RESUMEN
The neurohormone octopamine regulates many crucial physiological processes in insects and exerts its activity via typical G-protein coupled receptors. The roles of octopamine receptors in regulating behavior and physiology in Coleoptera (beetles) need better understanding. We used the red flour beetle, Tribolium castaneum, as a model species to study the contribution of the octopamine receptor to behavior and physiology. We cloned the cDNA of a ß-adrenergic-like octopamine receptor (TcOctß2R). This was heterologously expressed in human embryonic kidney (HEK) 293 cells and was demonstrated to be functional using an in vitro cyclic AMP assay. In an RNAi assay, injection of dsRNA demonstrated that TcOctß2R modulates beetle locomotion, mating duration, and fertility. These data present some roles of the octopaminergic signaling system in T. castaneum. Our findings will also help to elucidate the potential functions of individual octopamine receptors in other insects.
Asunto(s)
Locomoción/genética , Octopamina/genética , Receptores de Amina Biogénica/genética , Reproducción/genética , Tribolium/genética , Adrenérgicos/metabolismo , Secuencia de Aminoácidos , Animales , Línea Celular , Escarabajos/genética , AMP Cíclico/genética , Femenino , Células HEK293 , Humanos , Proteínas de Insectos/genética , Masculino , Interferencia de ARN/fisiología , ARN Bicatenario/genética , Receptores Acoplados a Proteínas G/genética , Alineación de SecuenciaRESUMEN
The biogenic amine octopamine (OA) and its precursor tyramine (TA) are involved in controlling a plethora of different physiological and behavioral processes. The tyramine-ß-hydroxylase (tßh) gene encodes the enzyme catalyzing the last synthesis step from TA to OA. Here, we report differential dominance (from recessive to overdominant) of the putative null tßhnM18 allele in 2 behavioral measures in Buridan's paradigm (walking speed and stripe deviation) and in proboscis extension (sugar sensitivity) in the fruit fly Drosophila melanogaster. The behavioral analysis of transgenic tßh expression experiments in mutant and wild-type flies as well as of OA and TA receptor mutants revealed a complex interaction of both aminergic systems. Our analysis suggests that the different neuronal networks responsible for the 3 phenotypes show differential sensitivity to tßh gene expression levels. The evidence suggests that this sensitivity is brought about by a TA/OA opponent system modulating the involved neuronal circuits. This conclusion has important implications for standard transgenic techniques commonly used in functional genetics.
Asunto(s)
Oxigenasas de Función Mixta/genética , Oxigenasas de Función Mixta/metabolismo , Alelos , Animales , Animales Modificados Genéticamente/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Femenino , Genotipo , Masculino , Mutación/genética , Octopamina/genética , Octopamina/metabolismo , Fenotipo , Receptores de Amina Biogénica/genética , Receptores de Amina Biogénica/metabolismo , Tiramina/metabolismoRESUMEN
Adrenergic signaling profoundly modulates animal behavior. For example, the invertebrate counterpart of norepinephrine, octopamine, and its biological precursor and functional antagonist, tyramine, adjust motor behavior to different nutritional states. In Drosophila larvae, food deprivation increases locomotor speed via octopamine-mediated structural plasticity of neuromuscular synapses, whereas tyramine reduces locomotor speed, but the underlying cellular and molecular mechanisms remain unknown. We show that tyramine is released into the CNS to reduce motoneuron intrinsic excitability and responses to excitatory cholinergic input, both by tyraminehonoka receptor activation and by downstream decrease of L-type calcium current. This central effect of tyramine on motoneurons is required for the adaptive reduction of locomotor activity after feeding. Similarly, peripheral octopamine action on motoneurons has been reported to be required for increasing locomotion upon starvation. We further show that the level of tyramine-ß-hydroxylase (TBH), the enzyme that converts tyramine into octopamine in aminergic neurons, is increased by food deprivation, thus selecting between antagonistic amine actions on motoneurons. Therefore, octopamine and tyramine provide global but distinctly different mechanisms to regulate motoneuron excitability and behavioral plasticity, and their antagonistic actions are balanced within a dynamic range by nutritional effects on TBH.
