and Taghert (2012) recently used an RNAi-mediated genetic approach to report that adenylate cyclase 3 (AC3) underlies PDF signaling in M cells. Surprisingly, disruption of AC3 does not alter PDF-R mediated only responses in non-PDF pacemakers (specifically, in the PDF-R(+) LNd). Moreover, AC3 disruptions in small LNv did not alter GPCR signaling by other ligands that elevate cAMP levels in these neurons (dopamine and the neuropeptide DH31). Hence, within small LNv, PDF-R

signaling occurs via discrete molecular pathways that are distinct from those controlled by other cAMP-elevating ligands. This provides a molecular mechanism underlying the observation that PDF-R activation in small LNv has potent effects on daily allocation of rest and activity, whereas DH31-R activation does not ( Choi et al., 2012). Furthermore, PDF-R association with a different AC(s) supports PDF signaling in the other circadian pacemakers. Thus critical pathways of circadian synchronization are mediated by highly specific second messenger components. These findings support a hypothesis that PDF signaling components within target cells are sequestered into “circadian signalosomes,” whose compositions differ between different pacemaker cell types ( Duvall and Taghert, 2012).