Flies were pre-warmed at 30°C such that the dTrpA1-activated cleaning movement was being performed at the time of dusting. They were shaken with or without dust and allowed to groom while their cleaning movements were recorded for behavioral analysis. (A) Representative manually annotated ethograms of GAL4 lines expressing dTrpA1 that were shaken with and without dust. The most common mutually exclusive movements performed by individual flies are displayed. R53A06 and R45G01 were not scored beyond 15 min because they had already progressed through the cleaning sequence. (B–E) Most head cleaning bouts occur at the beginning of grooming both in wild-type flies and when other cleaning movements are artificially activated. Number of cleaning movements for each major body part in 100 s intervals after flies were shaken without or with dust (plotted as the mean ± SEM; n = 5). Different lines displayed are: control (B), R40F04-GAL4/UAS-dTrpA1 (head cleaning) (C), R24B03-GAL4/UAS-dTrpA1 (abdominal cleaning) (D), R53A06-GAL4/UAS-dTrpA1 (wing cleaning) (E). Artificially activated movements are displayed as lines with filled circles at each time interval. Dust stimulated cleaning movements (not artificially activated) are shown as lines with no filled circles. All head cleaning movements are combined (labeled whole head). Friedman tests show a statistical difference between points in each head cleaning curve with dusted flies. This demonstrates that the numbers of head cleaning bouts at the beginning of grooming are higher than those at the end of the time course (p values for each line shown as undusted, dusted). Control (p = 0.817, p = 0.0004), R40F04 (p = 0.147, p = 0.00008), R24B03 (p = 0.3038, p = 0.0038), R53A06 (p = 0.0599, p = 0.0027). Other GAL4 lines showed similar results to these examples.