Asunto(s)
Oxigenasas de Función Mixta/genética , Neuronas Motoras/metabolismo , Octopamina/genética , Receptores de Amina Biogénica/genética , Tiramina/metabolismo , Animales , Conducta Animal/fisiología , Canales de Calcio Tipo L/genética , Canales de Calcio Tipo L/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Drosophila melanogaster/fisiología , Privación de Alimentos/fisiología , Larva/metabolismo , Larva/fisiología , Locomoción/genética , Locomoción/fisiología , Oxigenasas de Función Mixta/metabolismo , Neuronas Motoras/fisiología , Estado Nutricional/genética , Estado Nutricional/fisiología , Octopamina/metabolismo , Receptores de Amina Biogénica/metabolismo , Sinapsis/metabolismo , Sinapsis/fisiologíaRESUMEN
G protein-coupled receptors (GPCRs) are established drug targets. Despite their considerable appeal as targets for next-generation anthelmintics, poor understanding of their diversity and function in parasitic helminths has thwarted progress towards GPCR-targeted anti-parasite drugs. This study facilitates GPCR research in the liver fluke, Fasciola hepatica, by generating the first profile of GPCRs from the F. hepatica genome. Our dataset describes 147 high confidence GPCRs, representing the largest cohort of GPCRs, and the largest set of in silico ligand-receptor predictions, yet reported in any parasitic helminth. All GPCRs fall within the established GRAFS nomenclature; comprising three glutamate, 135 rhodopsin, two adhesion, five frizzled, one smoothened, and one secretin GPCR. Stringent annotation pipelines identified 18 highly diverged rhodopsins in F. hepatica that maintained core rhodopsin signatures, but lacked significant similarity with non-flatworm sequences, providing a new sub-group of potential flukicide targets. These facilitated identification of a larger cohort of 76 related sequences from available flatworm genomes, representing new members of existing groups (PROF1/Srfb, Rho-L, Rho-R, Srfa, Srfc) of flatworm-specific rhodopsins. These receptors imply flatworm specific GPCR functions, and/or co-evolution with unique flatworm ligands, and could facilitate the development of exquisitely selective anthelmintics. Ligand binding domain sequence conservation relative to deorphanised rhodopsins enabled high confidence ligand-receptor matching of seventeen receptors activated by acetylcholine, neuropeptide F/Y, octopamine or serotonin. RNA-Seq analyses showed expression of 101 GPCRs across various developmental stages, with the majority expressed most highly in the pathogenic intra-mammalian juvenile parasites. These data identify a broad complement of GPCRs in F. hepatica, including rhodopsins likely to have key functions in neuromuscular control and sensory perception, as well as frizzled and adhesion/secretin families implicated, in other species, in growth, development and reproduction. This catalogue of liver fluke GPCRs provides a platform for new avenues into our understanding of flatworm biology and anthelmintic discovery.
Asunto(s)
Evolución Molecular , Fasciola hepatica/genética , Genoma de los Helmintos , Receptores Acoplados a Proteínas G/genética , Rodopsina/genética , Acetilcolina/genética , Animales , Humanos , Neuropéptidos/genética , Octopamina/genética , Filogenia , Platelmintos/clasificación , Platelmintos/genética , Rodopsina/aislamiento & purificación , Alineación de Secuencia , Análisis de Secuencia de ARN , Serotonina/genéticaRESUMEN
It is important to design insecticides having both low drug resistance and less undesirable toxicity for desert locust control. Specific GPCRs of Schistocerca gregaria, especially ß-adrenergic-like octopamine receptor (SgOctßR), can be considered as its potential effective insecticide targets. However, either the unavailability of SgOctßR's structure or the inadequate capability of its sequence lead the development of insecticide for Schistocerca gregaria meets its plateau. To relax this difficulty, this paper develops a promising progressive structure simulation from SgOctßR's sequence, to its predicted structure of SgOctßR in vacuum, to its conformation as well as its complex with endogenous ligand octopamine in a solvent-membrane system. The combined approach of multiple sequence alignment, static structural characterization, and dynamic process of conformational change during binding octopamine reveal three important aspects. The first one is the characterization of SgOctßR's active pocket, including the attending secondary structure elements, its hydrophobic residues and nonpolar surface. The second one is the interaction with octopamine, especially the involved hydrogen bonds and an aromatic stacking of pi-pi interactions. The third one is the potential binding sites, including six highly conserved residues and one highly variable residue for locust insecticide design. This work is definitely helpful for the further structure-based drug design for efficient and eco-friendly insecticides, as well as site-directed mutagenesis biochemical research of SgOctßR.
