Response to stress in Drosophila is mediated by gender, age and stress paradigm

Abstract

All living organisms must maintain equilibrium in response to internal and external challenges within their environment. Changes in neural plasticity (alterations in neuronal populations, dendritic remodeling, and synaptic turnover) are critical components of the homeostatic response to stress, which has been strongly implicated in the onset of affective disorders. However, stress is differentially perceived depending on the type of stress and its context, as well as genetic background, age and sex; therefore, an individual's maintenance of neuronal homeostasis must differ depending upon these variables. We established Drosophila as a model to analyze homeostatic responses to stress. Sexually immature and mature females and males from an isogenic wild-type strain raised under controlled environmental conditions were exposed to four reproducible and high-throughput translatable stressors to facilitate the analysis of a large number of animals for direct comparisons. These animals were assessed in an open-field arena, in a light-dark box, and in a forced swim test, as well as for sensitivity to the sedative effects of ethanol. These studies establish that immature and mature females and males represent behaviorally distinct populations under control conditions as well as after exposure to different stressors. Therefore, the neural substrates mediating the stress response must be differentially expressed depending upon the hormonal status of the brain. In addition, an adaptive response to a given stressor in one paradigm was not predictive for outcomes in other paradigms.

Keywords: Ethanol sedation; forced swim test; light–dark box; open-field arena; sexual dimorphism; temporal dimorphism.

Fig. 1. Immature and mature female and male Drosophila display distinct motor patterns

Data were collected from min 15 – 20 in an open field arena, during which time the animals had habituated to the novel environment. A – C, motor pattern. A, mean distance moved; B, mean velocity; C, mean turn angle. D –E. Freezing behavior. D, duration not moving; E, frequency not moving. F – G, escape behavior. These included hops of velocity greater than 50 mm/s, as well as short bursts of flight. F, duration highly mobile; G, frequency of high mobility * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, two-way ANOVA followed by Bonferroni post tests. n = 40 for each population..

Fig. 2. Sexually mature males and females display dimorphic and distinct responses in three different paradigms relative to immature flies

A - B, light - dark box. A, Amount of time spent in light (min 10–15 of the 15 min observation period). B, Number of transitions between the light and dark sides of the arena during min 10–15 of the 15 min observation period. C – E, Forced Swim Test. C, latency to first bout of immobility; D, number of bouts; E, average bout duration. F – G, response to the sedative effects of ethanol. F, sec until sedation; G, sec until recovery from sedation. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, two-way ANOVA followed by Bonferroni post tests. n = 30 for each population.

Fig. 3. The effect of stressors on time spent in the lit arena in the Light-Dark Box

A, sexually immature females; B, sexually immature males; C, sexually mature females; D, sexually mature males. Statistical comparisons between control and stress conditions within a population, ** p < 0.01, one-way ANOVA followed by Dunnet’s post test.

Fig. 4. The effect of stressors on number of transitions in the Light-Dark Box

A, sexually immature females; B, sexually immature males; C, sexually mature females; D, sexually mature males. Statistical comparisons between control and stress conditions within a population, * p < 0.05, *** p < 0.001, one-way ANOVA followed by Dunnet’s post test.

Fig. 5. The effect of stressors on behavioral parameters of the Forced Swim Test

A – D, latency to first bout of immobility; E – H, number of bouts; I – L, bout duration. A, E, I sexually immature females; B, F, J sexually immature males; C, G, K sexually mature females; D, H, L, sexually mature males. Statistical comparisons between control and stress conditions within a population, * p < 0.05, ** p < 0.01, *** p < 0.001, one-way ANOVA followed by Dunnet’s post test. n = 30 for each population.

Fig. 6. The effect of stressors on immobility with increasing time during the Forced Swim Test

A, sexually immature females; B, sexually immature males; C, sexually mature females; D, sexually mature males. Statistical comparisons between control and stress conditions for each timepoint within a population, * p < 0.05, ** p < 0.01, *** p < 0.001, oneway ANOVA followed by Dunnet’s post test. n = 30 for each population.

Fig. 7. The effect of stressors on the sensitivity to the sedative effects of ethanol

A, sexually immature females; B, sexually immature males; C, sexually mature females; D, sexually mature males. Statistical comparisons between control and stress conditions within a population, ** p < 0.01, one-way ANOVA followed by Dunnet’s post test. n = 30 for each population.

Fig. 8. The effect of stressors on the rate of recovery from ethanol sedation

A, sexually immature females; B, sexually immature males; C, sexually mature females; D, sexually mature males. * p < 0.05, ** p < 0.01, ***, p < 0.001, one-way ANOVA followed by Dunnet’s post test. n = 30 for each population.