Investigating Drosophila Circadian Rhythms

Flies were raised in a 12 h:12 h light:dark (LD) cycle at 25°C in vials containing standard cornmeal medium. For these experiments, we use the following stocks: w¹¹¹⁸ (RRID:BDSC_5905), UAS-Kir2.1 (Nitabach et al., 2002 (link)), UAS-dTrpA1 (Rosenzweig et al., 2008 (link)), UAS-mCD8GFP, Clk856-GAL4 (Gummadova et al., 2009 (link)) and OK107-GAL4 (MB) driver) that were obtained from the Bloomington Stock Center. We used the same group of enhancer trap lines used by Gorostiza et al. (2014) (link): 3-86-GAL4, 11-8-GAL4, 4-12-GAL4, 4-93-GAL4, 5-133-GAL4, 4-59-GAL4, 5-43-GAL4 and 7-49-GAL4. These lines were a gift from U. Heberlein (Janelia Farm, USA). For the optical imaging experiments we used the following fly lines: pdf-LexA (Shang et al., 2008 (link)), UAS-GCaMP3 (Tian et al., 2009 (link)), LexAop-P2X2, pdf-GAL4 (Renn et al., 1999 (link)) (RRID:BDSC_6900), Clk4.1-GAL4 (Zhang et al., 2010 (link)), Clk4.5-GAL4, Mai179-GAL4>pdf-GAL80 and tim-GAL4>pdf-GAL80 (Emery et al., 1998 (link)) (RRID:BDSC_7126). We generated the experimental fly lines crossing the different GAL4s to the pdf-LexA,UAS-GCaMP3>LexAop-P2X2 line. The UAS-GCaMP3 was obtained from Janelia Farm and the LexAop-P2X2 was a gift from O. Shafer (University of Michigan) (Yao et al., 2012 (link)). All experimental protocols were performed in accordance with relevant guidelines and ethical regulations of our institution.