Asunto(s)
Adrenérgicos/química , Insecticidas/química , Octopamina/química , Receptores de Amina Biogénica/química , Animales , Sitios de Unión , Resistencia a Medicamentos/genética , Saltamontes/química , Ligandos , Mutagénesis , Octopamina/genética , Receptores de Amina Biogénica/genéticaRESUMEN
The monoamines octopamine (OA) and tyramine (TA) modulate numerous behaviours and physiological processes in invertebrates. Nevertheless, it is not clear whether these invertebrate counterparts of norepinephrine are important regulators of metabolic and life history traits. We show that flies (Drosophila melanogaster) lacking OA are more resistant to starvation, while their overall life span is substantially reduced compared with control flies. In addition, these animals have increased body fat deposits, reduced physical activity and a reduced metabolic resting rate. Increasing the release of OA from internal stores induced the opposite effects. Flies devoid of both OA and TA had normal body fat and metabolic rates, suggesting that OA and TA act antagonistically. Moreover, OA-deficient flies show increased insulin release rates. We inferred that the OA-mediated control of insulin release accounts for a substantial proportion of the alterations observed in these flies. Apparently, OA levels control the balance between thrifty and expenditure metabolic modes. Thus, changes in OA levels in response to external and internal signals orchestrate behaviour and metabolic processes to meet physiological needs. Moreover, chronic deregulation of the corresponding signalling systems in humans may be associated with metabolic disorders, such as obesity or diabetes.
Asunto(s)
Proteínas de Drosophila/genética , Oxigenasas de Función Mixta/genética , Octopamina/genética , Inanición/genética , Tiramina/metabolismo , Tirosina Descarboxilasa/genética , Animales , Conducta Animal/fisiología , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Redes y Vías Metabólicas/genética , Octopamina/deficiencia , Octopamina/metabolismo , FenotipoRESUMEN
The effect of strong hypomorphic mutation of the insulin-like protein gene (dilp6) on metabolism of octopamine (one of the main biogenic amines in insects) was studied in Drosophila melanogaster males and females. The activity of tyrosine decarboxylase (the key enzyme of octopamine synthesis) and the activity of octopamine-dependent N-acetyltransferase (the enzyme of its degradation) were measured. It was demonstrated that the activity of both studied enzymes is decreased under normal conditions in the dilp6 41 mutants (as we previously demonstrated, this is correlated with an increased level of octopamine). It was also found that hypomorphic mutation of the dilp6 gene decreases the intensity of tyrosine decarboxylase response to heat stress. Thus, it was demonstrated for the first time that insulin-like DILP6 protein in drosophila influences the level of octopamine (regulating the activity of the enzyme degrading octopamine).
Asunto(s)
Proteínas de Drosophila , Respuesta al Choque Térmico/fisiología , Octopamina , Somatomedinas , Animales , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster , Octopamina/genética , Octopamina/metabolismo , Somatomedinas/genética , Somatomedinas/metabolismoRESUMEN
Ovulation is essential for the propagation of the species and involves a proteolytic degradation of the follicle wall for the release of the fertilizable oocyte. However, the precise mechanisms for regulating these proteolytic events are largely unknown. Work from our lab and others have shown that there are several parallels between Drosophila and mammalian ovulation at both the cellular and molecular levels. During ovulation in Drosophila, posterior follicle cells surrounding a mature oocyte are selectively degraded and the residual follicle cells remain in the ovary to form a corpus luteum after follicle rupture. Like in mammals, this rupturing process also depends on matrix metalloproteinase 2 (Mmp2) activity localized at the posterior end of mature follicles, where oocytes exit. In the present study, we show that Mmp2 activity is regulated by the octopaminergic signaling in mature follicle cells. Exogenous octopamine (OA; equivalent to norepinephrine, NE) is sufficient to induce follicle rupture when isolated mature follicles are cultured ex vivo, in the absence of the oviduct or ovarian muscle sheath. Knocking down the alpha-like adrenergic receptor Oamb (Octoampine receptor in mushroom bodies) in mature follicle cells prevents OA-induced follicle rupture ex vivo and ovulation in vivo. We also show that follicular OA-Oamb signaling induces Mmp2 enzymatic activation but not Mmp2 protein expression, likely via intracellular Ca2+ as the second messenger. Our work develops a novel ex vivo follicle rupture assay and demonstrates the role for follicular adrenergic signaling in Mmp2 activation and ovulation in Drosophila, which is likely conserved in other species.
