Figure 3. Behavioral responses of mir-124 mutants to environmental perturbations. A, B, Light-resetting analysis. A, Following LD entrainment, a 10′ light pulse was delivered at Zeitgeber time 21 (i.e., 3 h prematurely) before moving to DD (see Materials and Methods). The light pulse correspondingly shifts the evening peak in all genotypes analyzed. B, Quantification of the subjective evening peak in the first 2 d following shift to DD conditions, without and with a premature light pulse. C, D, Analysis of temperature cycling in DD; black bars indicate 20°C; white bars indicate 29°C. The evening activity peak of mir-124 mutants is still advanced under temperature cycling. E, F, Analysis of temperature-entrained animals, following release to constant conditions (25°C). Note that the phase of all genotypes shifts, but mir-124 mutants exhibit greater phase advance. These assays use the same cohorts from C and D, except dead animals were excluded from analysis; mir-124 mutants have increased lethality following TC. n = number of male animals analyzed. We used Mann–Whitney statistical tests for pairwise comparisons. For light-resetting analysis, we indicate significance values between controls and mutants in both conditions, and for each cognate genotype with and without light pulse (shown as color-coded p values). Error bars indicate SEM.