(a) Fighting frequencies of flies with sbb or atro knocked down in the PI using the 50Y driver and control lines containing only one component of the binary expression system. Only 50Y>AtroIR and 50Y>AtroRNAi flies have significantly increased fighting frequencies (Kruskal–Wallis ANOVA, **P<0.01 and ***P<0.001, n=minimum of 70 pairs for each genotype). (b) Immunofluorescence image of a slice of an adult male brain stained with anti-Atro antibody. The PI is indicated with a white arrowhead. Scale bar=100 μm. (c) Fighting frequencies of flies in which tll alone is knocked down, in which tll and atro are simultaneously knocked down in the pars intercerebralis using the 50Y driver, and control lines containing only one component of the GAL4/UAS system. Asterisks denote statistically significant higher medians in the tll and tll/Atro knockdown males compared with the single binary component controls (Kruskal–Wallis ANOVA, ***P<0.001, n=minimum of 60 pairs for each genotype). Bar graphs represent means±s.e.m.