Asunto(s)
Proteínas de Drosophila/genética , Drosophila/crecimiento & desarrollo , Metaloproteinasa 2 de la Matriz/genética , Folículo Ovárico/crecimiento & desarrollo , Ovulación/genética , Receptores de Neurotransmisores/genética , Animales , Señalización del Calcio/genética , Cuerpo Lúteo/crecimiento & desarrollo , Cuerpo Lúteo/metabolismo , Drosophila/genética , Proteínas de Drosophila/biosíntesis , Femenino , Humanos , Metaloproteinasa 2 de la Matriz/biosíntesis , Octopamina/genética , Oocitos/crecimiento & desarrollo , Oocitos/metabolismo , Folículo Ovárico/metabolismo , Oviductos/metabolismo , Proteolisis , Receptores de Neurotransmisores/biosíntesis , Transducción de SeñalRESUMEN
Flexible behaviour allows organisms to respond appropriately to changing environmental and social conditions. In the subsocial beetle Nicrophorus vespilloides, females tolerate conspecifics when mating, become aggressive when defending resources, and return to social tolerance when transitioning to parenting. Given the association between octopamine and aggression in insects, we hypothesized that genes in the octopaminergic system would be differentially expressed across different social and reproductive contexts. To test this in N. vespilloides, we first obtained the sequences of orthologues of the synthetic enzymes and receptors of the octopaminergic system. We next compared relative gene expression from virgin females, mated females, mated females alone on a resource required for reproduction and mated females on a resource with a male. Expression varied for five receptor genes. The expression of octopamine ß receptor 1 and octopamine ß receptor 2 was relatively higher in mated females than in other social conditions. Octopamine ß receptor 3 was influenced by the presence or absence of a resource and less by social environment. Octopamine α receptor and octopamine/tyramine receptor 1 gene expression was relatively lower in the mated females with a resource and a male. We suggest that in N. vespilloides the octopaminergic system is associated with the expression of resource defence, alternative mating tactics, social tolerance and indirect parental care.
Asunto(s)
Escarabajos/genética , Escarabajos/fisiología , Agresión , Animales , Secuencia de Bases , Femenino , Expresión Génica/fisiología , Masculino , Datos de Secuencia Molecular , Octopamina/biosíntesis , Octopamina/genética , Reproducción , Conducta Sexual Animal , Conducta SocialRESUMEN
Recent evidence suggests that flies' sensitivity to large-field optic flow is increased by the release of octopamine during flight. This increase in gain presumably enhances visually mediated behaviors such as the active regulation of forward speed, a process that involves the comparison of a vision-based estimate of velocity with an internal set point. To determine where in the neural circuit this comparison is made, we selectively silenced the octopamine neurons in the fruit fly Drosophila, and examined the effect on vision-based velocity regulation in free-flying flies. We found that flies with inactivated octopamine neurons accelerated more slowly in response to visual motion than control flies, but maintained nearly the same baseline flight speed. Our results are parsimonious with a circuit architecture in which the internal control signal is injected into the visual motion pathway upstream of the interneuron network that estimates groundspeed.
Asunto(s)
Drosophila/fisiología , Vuelo Animal , Octopamina/metabolismo , Animales , Drosophila/genética , Expresión Génica , Silenciador del Gen , Percepción de Movimiento , Neuronas/metabolismo , Octopamina/genética , Flujo Optico , Visión OcularRESUMEN
After Drosophila males are rejected by mated females, their subsequent courtship is inhibited even when encountering virgin females. Molecular mechanisms underlying courtship conditioning in the CNS are unclear. In this study, we find that tyramine ß hydroxylase (TßH) mutant males unable to synthesize octopamine (OA) showed impaired courtship conditioning, which could be rescued by transgenic TßH expression in the CNS. Inactivation of octopaminergic neurons mimicked the TßH mutant phenotype. Transient activation of octopaminergic neurons in males not only decreased their courtship of virgin females, but also produced courtship conditioning. Single cell analysis revealed projection of octopaminergic neurons to the mushroom bodies. Deletion of the OAMB gene encoding an OA receptor expressed in the mushroom bodies disrupted courtship conditioning. Inactivation of neurons expressing OAMB also eliminated courtship conditioning. OAMB neurons responded robustly to male-specific pheromone cis-vaccenyl acetate in a dose-dependent manner. Our results indicate that OA plays an important role in courtship conditioning through its OAMB receptor expressed in a specific neuronal subset of the mushroom bodies.
