Ethanol‐Regulated Genes That Contribute to Ethanol Sensitivity and Rapid Tolerance in Drosophila

Abstract

Background Increased ethanol intake, a major predictor for the development of alcohol use disorders, is facilitated by the development of tolerance to both the aversive and pleasurable effects of the drug. The molecular mechanisms underlying ethanol tolerance development are complex and are not yet well understood. Methods To identify genetic mechanisms that contribute to ethanol tolerance, we examined the time course of gene expression changes elicited by a single sedating dose of ethanol in Drosophila, and completed a behavioral survey of strains harboring mutations in ethanol-regulated genes. Results Enrichment for genes in metabolism, nucleic acid binding, olfaction, regulation of signal transduction, and stress suggests that these biological processes are coordinately affected by ethanol exposure. We also detected a coordinate up-regulation of genes in the Toll and Imd innate immunity signal transduction pathways. A multi-study comparison revealed a small set of genes showing similar regulation, including increased expression of 3 genes for serine biosynthesis. A survey of Drosophila strains harboring mutations in ethanol-regulated genes for ethanol sensitivity and tolerance phenotypes revealed roles for serine biosynthesis, olfaction, transcriptional regulation, immunity, and metabolism. Flies harboring deletions of the genes encoding the olfactory co-receptor Or83b or the sirtuin Sir2 showed marked changes in the development of ethanol tolerance. Conclusions Our findings implicate novel roles for these genes in regulating ethanol behavioral responses.

The FlyBar: Administering Alcohol to Flies

“…Observations beyond 1 hr should be avoided when possible because of the potential of rapid tolerance buildup in the flies [22][23][24] This protocol depends heavily on behavioral observations, so strict adherence to a standardized protocol is essential to avoid drift in behavioral observations over time. If possible, behavioral observations should be performed so that the observer is blind to the genotype or time point being tested.…”

“…The flies did not reach the 50% sedation mark within the hour as the observations were made under our standard LoRR conditions in order to make a direct comparison possible. Observations beyond the hour are problematic due to the buildup of rapid tolerance [22][23][24] . In this experiment, less than 25% of the flies at either circadian time point were sedated at 40 min, indicating that there is a difference in leading edge sedation sensitivity between these groups.…”

“…Observations are limited to 60 min to avoid the effects of rapid tolerance buildup [22][23][24] . The w 1118 mutants display significantly reduced LoRR sensitivity in response to ethanol vapor than does Canton-S (ANOVA between subjects F 1,10 = 57.12, p<0.001, N = 6).…”

Transcriptional Networks for Alcohol Sensitivity in Drosophila melanogaster

“…When exposed to alcohol, flies initially become hyperactive, but soon lose postural control and fall through the inebriometer at a rate that is correlated with their sensitivity. Following recovery, a second exposure reveals the development of tolerance reflected in a longer knockdown time (Moore et al 1998;Scholz et al 2000;Singh and Heberlein 2000;Cowmeadow et al 2005Cowmeadow et al , 2006Scholz et al 2005;Wen et al 2005;Morozova et al 2006Morozova et al , 2009Kong et al 2009). …”

Reduction in the cumulative effect of stress-induced inbreeding depression due to intragenerational purging in Drosophila melanogaster

“…Both heat and ethanol stress also induce upregulation of heat shock proteins in Drosophila Kong et al, 2010), however, this increased expression can also be costly for growth, development and longevity (see reviews in Sorensen et al, 2003). Therefore, irreversible damage to basic cellular functioning and long-term costs associated with mounting molecular stress defenses could offset in part the fitness benefits associated with stress-induced purging of genetic load under abiotic heat and ethanol stress.…”