Quantification of food intake in Drosophila - PubMed (original) (raw)
Quantification of food intake in Drosophila
Richard Wong et al. PLoS One. 2009.
Abstract
Measurement of food intake in the fruit fly Drosophila melanogaster is often necessary for studies of behaviour, nutrition and drug administration. There is no reliable and agreed method for measuring food intake of flies in undisturbed, steady state, and normal culture conditions. We report such a method, based on measurement of feeding frequency by proboscis-extension, validated by short-term measurements of food dye intake. We used the method to demonstrate that (a) female flies feed more frequently than males, (b) flies feed more often when housed in larger groups and (c) fly feeding varies at different times of the day. We also show that alterations in food intake are not induced by dietary restriction or by a null mutation of the fly insulin receptor substrate chico. In contrast, mutation of takeout increases food intake by increasing feeding frequency while mutation of ovo(D) increases food intake by increasing the volume of food consumed per proboscis-extension. This approach provides a practical and reliable method for quantification of food intake in Drosophila under normal, undisturbed culture conditions.
Conflict of interest statement
Competing Interests: The authors have declared that no competing interests exist.
Figures
Figure 1. Measurements of blue label uptake after 30 minutes of feeding and the proportion of feeding events observed during this period, where one circle represents one vial containing 5 flies.
Trend lines represent the relationship between the volume of food ingested and the observed proportion of flies feeding (V/O) described in Table 1. Dashed lines represent open circles. All flies were female unless stated, were 7 days old and were allowed to mate for 48 hours after eclosion (NF = the number of flies per condition, NV = the number of vials per condition). (a) A linear (V/O) relationship existed in mated Dahomey females (NF = 210, NV = 42). (b) The V/O relationships of mated Dahomey males and females did not differ significantly, although females were found to have fed at a greater frequency than males during the 30 minutes (NF = 200, NV = 40). The gradient for males did not differ significantly from that for females but had a lower intercept. (c) DR fed and full fed Dahomey females shared the same V/O relationship and no difference in feeding between dietary conditions was found with the combined assay (NF = 75, NV = 15). (d) The V/O relationship was the same in chico 1 heterozygotes and in the Dahomey control. No difference in feeding between genotypes was found with the combined assay (NF = 90, NV = 18). (e) The V/O relationship was the same in takeout 1 and in Canton-S females, even though takeout 1 flies were found to feed at a higher frequency than Canton-S controls (NF = 60, NV = 12). (f) Both ovo D1 and white Dahomey females had a positive V/O relationship, but ovo D1 flies had a significantly greater gradient and intercept, and therefore increased the volume of food ingested per proboscis-extension more quickly than white Dahomey females (NF = 200, NV = 40).
Figure 2. The relationship between blue label uptake and observed feeding events did not change for flies of advancing age.
Circles represent measurements of blue label uptake after 30 minutes of feeding and the proportion of feeding events observed during this period. One circle represents one vial containing 5 flies. Experiments were conducted with mated Dahomey females. Assays occurred at 4 different ages: on days 7, 21, 35 and 50 after eclosion. Each assay used 60 flies (12 vials) that were taken from a population that began with 500 individuals. Solid lines represent the significant (P<0.0001) V/O relationship with a gradient coefficient of 160.36 (S.E. = 31.39) and intercept of 2.89 (S.E. = 3.45), dashed lines represent the line of best fit for each age class.
Figure 3. Possible factors that influence feeding frequency.
(a) The proportion of time spent feeding of 7-day old mated females over a 2-hour period at varying times after lights-on. Females were housed alone, or in groups of 2, 5 or 10 (the number of flies for each condition = 30, with 30 vials for single flies, 15 vials for groups of 2, 6 vials for groups of 5 and 3 vials for groups of 10). We found that increasing the number of flies per vial increased the feeding frequency of each fly, and overall, flies fed more frequently in the afternoon and evening. We calculated the proportion of time spent feeding by summing the scored feeding events divided by the total number of feeding opportunities, which is unaffected by the difference in the number of vials per condition (b) The proportion of time spent feeding for flies fed different yeast-based diets. Flies were fed two similar diets containing either a water-soluble yeast extract (CSYExtract) or lyophilised yeast (SYBrewer's) at two different concentrations (DR = Dietary Restriction, FF = Full Fed). While feeding frequency was sensitive to the concentration of yeast extract in the diet, it was unchanged by the concentration of lyophilised yeast (NF = 60 and NV = 12 per condition: ** = P<0.005, and error bars = S.E.).
Figure 4. The proportion of time spent feeding for DR (open circles) and full fed (FF) flies (closed circles) on different days of their lifespan.
Survivorship curves are indicated with a solid grey line (DR) and a solid black line (FF) flies. Median lifespan: DR = 70 days, FF = 65 days. Proboscis-extension assays used 150 flies (30 vials) per condition. Flies were maintained in populations that began with 1500 individuals per condition (error bars = S.D.).
Figure 5. The proportion of time spent feeding during a proboscis-extension assay for DR (open circle) and fully fed (closed circle) once-mated 14-day old females.
Flies were maintained on different diets throughout their lifespan. DR females did not differ from fully fed females in feeding frequency. The assay began immediately when the observer arrived. Note the lower proportion of flies feeding during the first 30 minutes of the assay, which may relate to the appearance of the observer in the room (NF = 100; NV = 20).
Figure 6. The observed proportion of time spent feeding for Dahomey (control) flies (closed circles) and chico 1 heterozygotes (open circles) on different days of their lifespan, obtained by dividing the number of flies observed feeding by the total number of flies present.
Two observers alternately performed assays on the same population of flies. No significant interaction (P = 0.151) between the observers' data was found. Assays used 50 flies (10 vials) per condition, flies were maintained in populations that began with 500 individuals per condition; error bars = S.D.
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