Select Drosophila glomeruli mediate innate olfactory attraction and aversion

Abstract

Fruitflies show robust attraction to food odours, which usually excite several glomeruli. To understand how the representation of such odours leads to behaviour, we used genetic tools to dissect the contribution of each activated glomerulus. Apple cider vinegar triggers robust innate attraction at a relatively low concentration, which activates six glomeruli. By silencing individual glomeruli, here we show that the absence of activity in two glomeruli, DM1 and VA2, markedly reduces attraction. Conversely, when each of these two glomeruli was selectively activated, flies showed as robust an attraction to vinegar as wild-type flies. Notably, a higher concentration of vinegar excites an additional glomerulus and is less attractive to flies. We show that activation of the extra glomerulus is necessary and sufficient to mediate the behavioural switch. Together, these results indicate that individual glomeruli, rather than the entire pattern of active glomeruli, mediate innate behavioural output.

Figure 1. Flies are robustly attracted to apple cider vinegar, which excites six glomeruli

a, Path of a single fly with 3 ppm vinegar in the upper left quadrant. b, c, Density plot of 20 w¹¹¹⁸ and Or83b^−/− flies. d, Performance index of w¹¹¹⁸ and Or83b^−/−flies. *** indicates P < 0.001; T-test. e, g, Pre-stimulation images showing glomerular structure. f, h, Responses to 3 ppm vinegar in flies bearing the GH146-Gal4 and UAS-GCaMP transgenes. i, Quantification of ΔF/F for all six glomeruli over ten flies. Error bars indicate s.e.m.

Figure 2. Silencing DM1 or VA2 reduces attraction to 3 ppm vinegar

a–f, Density plots composed of 20 flies each. g, Performance indices for flies bearing the OrX-Gal4 and UAS-shi^ts transgenes at permissive and nonpermissive temperatures. ANOVA followed by Tukey’s test was performed on PI values from flies of the experimental group at the permissive and nonpermissive temperatures, and the corresponding genetic background controls at the nonpermissive temperature. ***, P < 0.001; and **, P < 0.01. Error bars indicate s.e.m.

Figure 3. Restoring Or83b in DM1 and VA2 ORNs restores attraction to control levels

a–f, Density plots of 20 flies responding to 3 ppm vinegar. g, Performance indices of flies in which Or83b is selectively restored in individual ORN types. Comparisons between groups were made using ANOVA followed by Tukey’s test. Significant differences (P < 0.05) are denoted by letters. Error bars indicate s.e.m.

Figure 4. Vinegar becomes less attractive and activates an additional glomerulus at high concentrations

a, Performance indices of w¹¹¹⁸ flies at various concentrations of vinegar. PI values were compared using ANOVA followed by Tukey’s test. Significant differences (P < 0.05) are denoted by letters. b,c, Density plots of w¹¹¹⁸ behavior in response to 12 ppm and 32 ppm vinegar. d, Responses to 12 ppm and 32 ppm vinegar in flies bearing the GH146-Gal4 and UAS-GCaMP transgenes. e, Average ΔF/F. ** indicates P < 0.01; T-test. Error bars indicate s.e.m.

Figure 5. DM5 mediates the decrease in attraction in response to 32 ppm vinegar

a, Density plot of twenty flies in which the DM5 ORNs are silenced. b, Density plot of 20 DM5 rescue flies. c, Behavioral responses to 32 ppm vinegar for flies in which DM5 and DM1 are silenced and selectively rescued. For silencing experiments, we performed the same statistical analysis as for Figure 2. DM5 rescue and DM1 rescue flies were compared to Or83b^−/− flies by T-test. ***, P < 0.001; and **, P < 0.01.

Figure 6. DM1 and DM5 mediate attraction and aversion in response to ethyl butyrate

a, Performance indices in response to 7 ppm ethyl butyrate for DM1 and DM5 rescue flies. ***, P < 0.001; T-test. b, Ectopic expression of Or22a in Or85a ORNs reduced attraction to 12 ppm vinegar. PI values were compared using ANOVA followed by Tukey’s test. *** indicates P < 0.001.