Article The Leucokinin Pathway and Its Neurons Regulate Meal Size in Drosophila

Department of Molecular, Cellular and DevelopmentalBiology, Yale University, KBT 610, PO Box 208103,New Haven, CT 06511, USASummaryBackground: Total food intake is a function of meal size andmeal frequency, and adjustments to these parameters allowanimals to maintain a stable energy balance in changingenvironmental conditions. The physiological mechanisms thatregulate meal size have been studied in blowflies but have notbeen previously examined in Drosophila.Results:Here we show that mutations in the leucokininneuro-peptide (leuc) and leucokinin receptor (lkr) genes causephenotypes in which Drosophila adults have an increase inmeal size and a compensatory reduction in meal frequency.Because mutant flies take larger but fewer meals, their caloricintake is the same as that of wild-type flies. The expressionpatternsoftheleucandlkrgenesidentifysmallgroupsofbrainneurons that regulate this behavior. Leuc-containing presyn-aptic terminals are found close to Lkr neurons in the brainand ventral ganglia, suggesting that they deliver Leuc peptideto these neurons. Lkr neurons innervate the foregut. Flies inwhich Leuc or Lkr neurons are ablated have defects identicalto those of leucokinin pathway mutants.Conclusions: Our data suggest that the increase in meal sizein leuc and lkr mutants is due to a meal termination defect,perhaps arising from impaired communication of gut disten-sion signals to the brain. Leucokinin and the leucokininreceptor are homologous to vertebrate tachykinin and itsreceptor, and injection of tachykinins reduces food consump-tion.Ourresultssuggestthattherolesofthetachykininsystemin regulating food intake might be evolutionarily conservedbetween insects and vertebrates.IntroductionIn mammals, nutrient intake is regulated to keep body weightconstant over long periods of time. Most animals consumefood in discrete bouts called meals, and total food intake isa function of both meal size and meal frequency. Identificationofthepathwaysthatregulatethesemeal-relatedparametersisessential for the understanding of the relationships betweenbody weight regulation and caloric intake [1, 2].Signals that control meal size and frequency fall into threecategories: those that initiate a meal, those that maintainfeeding once a meal has begun, and those that terminatea meal. In hungry mammals, the smell and taste of food initiatefeeding. As feeding continues, the level of gastric distension isconveyed to the brain via stomach wall stretch receptors.When the extent of stomach distension passes a threshold,the meal is likely to terminate [3–5]. Also, during the courseof a meal, some nutrients are absorbed in the small intestine,allowing a postgastric evaluation of the caloric content ofingested food that can also contribute to meal termination [6].The steps involved in physiological regulation of feedingbehavior in flies have been elucidated primarily throughstudies on the blowfly Phormia regina [7, 8]. As the hungryfly walks, taste hairs on its legs sample the surface. Whena food source is detected, the fly extends its proboscis andbegins to feed. During ingestion, liquid food passes throughthe foregut into a collapsible food-storage sac called thecrop. Eventually, the fly becomes satiated and stops feeding.A number of factors contribute to termination of a feedingbout and thus determine meal size. First, stretch receptorsmonitoring gut distension provide a negative feedback signalto the brain. Second, neurons in the taste hairs habituate andbecome less responsive to food [7, 8].Leucokinin(Leuc)isamyotropicneuropeptidefoundinmostinvertebratespecies[9].Itwasinitiallyidentifiedasaneurohor-mone that increases Malpighian tubule fluid secretion andhindgut motility in some insect species [10–14]. The biologicalactivity of leucokinin requires an amidated C-terminal penta-peptide motif called FXXWG-amide, a feature that it shareswith the related vertebrate tachykinin neuropeptides. Thetachykinin family includes substance P, substance K/neuroki-nin A, and neuropeptide K /neurokinin B [14, 15]. Althoughthe Drosophila genome encodes another peptide whosesequence is somewhat closer to vertebrate tachykinins thanis leucokinin’s [16], the observation that the Drosophila leuco-kinin receptor, Lkr, is homologous to vertebrate tachykininreceptors confirms the homology between the leucokininand tachykinin pathways [17].Here, we report that the leucokinin pathway is involved inmeal size regulation in Drosophila. Flies with reduced leucoki-nin pathway signaling as a result of mutations in the genesencoding either the leucokinin neuropeptide (leuc) or theleucokinin receptor (lkr) have an abnormal increase in mealsize. This increase is associated with a reduction in mealfrequency that causes mutant flies to consume the same totalamount of food as wild-type flies. The functions of the leucoki-ninpathwayinregulationofmealsizeareexecutedinneurons,because pan-neuronal expression of leuc or lkr rescues thephenotypes. leuc and lkr are expressed in distinct patternsof neurons, and ablation of these neurons phenocopies theeffects of the leuc and lkr mutations.Resultsleuc and lkr Mutant Flies Eat Excessively after StarvationTo obtain insights into the molecular mechanisms involved incontrol of meal size, we performed a screen for mutationsthat cause adults of thegenetically tractable insect Drosophilamelanogaster to consume abnormally large amounts of food.A number of different assays have been used to monitorfood consumption in Drosophila [18, 19]. For our screen, we