Associative learning in larval and adult Drosophila is impaired by the dopamine-synthesis inhibitor 3-Iodo-L-tyrosine.

Across the animal kingdom, dopamine plays a crucial role in conferring reinforcement signals that teach animals about the causal structure of the world. In the fruit fly Drosophila melanogaster, dopaminergic reinforcement has largely been studied using genetics, whereas pharmacological approaches have received less attention. Here, we apply the dopamine-synthesis inhibitor 3-Iodo-L-tyrosine (3IY), which causes acute systemic inhibition of dopamine signaling, and investigate its effects on Pavlovian conditioning. We find that 3IY feeding impairs sugar-reward learning in larvae while leaving task-relevant behavioral faculties intact, and that additional feeding of a precursor of dopamine (L-3,4-dihydroxyphenylalanine, L-DOPA), rescues this impairment. Concerning a different developmental stage and for the aversive valence domain. Moreover, we demonstrate that punishment learning by activating the dopaminergic neuron PPL1-γ1pedc in adult flies is also impaired by 3IY feeding, and can likewise be rescued by L-DOPA. Our findings exemplify the advantages of using a pharmacological approach in combination with the genetic techniques available in D. melanogaster to manipulate neuronal and behavioral function.

Muscle-derived Dpp regulates feeding initiation via endocrine modulation of brain dopamine biosynthesis.

In animals, the brain regulates feeding behavior in response to local energy demands of peripheral tissues, which secrete orexigenic and anorexigenic hormones. Although skeletal muscle is a key peripheral tissue, it remains unknown whether muscle-secreted hormones regulate feeding. In Drosophila, we found that decapentaplegic (dpp), the homolog of human bone morphogenetic proteins BMP2 and BMP4, is a muscle-secreted factor (a myokine) that is induced by nutrient sensing and that circulates and signals to the brain. Muscle-restricted dpp RNAi promotes foraging and feeding initiation, whereas dpp overexpression reduces it. This regulation of feeding by muscle-derived Dpp stems from modulation of brain tyrosine hydroxylase (TH) expression and dopamine biosynthesis. Consistently, Dpp receptor signaling in dopaminergic neurons regulates TH expression and feeding initiation via the downstream transcriptional repressor Schnurri. Moreover, pharmacologic modulation of TH activity rescues the changes in feeding initiation due to modulation of dpp expression in muscle. These findings indicate that muscle-to-brain endocrine signaling mediated by the myokine Dpp regulates feeding behavior.

Electrochemical Measurements of Acetylcholine-Stimulated Dopamine Release in Adult Drosophila melanogaster Brains.

The fruit fly, Drosophila melanogaster, is a popular model organism for studying neurological processes and diseases due to the availability of sophisticated genetic tools. While endogenous neurotransmitter release has been characterized in Drosophila larvae, here, we measured endogenous dopamine release in isolated adult Drosophila brains for the first time. Dopamine was measured with fast-scan cyclic voltammetry (FSCV), and acetylcholine or nicotine were used as the stimulus, as both interact with nicotinic acetylcholine receptors (nAChRs) to evoke endogenous dopamine release. Stimulations with 10 pmol of acetylcholine elicited 0.26 ± 0.05 µM dopamine, while 70 fmol nicotine stimulations evoked 0.29 ± 0.03 µM in the central complex. Nicotine-stimulated dopamine release lasted much longer than acetylcholine-stimulated release. Dopamine release is reduced in the presence of nAChR antagonist α-bungarotoxin and the sodium channel blocker tetrodotoxin, indicating release is mediated by nAChRs and exocytosis. The identity of dopamine was confirmed by using 3-iodotyrosine, a dopamine synthesis inhibitor, and by confirming that release was not changed in octopamine synthesis mutant flies, Tdc2 RO54. Additionally, the half-decay time ( t50) in fumin (67 ± 15 s), dopamine transporter mutant flies, was larger than in wild-type flies (16 ± 3.7 s) further proving that acetylcholine stimulation evokes dopamine release. This study demonstrates that stimulation of nAChRs can be used to elicit endogenous dopamine release in adult fly brains, which will be a useful technique for future studies probing dopamine changes during aging or in neurodegenerative diseases.

The NMDA Receptor Promotes Sleep in the Fruit Fly, Drosophila melanogaster.

