Rapid and Chronic: Two Distinct Forms of Ethanol Tolerance in Drosophila

Berger, Karen H.; Heberlein, Ulrike; Moore, Monica

doi:10.1097/01.alc.0000141817.15993.98

Cited by 81 publications

(109 citation statements)

References 37 publications

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“…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).…”

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confidence: 99%

“…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.…”

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The FlyBar: Administering Alcohol to Flies

Linde

Fumagalli

Roman

et al. 2014

JoVE

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Fruit flies (Drosophila melanogaster) are an established model for both alcohol research and circadian biology. Recently, we showed that the circadian clock modulates alcohol sensitivity, but not the formation of tolerance. Here, we describe our protocol in detail. Alcohol is administered to the flies using the FlyBar. In this setup, saturated alcohol vapor is mixed with humidified air in set proportions, and administered to the flies in four tubes simultaneously. Flies are reared under standardized conditions in order to minimize variation between the replicates. Three-day old flies of different genotypes or treatments are used for the experiments, preferably by matching flies of two different time points (e.g., CT 5 and CT 17) making direct comparisons possible. During the experiment, flies are exposed for 1 hr to the pre-determined percentage of alcohol vapor and the number of flies that exhibit the Loss of Righting reflex (LoRR) or sedation are counted every 5 min. The data can be analyzed using three different statistical approaches. The first is to determine the time at which 50% of the flies have lost their righting reflex and use an Analysis of the Variance (ANOVA) to determine whether significant differences exist between time points. The second is to determine the percentage flies that show LoRR after a specified number of minutes, followed by an ANOVA analysis. The last method is to analyze the whole times series using multivariate statistics. The protocol can also be used for non-circadian experiments or comparisons between genotypes. Video LinkThe video component of this article can be found at

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confidence: 99%

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The FlyBar: Administering Alcohol to Flies

Linde

Fumagalli

Roman

et al. 2014

JoVE

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“…27 Previous work in Drosophila has shown that the development of chronic tolerance is blocked by cyclohexamide treatment, suggesting that it requires protein synthesis. 30 Our observation of reduced expression from the associated haplotype suggests that this downregulation of a likely transcriptional repressor may relax the control of expression of downstream loci, facilitating neuroplastic changes in response to ethanol exposure.…”

Section: Discussionmentioning

confidence: 83%

Alcohol dependence is associated with the ZNF699 gene, a human locus related to Drosophila hangover, in the Irish affected sib pair study of alcohol dependence (IASPSAD) sample

et al. 2006

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Because tolerance is an important aspect of alcohol dependence (AD) in humans, recent evidence showing that the Drosophila gene hang is critically involved in the development of alcohol tolerance in the fly suggests that variation in related human loci might be important in the etiology of alcohol-related disorders. The orthology of hang in mammals is complex, but a number of human gene products (including ZNF699) with similar levels of amino-acid identity (18-26%) and similarity (30-41%), are consistently identified as the best matches with the translated hang sequence. We tested for association between the dichotomous clinical phenotype of alcohol dependence and seven single nucleotide polymorphisms (SNPs) in ZNF699 in our sample of 565 genetically independent cases and 496 siblings diagnosed with AD, and 609 controls. In analyses of genetically independent cases and controls, four of the seven single markers show strong evidence for association with AD (0.00003 < Fisher's exact P < 0.001), and the most significant single marker, rs7254880, tags an associated haplotype with frequency 0.071 in cases compared to 0.034 in controls (v 2 15.563, P < 0.00008, 5000 permutation P < 0.001, OR 2.17); inclusion of affected siblings gives similar results. Expression analyses conducted in independent postmortem brain samples show that expression of ZNF699 mRNA is significantly reduced in the dorsolateral prefrontal cortex of individuals carrying this haplotype compared with other observed haplotype combinations.

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“…The inebriometer has been used most commonly in the past and is the best suited to screening large numbers of mutations. However, it can only measure the knockdown phase of intoxication (Leibovitch et al, 1995;Moore et al, 1998;Singh and Heberlein, 2000;Berger et al, 2004). As has been demonstrated with other assays, flies become hyperactive when exposed to ethanol before becoming sedated (Moore et al, 1998).…”

Section: Introductionmentioning

confidence: 99%

“…In its first published study, one of the trial runs showed a significant row effect (Parr et al, 2001). Direct visual observation of the negative geotactic response and postural control has been used by multiple labs, including ours (Berger et al, 2004;Ghezzi et al, 2004;Wen et al, 2005;Cowmeadow et al, 2005). While this is certainly a thorough way to quantify sedation, it is also labor-intensive and therefore not well suited to the large volume of measurements inherent in a genetic screen.…”

Section: Introductionmentioning

confidence: 99%

Computer automated movement detection for the analysis of behavior

Ramazani¹,

Krishnan²,

Bergeson³

et al. 2007

Journal of Neuroscience Methods

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Currently, measuring ethanol behaviors in flies depends on expensive image analysis software or time intensive experimenter observation. We have designed an automated system for the collection and analysis of locomotor behavior data, using the IEEE 1394 acquisition program dvgrab, the image toolkit ImageMagick and the programming language Perl. In the proposed method, flies are placed in a clear container and a computer-controlled camera takes pictures at regular intervals. Digital subtraction removes the background and non-moving flies, leaving white pixels where movement has occurred. These pixels are tallied, giving a value that corresponds to the number of animals that have moved between images. Perl scripts automate these processes, allowing compatibility with highthroughput genetic screens. Four experiments demonstrate the utility of this method, the first showing heat-induced locomotor changes, the second showing tolerance to ethanol in a climbing assay, the third showing tolerance to ethanol by scoring the recovery of individual flies, and the fourth showing a mouse's preference for a novel object. Our lab will use this method to conduct a genetic screen for ethanol induced hyperactivity and sedation, however, it could also be used to analyze locomotor behavior of any organism.

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Rapid and Chronic: Two Distinct Forms of Ethanol Tolerance in Drosophila

Cited by 81 publications

References 37 publications

The FlyBar: Administering Alcohol to Flies

The FlyBar: Administering Alcohol to Flies

Alcohol dependence is associated with the ZNF699 gene, a human locus related to Drosophila hangover, in the Irish affected sib pair study of alcohol dependence (IASPSAD) sample

Computer automated movement detection for the analysis of behavior

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