LNv and Morning Anticipation Figure 1. Large LNv highest action potential firing rates occur in the morning. (left, center panels) Representative traces of whole-cell current-clamp recordings of control large LNv (pdfGal4/dORK-NC1) taken during the day (left panel, ZT 1) and night (middle panel, ZT 22). Firing frequency (Hz) of 67 individual large LNv neurons plotted according to the time the recording was made, to the nearest half hour from whole brain preparations of flies aged 1–7 days (right panel). Recordings during the day displayed an overall higher firing frequency (3.260.4 Hz, n = 43, ZT 0–12) than those taken at night (2.460.4 Hz, n = 25, ZT 13–24), the dashed vertical line indicates the time of light-dark transition. The highest large LNv average firing frequencies are observed in the morning (4.561.5 Hz, n = 10, ZT 0–2) while the lowest large LNv average firing frequencies tend to occur at night. doi:10.1371/journal.pone.0011628.g001 Figure 2. Expression of Q128-Htt in LNv causes loss of circadian locomotor behavior. (A) Representative activity/rest records (actograms) of individual male flies expressing either control Q0-Htt (upper panels) or Q128-Htt (lower panels) in the LNv circadian pacemaker neurons. Left panels (yw) are genetic background controls where the UAS- PolyQ is not driven by GAL4, while right panels show flies where the expression of Q0- Htt or Q128-Htt is driven by the pdf-GAL4 driver in the large and small LNv in the adult brain. Flies were entrained in 12:12 h light/dark (LD12:12) regime from age day 2 to 6 following which they were subjected to constant darkness (DD) starting at day 7. The black and white horizontal bars above the top panels indicate the times during which the lights were OFF or ON, respectively, during the initial five days in LD, while the lower black bar indicates darkness during days 7–27. The x-axis indicates the time of day while y-axis indicates consecutive days. Genetic background controls of both Q0-Htt and Q128-Htt lines are predominantly rhythmic throughout the assay in DD. While pdf GAL4 driven Q0-Htt expressing flies are not significantly different from their genetic background controls, nearly all Q128-Htt flies are arrhythmic in DD. (B) Percentage of flies with arrhythmic activity/rest pattern during DD day 7–13, day 14–20 and day 21–27 for pdfGAL4 driven PolyQ expressing flies (right bars) and their genetic controls (left bars) There is no significant difference in fraction of arrhythmic flies from controls when Q0-Htt is expressed in LNv (upper panel), whereas nearly all Q128-Htt expressing flies are behaviorally arrhythmic for all age groups tested (lower panel). * indicates significant differences at p,0.01; Fisher’s exact p test. doi:10.1371/journal.pone.0011628.g002 PLoS ONE | www.plosone.org 3 July 2010 | Volume 5 | Issue 7 | e11628