Considerable evidence indicates that sleep is essential for learning and memory. Drosophila melanogaster has emerged as a novel model for studying sleep. We previously found a short sleeper mutant, fumin (fmn), and identified its mutation in the dopamine transporter gene. We reported similarities in the molecular basis of sleep and arousal regulation between mammals and Drosophila. In aversive olfactory learning tasks, fmn mutants demonstrate defective memory retention, which suggests an association between sleep and memory. In an attempt to discover additional sleep related genes in Drosophila, we carried out a microarray analysis comparing mRNA expression in heads of fmn and control flies and found that 563 genes are differentially expressed. Next, using the pan-neuronal Gal4 driver elav-Gal4 and UAS-RNA interference (RNAi) to knockdown individual genes, we performed a functional screen. We found that knockdown of the NMDA type glutamate receptor channel gene (Nmdar1) (also known as dNR1) reduced sleep. The NMDA receptor (NMDAR) plays an important role in learning and memory both in Drosophila and mammals. The application of the NMDAR antagonist, MK-801, reduced sleep in control flies, but not in fmn. These results suggest that NMDAR promotes sleep regulation in Drosophila.

Dopamine modulates metabolic rate and temperature sensitivity in Drosophila melanogaster.

Homeothermal animals, such as mammals, maintain their body temperature by heat generation and heat dissipation, while poikilothermal animals, such as insects, accomplish it by relocating to an environment of their favored temperature. Catecholamines are known to regulate thermogenesis and metabolic rate in mammals, but their roles in other animals are poorly understood. The fruit fly, Drosophila melanogaster, has been used as a model system for the genetic studies of temperature preference behavior. Here, we demonstrate that metabolic rate and temperature sensitivity of some temperature sensitive behaviors are regulated by dopamine in Drosophila. Temperature-sensitive molecules like dTrpA1 and shi(ts) induce temperature-dependent behavioral changes, and the temperature at which the changes are induced were lowered in the dopamine transporter-defective mutant, fumin. The mutant also displays a preference for lower temperatures. This thermophobic phenotype was rescued by the genetic recovery of the dopamine transporter in dopamine neurons. Flies fed with a dopamine biosynthesis inhibitor (3-iodo-L-tyrosine), which diminishes dopamine signaling, exhibited preference for a higher temperature. Furthermore, we found that the metabolic rate is up-regulated in the fumin mutant. Taken together, dopamine has functions in the temperature sensitivity of behavioral changes and metabolic rate regulation in Drosophila, as well as its previously reported functions in arousal/sleep regulation.

Requirement of a dopaminergic neuronal phenotype for toxicity of low concentrations of 1-methyl-4-phenylpyridinium to human cells.

LUHMES cells are conditionally-immortalized non-transformed human fetal cells that can be differentiated to acquire a dopaminergic neuron-like phenotype under appropriate growth conditions. After differentiation by GDNF and cyclic adenosine monophosphate, LUHMES were sensitive to 1-methyl-4-phenylpyridinium (MPP(+)) toxicity at < or =5 microM, but resistant to the parental compound 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The high homogeneity and purity of the cultures allowed the detection of metabolic changes during the degeneration. Cellular ATP dropped in two phases after 24 and 48 h; cellular glutathione (GSH) decreased continuously, paralleled by an increase in lipid peroxidation. These events were accompanied by a time-dependent degeneration of neurites. Block of the dopamine transporter by GBR 12909 or mazindol completely abrogated MPP(+) toxicity. Inhibition of de novo dopamine synthesis by alpha-methyl-l-tyrosine or 3-iodo-l-tyrosine attenuated toxicity, but did not reduce the initial drop in ATP. Inhibition of mixed lineage kinases by CEP1347 completely prevented the MPP(+)-induced loss of viability and intracellular GSH, but failed to attenuate the initial drop of ATP. For the quantitative assessment of neurite degeneration, an automated imaging-based high content screening approach was applied and confirmed the findings made by pharmacological interventions in this study. Our data indicate that inhibition of mitochondrial ATP synthesis is not sufficient to trigger cell death in MPP(+)-treated LUHMES.

Interaction of dopamine, female pheromones, locomotion and sex behavior in Drosophila melanogaster.

