Alcohol potentiates a pheromone signal in flies

Abstract

For decades, numerous researchers have documented the presence of the fruit fly or Drosophila melanogaster on alcohol-containing food sources. Although fruit flies are a common laboratory model organism of choice, there is relatively little understood about the ethological relationship between flies and ethanol. In this study, we find that when male flies inhabit ethanol-containing food substrates they become more aggressive. We identify a possible mechanism for this behavior. The odor of ethanol potentiates the activity of sensory neurons in response to an aggression-promoting pheromone. Finally, we observed that the odor of ethanol also promotes attraction to a food-related citrus odor. Understanding how flies interact with the complex natural environment they inhabit can provide valuable insight into how different natural stimuli are integrated to promote fundamental behaviors.

Keywords: D. melanogaster; cis-vaccenyl acetate; complex environment; neuroscience; odor mixture; or67d; social behavior.

Figure 1.. Alcohol odor increases aggression in male flies.

(a) Traces of T1 sensilla recordings with a 300 ms exposure to air or vapor from 30% ethanol. (b) Hemolymph ethanol concentration (mg/mL) in flies in aggression arenas for 30 mins show no significant increases except with 20% ethanol (one-way ANOVA with Dunnett’s p<0.0001, n = 11–12). (c) Time spent fighting on ethanol-containing food (Control vs. 5% p=0.038 Kruskal-Wallis test with Dunn’s correction, n = 10–20). (d) Number of fights on ethanol-containing food (Control vs. 5% p=0.0012, statistical tests as in c). (e) Lunges on ethanol-containing food (Control vs. 5% p=0.0225, statistical tests as in c). (f) Latency to lunge (p=0.009, Log-rank Mantel-Cox with Bonferroni correction). (g) Cumulative latency of flies that lunged during the test (Control vs. 10% p=0.009, Log-rank Mantel-Cox with Bonferroni correction). (h) Locomotion as measured by line crossings during the test (Control vs. 20% p=0.0048, statistical tests as in 1 c). p<0.05 *; p<0.01 **; p<0.001 ***. Error bars denote SEM.

Figure 2.. Alcohol odor potentiates the response to cVa.

(a) Experimental timeline and diagram of recording site on fly antenna. (b) Traces of cVa-sensing T1 neurons. The red bar denotes 300 ms cVa exposure. (c) Δ Spikes calculated as cVa-induced activity (1 s during and after cVa) – spontaneous activity (paired two-tailed t-test, p=0.002, n = 7,15,15). (d) Spontaneous activity before, during, and after ethanol exposure. Spontaneous activity calculated as the total number of spikes 10 s prior to cVa delivery/10 s (Pre- vs. Post-5% p=0.002, Pre- vs. Post-30% p<0.0001, During vs. Post-30% p=0.0152, Kruskal-Wallis test with Dunn’s correction). (e), (f), (g) Averaged spikes over time for air, 5% ethanol, and 30% ethanol, respectively. Red bar denotes 300 ms cVa exposure (h) Time constant (τ) of decay of the cVa-induced spikes (Mann-Whitney test, p=0.0043). p>0.05, n.s. (not significant); p<0.05 *; p<0.01 **; p<0.0001 ****. Error bars denote SEM.

Figure 3.. Ethanol increases attraction to and potentiates the neuronal response of a food related odor.

(a) Paradigm used to evaluate farnesol responses pre- and post-ethanol treatment. Responses are shown in panels b-e. (b) Δ Spikes of farnesol-induced activity in ai2 sensilla, calculated as in 2b (paired two-tailed t-test, p=0.0059, n = 9–10). (c) Spontaneous activity before, during, and after ethanol exposure, calculated as in 2 c (Pre- vs. During Ethanol p=0.017, Pre- vs. Post-Ethanol p<0.0001, Kruskal-Wallis test with Dunn’s correction). (d, e) Traces from ai2 farnesol-sensing neurons. The red bar denotes a 300 ms farnesol exposure. (f) Graphic of the two-choice olfactory trap assay used to measure relative attraction to odors. (g) Preference Indices calculated as Number of flies in Choice 1-Number of flies in Choice 2/Total Number of flies. p>0.05, n.s. (not significant); p<0.05 *; p<0.01 **; p<0.0001 ****. Error bars denote SEM.

Author response image 1.

Author response image 2.