Dimorphic Stress Hormones Figure 1. Manipulations of corazonin neurons alter levels of corazonin immunoreactivity. (A) Representative image of an adult brain stained with anti-corazonin expressing a GFP transgene in corazonin cells showing co-localization of immunosignals and GFP expression. (B) Representative image of an adult brain expressing the ork transgene. (C) Lack of corazonin immunosignals in brains expressing the reaper construct which induces apoptosis. For this image, we increased the brightness of the background to show that no cell bodies were stained with the corazonin antibody. (D) Representative brain from fly expressing the NaChBac transgene in corazonin neurons, which is aimed to facilitate peptide secretion, and note less intense staining in the soma of corazonin expressing neurons. (E). Quantification of immunosignals in each of these different treatments was performed on ten brains per genotype per sex. Images were collected under the same microscope settings. For quantification, a region of interest was drawn to include corazonin neuronal cell bodies and divided by the pixel intensity of a similar sized area of the brain lacking corazonin immunosignals. For animals expressing the reaper construct, there was no evidence of any corazonin immunolabels, preventing this analysis. Asterisks indicate significant differences (p,0.05) between the Crz-GFP ratios as determined by ANOVA. doi:10.1371/journal.pone.0009141.g001 (P,0.001, F = 80.75, R2 = 0.9735, Two-Way ANOVA). Intro- duction of the transformer element along with both the ork and NaChBac elements reversed or partially reversed the effects on locomotion exhibited by males expressing either element alone (Figure 3B). We then tested the impact of feminizing corazonin cells on survival during stress. Similarly, feminization of corazonin neurons in males bearing the NaChBac element partially reversed the stress sensitivity exhibited in males expressing the NaChBac element and tempered the phenotypes associated with male corazonin ork expression. Specifically, the median survival for silenced corazonin neurons was 51.960.7 hrs for starvation, 95.163.0 hrs for osmotic stress, and 180.064.1 hrs for oxidative stress. Feminiza- tion of this same genotype led to diminished effects of the silencing, with median survival being 48.060.2 hrs for starvation, 69.963.1 hrs for osmotic stress, and 132.060.6 for oxidative PLoS ONE | www.plosone.org 4 February 2010 | Volume 5 | Issue 2 | e9141