The regulation of female hydrocarbons and courtship behavior by dopamine and their relationship with locomotion, were investigated in Drosophila melanogaster. Ddc mutants and wild-type female flies treated with tyrosine hydroxylase inhibitors (alpha-methyltyrosine or 3-iodotyrosine) had fewer diene hydrocarbons (female pheromones) and there was a total (Ddc), partial (alpha-methyltyrosine) or no (3-iodotyrosine) rescue of hydrocarbon pattern after dopamine ingestion. There was a correlation between female pheromone level and male courtship intensity for these dopamine-depleted or rescued flies. Female locomotion was decreased in flies treated with tyrosine hydroxylase inhibitors and restored by dopamine, showing that decreased mobility of the female has little importance on male courtship. However, male courtship was inhibited by an increased mobility of dopamine-supplemented females. Tanning, which is altered in dopamine-deficient flies and in tan and ebony mutants, seemed to have no significant influence on female pheromones. Females with increased quantities of dopamine (by ingestion) exhibited larger quantities of pheromones. However, Catsup mutants did not, probably as a result of defects in the epidermis. The Dat mutation, which resulted in more dopamine being produced in the brain, showed no pheromone modification. Together, these data show a complex interaction between dopamine, female hydrocarbons, locomotion and male courtship behavior.

Modifying effects of iodine on the immunogenicity of thyroglobulin peptides.

We have previously shown that iodotyrosyl formation within thyroglobulin (Tg) generates neoantigenic determinants that are immunopathogenic. In the current study, we have examined iodination effects on three tyrosyl-containing Tg peptides that are immunogenic in their non-iodinated form. We found that iodotyrosyl formation can enhance (p179, a.a. 179-194), suppress (p2540, a.a. 2540-2554), or not alter (p2529, a.a. 2529-2545) the immunogenic profiles of these peptides at the T-cell level. On the other hand, iodination did not alter the MHC-restriction profile of p2529 and p2540 (A(k)-binders) or p179 (A(k)- and E(k)-binder) and did not significantly influence the pathogenicity of these determinants. At the B-cell level, addition of an iodine atom on Y192 in p179 generated a neoantigenic determinant, but analogous effects were not discernible in p2529 or p2540. Our results demonstrate that iodotyrosyl formation can exert variable effects on the immunogenic behavior of Tg epitopes which may not always result in enhanced pathology. These findings also suggest that variations in the iodine content of Tg may significantly alter the hierarchy of antigenic determinants, to which the immune system may or may not be tolerant.

Protein radical formation on thyroid peroxidase during turnover as detected by immuno-spin trapping.

Thyroid peroxidase (TPO) is a 933 amino acid residue, heme-containing, integral membrane glycoprotein that catalyzes two steps in the maturation of the thyroid hormone precursor. As with other peroxidases, these reactions require hydrogen peroxide and initial enzyme oxidation. Previous researchers studied the oxidative state of the TPO heme moiety using spectrophotometric and catalytic analyses. We use a novel antiserum to 5,5-dimethyl-1-pyrroline N-oxide (DMPO) to detect radical-derived DMPO spin-trapped TPO. Our work reveals that TPO generates radical adducts in the presence of H2O2, but that the generation of these adducts can be suppressed by the addition of substrates and inhibitors. Chemical alteration of the tyrosine residues of TPO greatly reduces the generation of TPO-DMPO adducts. Iodide strongly suppresses the H2O2-generated production of TPO radical adducts and protects the enzyme from loss of enzyme activity. Because the normal catalytic mechanism of TPO involves the production of radical species, TPO is potentially more susceptible to oxidative damage than most enzymes which do not require H2O2 as a substrate. We hypothesize that oxidatively damaged TPO may trigger the production of anti-TPO autoantibodies, resulting in the development of autoimmune thyroid disorders. Evidence that correlates iodine deficiencies with development of thyroid autoimmune disorders supports this conjecture.

Effects of dopamine on juvenile hormone metabolism and fitness in Drosophila virilis.

The effects of dopamine (DA) on juvenile hormone (JH) metabolism and fitness (estimated as fecundity and viability levels under heat stress (38 degrees C)) in Drosophila virilis have been studied. An increase of DA level obtained by feeding with DA reduced fitness of wild-type (wt) flies under stress, and decreased JH degradation in young wt females while increasing it in sexually mature wt females. A decrease in DA levels resulted from 3-iodo-tyrosine treatment and caused a decrease in JH degradation in sexually mature wt and heat sensitive (hs) mutant females (DA level in hs females is twice as high in wt females). A dramatic decrease in viability under stress and fecundity under normal conditions in wt, but not hs, females was observed. 3-iodo-tyrosine treatment also reduced the number of oocytes at stages 8-14, delayed oocyte transition to stage 10 and resulted in the accumulation of mature eggs in wt females. It delayed maturation of wt, but not hs, males as well, but did not affect their fertility. This advances our understanding of the regulation of JH metabolism by DA in Drosophila and suggests a crucial role for the basal DA level in fitness.

Loss of flight and associated neuronal rhythmicity in inositol 1,4,5-trisphosphate receptor mutants of Drosophila.

Coordinated flight in winged insects requires rhythmic activity of the underlying neural circuit. Here, we show that Drosophila mutants for the inositol 1,4,5-trisphosphate (InsP(3)) receptor gene (itpr) are flightless. Electrophysiological recordings from thoracic indirect flight muscles show increased spontaneous firing accompanied by a loss of rhythmic flight activity patterns normally generated in response to a gentle puff of air. In contrast, climbing speed, the jump response, and electrical properties of the giant fiber pathway are normal, indicating that general motor coordination and neuronal excitability are much less sensitive to itpr mutations. All mutant phenotypes are rescued by expression of an itpr(+) transgene in serotonin and dopamine neurons. Pharmacological and immunohistochemical experiments support the idea that the InsP(3) receptor functions to modulate flight specifically through serotonergic interneurons. InsP(3) receptor action appears to be important for normal development of the flight circuit and its central pattern generator.

Ontogenic and adult whole body distribution of aminopeptidase N in rat investigated by in vitro autoradiography.

Aminopeptidase N (APN), which is widely distributed in mammalian tissues, is able to cleave numerous regulatory peptides. The selective inhibitor of APN, [(125)I] RB129, has been used to study the distribution of this exopeptidase during rat prenatal development and adult life by in vitro whole-body autoradiography. In the central nervous system, APN shows a weak labeling compared to the major part of the non-nervous tissues in the embryo and in the adult. APN is progressively expressed in kidney, intestine, heart, lung, sensory organs, eye, and thymus. In organs such as the liver, the cartilages and the bones, altered levels of APN expression are observed during the development, or in the embryo compared to the adult, suggesting a role of APN during the liver haematopoiesis and bone growth. At this time, all the physiological functions of APN are still incompletely known, however its developmental pattern of expression strongly suggests a function of modulation of this enzyme during the development, next in physiological and/or pathological situations in adult. In this way, APN could represent a new therapeutic target in pathological processes, such as tumoral proliferation and/or angiogenesis associated with cancer development, where an increase in the level of this enzyme has been observed.

The effects of dopamine synthesis inhibitors and dopamine antagonists on regeneration in the hydra Hydra attenuata.

The effects of catecholamine synthesis inhibitors (alpha-methyltyrosine, 3-iodotyrosine, and alpha-methyl-DOPA) and dopamine receptor blockers (haloperidol and spiperone) on the regeneration of apical, gastral, and basal fragments of the hydra Hydra attenuata were studied. These experiments showed that alpha-methyltyrosine and 3-iodotyrosine significantly inhibited regeneration but did not produce morphological anomalies. Alpha-Methyl-DOPA produce less inhibition of regeneration, but induced ectopic tentacles and outgrowths in gastral regenerates. Haloperidol and spiperone had no significant effect on the rate of regeneration but induced significant numbers of morphogenetic anomalies in gastral regenerates. Apical and basal regenerates, which retained their natural organizers (the head and base respectively) never yielded morphogenetic anomalies in the presence of either dopamine receptor blockers or dopamine synthesis inhibitors. The possible role of neurotransmitters. particularly dopamine, in morphogenesis in hydras is discussed.

[The action of catecholamine-synthesis inhibitors and of spiperone on sea urchin and mouse embryos]. / Deistvie ingibitorov sinteza katekholaminov i spiperona na zarodyshi morskikh ezhei i myshei.

We studied the effects of three inhibitors of catecholamine synthesis on the development of sea urchins Sphaerechinus granularis and Paracentrotus lividus. These drugs affected the early embryogenesis, which was expressed in inhibition of the cleavage divisions, appearance of abnormal embryos, and developmental arrest. The addition of arachidonic acid amide and dopamine to the incubation medium weakened the effects of the inhibitors. Spiperone induced developmental defects in preimplantation mouse embryos and sea urchin embryos. Arachidonic acid amide with dopamine exerted a protective effect against spiperone when introduced to sea urchin embryos at the blastula or late gastrula stages, rather than after fertilization. In murine embryos, this amide induced developmental defects and arrest itself and its effect was reversible. Possible mechanisms underlying the effects of these drugs are discussed.

Binding properties of a highly potent and selective iodinated aminopeptidase N inhibitor appropriate for radioautography.

Aminopeptidase N (APN) is a zinc metallopeptidase involved in the inactivation of biologically active peptides. The knowledge of its precise distribution is crucial to investigate its physiological role. This requires the use of appropriate probes such as the recently developed highly potent and selective radiolabeled APN inhibitor 2(S)-benzyl-3-[hydroxy(1'(R)-aminoethyl)phosphinyl]propanoyl-L-3-[ (12 5)I]iodotyrosine ([(125)I]RB 129). Its binding properties were investigated using rat brain homogenates (K(d)=3.4 nM) or APN expressed in COS-7 cells (K(d)=0.9 nM). The specific binding was 95% at [K(d)], and preliminary autoradiography in intestine is promising. The decreased affinity of [(125)I]RB 129 (=10(-6) M) for the E(350)D APN mutant, supports the critical role of E(350) in the amino-exopeptidase action of APN.

[Effect of inhibitors of synthesis of dopamine and its antagonists on Hydra attenuata regeneration]. / Vliianie ingibitorov sinteza i antagonistov dofamina na regeneratsiiu gidry Hydra attenuata.

Obvious inhibition of the hydra regeneration with no subsequent morphological abnormalities, was shown to occur when using alpha-methylthyrosine and 3 Jthyrosine. alpha-Methyldopa induced a slight inhibition but a considerable morphological change: ectopic tentacles, projections, bipolar forms in the gastric fragment. The apical and basal fragments did not suffer. The role of neurotransmitters in the hydra morphogenesis is discussed.

Dopamine and serotonin in the larval CNS of a drosophilid fly, Chymomyza costata: are they involved in the regulation of diapause?

Developmental profiles of dopamine (DA) and serotonin (5-hydroxytryptamine, 5-HT) in the larval CNS of Chymomyza costata were measured by HPLC using electrochemical detection. Larvae of two strains, wild-type (W) and nondiapause mutant (M), were maintained either under long-day (LD, inducing pupariation) or short-day (SD, inducing diapause in W-strain) photoperiods. The levels of DA ranged from 10 fmol/CNS (early 3rd instar larvae) to 60 fmol/CNS (150-day-old diapausing larvae); the range for 5-HT was from 10 fmol/CNS to 75 fmol/CNS in the same larvae. During the 3rd larval instar, which is the decisive stage for photoperiodic induction of diapause, no differences were found in DA developmental profiles between different strains or conditions. Some differences were found in 5-HT developmental profiles, but only after the end of sensitive stage, and were therefore regarded as insignificant for regulation of developmental mode. Similarly, no clear correlations between the developmental profiles of DA and 5-HT and the course of developmental changes during the maintenance and termination of a few-month-long larval diapause were observed. Furthermore, the DA and 5-HT levels in the CNS were pharmacologically manipulated by feeding the larvae with either precursors or enzyme inhibitors of DA and 5-HT biosynthesis. Although retardations of growth and development were observed, the treated larvae retained full capacity for the photoperiodic response, irrespective of the level of DA or 5-HT in their CNS. Larvae with their 5-HT depleted to trace levels survived and were capable of diapause induction, maintenance, and termination. Depletion of DA to trace levels resulted in 100% mortality. Collectively, the present study indicates that 5-HT in the CNS is dispensable for the photoperiodic response in C. costata. More information is needed to elucidate the potential role of DA.

Acute effects of thyroid hormone analogs on sodium currents in neonatal rat myocytes.

We previously reported that T3(3,3',5-triiodo-L-thyronine) acutely increases sodium currents (INa) in neonatal rat myocytes. Here we compare the effects of several thyroid hormone analogs, including T4(3,3',5,5'-tetraiodo-L-thyronine), rT3(3,3',5'-triiodo-L-thyronine), D-T3(3,3',5-triiodo-D-thyronine), 3,5-T2(3,5-diiodo-L-thyronine), DIT (3,5-diiodo-L-tyrosine), MIT (3-monoiodo-L-tyrosine), tetrac (3,3',5,5'-tetraiodo-thyroacetic acid), triac (3, 3',5-triiodo-thyroacetic acid), and tyrosine, on INa in cultured neonatal rat myocytes (n ranged from 9 to 28 for each comparison). T4, T3, 3,5-T2, and DIT (10 n m) all increased current density relative to control to a similar degree: to 1.22+/-0.2, 1.21+/-0.03, 1.16+/-0.02 and 1.16+/-0.03, respectively, P<0.05. In contrast, thyroid hormone analogs with an altered side group of the inner iodophenyl ring, including tetrac, triac, and D-T3, had no effect on INa nor did rT3, MIT or tyrosine. Pretreatment with rT3 inhibited the effects of T4, T3, 3,5-T2, and DIT. Conversely, the dose-dependent inhibitory effect of amiodarone, an iodinated benzofuran derivative that antagonizes thyroid hormone actions, on INa was blocked when myocytes were pretreated with T3(100 n m, n=3), suggesting an interaction of T3 with amiodarone. The enhancement of INa by T3 and 3, 5-T2 could not be blocked by propranolol, suggesting that the effects are not mediated through beta -adrenergic signaling pathways. In conclusion, the present results suggest that the acute effects of thyroid hormone and analogs on cardiac INa are mediated by a non-genomic thyroid hormone receptor with a unique structure-activity relationship.

Effects of iodotyrosines, thyronines, iodothyroacetic acids and thyromimetic analogues on in vitro copper-induced oxidation of low-density lipoproteins.

We studied the effect of different thyroid compounds [(I2, monoiodo-L-tyrosine (MIT), diiodo-L-tyrosine (DIT), L-thyronine (T0), 3,5-diiodo-L-thyronine (T2), 3,5,3'-triiodo-L-thyronine (T3), 3,3',5'-triiodo-L-thyronine (rT3), 3,5,3',5'-tetraiodo-L-thyronine (T4), 3,5-diiodothyroacetic acid (TA2), 3,5,3'-triiodothyroacetic acid (TA3) and 3,5,3',5'-tetraiodothyroacetic acid (TA4)] or thyromimetics [(3,5-dimethyl-3'-isopropyl-L-thyronine (DIMIT) and 3,5-diiodo-3'-isopropyl-thyroacetic acid (IpTA2)] on in vitro copper-induced oxidation of low-density lipoproteins (LDL). Human native LDL (0.05 g protein/L) oxidation was induced by 2.5 micromol/L of CuCl2. Conjugated dienes were measured spectrophotometrically for up to 10 hr. The length of the lag phase (Tlag), maximum velocity of the reaction (Vmax) and the maximum amount of generated dienes were obtained from kinetic data. T3 increased Tlag and decreased Vmax with a dependence upon concentration (0 to 3 micromol/L). There was no difference between the Dmax obtained with Cu2+ alone or in the presence of the various compounds (1 micromol/L). I2, MIT and DIT did not modify any parameter of the oxidation kinetic. T0 and T2 had the same antioxidant efficiency as T3, whereas T4 only decreased Vmax. rT3 increased Tlag less than did T3, whereas DIMIT was the thyronine that had the most important effect. TA2 and TA, were the most efficient antioxidant compounds. TA4 decreased Tlag less than TA3 did, whereas IpTA2 had an effect weaker than that of the physiological acetic derivatives. The data suggest that thyroid hormones and derivatives have LDL-antioxidant properties, their importance being related to their 4'-hydroxy diphenyl ether structure and depending upon the nature and the position of substituents in this structure.

Dopamine and mushroom bodies in Drosophila: experience-dependent and -independent aspects of sexual behavior.

Depletion of dopamine in Drosophila melanogaster adult males, accomplished through systemic introduction of the tyrosine hydroxylase inhibitor 3-iodo-tyrosine, severely impaired the ability of these flies to modify their courtship responses to immature males. Mature males, when first exposed to immature males, will perform courtship rituals; the intensity and duration of this behavior rapidly diminishes with time. Dopamine is also required for normal female sexual receptivity; dopamine-depleted females show increased latency to copulation. One kilobase of 5' upstream information from the Drosophila tyrosine hydroxylase (DTH) gene, when fused to the Escherichia coli beta-galactosidase reporter and transduced into the genome of Drosophila melanogaster, is capable of directing expression of the reporter gene in the mushroom bodies, which are believed to mediate learning acquisition and memory retention in flies. Ablation of mushroom bodies by treatment of newly hatched larva with hydroxyurea resulted in the inability of treated mature adult males to cease courtship when placed with untreated immature males. However, functional mushroom bodies were not required for the dopaminergic modulation of an innate behavior, female sexual receptivity. These data suggest that dopamine acts as a signaling molecule within the mushroom bodies to mediate a simple form of learning.