Asunto(s)
Condicionamiento Psicológico , Cortejo , Proteínas de Drosophila/genética , Técnicas de Sustitución del Gen , Octopamina/genética , Receptores de Neurotransmisores/genética , Conducta Sexual Animal/fisiología , Animales , Condicionamiento Psicológico/fisiología , Proteínas de Drosophila/fisiología , Drosophila melanogaster , Femenino , Técnicas de Sustitución del Gen/métodos , Masculino , Octopamina/fisiología , Receptores de Neurotransmisores/fisiologíaRESUMEN
Sleep length and metabolic dysfunction are correlated, but the causal relationship between these processes is unclear. Octopamine promotes wakefulness in the fly by acting through the insulin-producing cells (IPCs) in the fly brain. To determine if insulin signaling mediates the effects of octopamine on sleep:wake behavior, we assayed flies in which insulin signaling activity was genetically altered. We found that increasing insulin signaling does not promote wake, nor does insulin appear to mediate the wake-promoting effects of octopamine. Octopamine also affects metabolism in invertebrate species, including, as we show here, Drosophila melanogaster. Triglycerides are decreased in mutants with compromised octopamine signaling and elevated in flies with increased activity of octopaminergic neurons. Interestingly, this effect is mediated at least partially by insulin, suggesting that effects of octopamine on metabolism are independent of its effects on sleep. We further investigated the relative contribution of metabolic and sleep phenotypes to the starvation response of flies with altered octopamine signaling. Hyperactivity (indicative of foraging) induced by starvation was elevated in octopamine receptor mutants, despite their high propensity for sleep, indicating that their metabolic state dictates their behavioral response under these conditions. Moreover, flies with increased octopamine signaling do not suppress sleep in response to starvation, even though they are normally hyper-aroused, most likely because of their high triglyceride levels. Together, these data suggest that observed correlations between sleep and metabolic phenotypes can result from shared molecular pathways rather than causality, and environmental conditions can lead to the dominance of one phenotype over the other.
Asunto(s)
Octopamina/metabolismo , Transducción de Señal/fisiología , Sueño/fisiología , Animales , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster , Insulina/genética , Insulina/metabolismo , Mutación , Octopamina/genética , Receptores de Amina Biogénica/genética , Receptores de Amina Biogénica/metabolismo , Triglicéridos/genética , Triglicéridos/metabolismoRESUMEN
Amines are one class of signaling molecules used by nervous systems. In crustaceans, four amines are recognized: dopamine, histamine, octopamine, and serotonin. While much is known about the physiological actions of amines in crustaceans, little is known about them at the molecular level. Recently, we mined the Daphnia pulex genome for proteins required for histaminergic signaling. Here, we expand this investigation, mining the D. pulex genome for proteins necessary for dopamine, octopamine and serotonin signaling. Using known Drosophila protein sequences, the D. pulex database was queried for genes encoding homologs of amine biosynthetic enzymes, receptors and transporters. Among the proteins identified were the biosynthetic enzymes tryptophan-phenylalanine hydroxylase (dopamine, octopamine and serotonin), tyrosine hydroxylase (dopamine), DOPA decarboxylase (dopamine and serotonin), tyrosine decarboxylase (octopamine), tyramine ß-hydroxylase (octopamine) and tryptophan hydroxylase (serotonin), as well as receptors for each amine and several amine transporters (dopamine and serotonin). Comparisons of the Daphnia proteins with their Drosophila queries showed high sequence identity/similarity, particularly in domains required for function. The data presented in this study provide the first molecular descriptions of dopamine, octopamine and serotonin signaling systems in Daphnia, and provide foundations for future molecular, biochemical, anatomical, and physiological investigations of aminergic signaling in this species.
Asunto(s)
Daphnia/genética , Daphnia/metabolismo , Dopamina/metabolismo , Genómica , Octopamina/metabolismo , Serotonina/metabolismo , Secuencia de Aminoácidos , Animales , Daphnia/química , Dopamina/genética , Datos de Secuencia Molecular , Octopamina/genética , Proteoma/química , Proteoma/genética , Proteoma/metabolismo , Alineación de Secuencia , Serotonina/genética , Transducción de SeñalRESUMEN
There is ample evidence linking octopamine (OA) and tyramine (TA) to several neurophysiological functions in arthropods. In our laboratory we use the freshwater prawn Macrobrachium rosenbergii to study the neural basis of aggressive behavior. As a first step towards understanding the possible role of these amines and their receptors in the modulation of interactive behaviors, we have cloned a putative octopamine/tyramine receptor. The predicted sequence of the cloned OA/TA(Mac) receptor consists of 1,579 base pairs (bp), with an open reading frame of 1,350bp that encodes a 450 amino acid protein. This putative protein displays sequence identities of 70% to an Aedes aegypti mosquito TA receptor, followed by 60% to a Stegomyia aegypti mosquito OA receptor, 59% and 58% to the migratory locust TA-1 and -2 receptors respectively, and 57% with the silkworm OA receptor. We also mapped the OA/TA(Mac) receptor distribution by in-situ hybridization to the receptor's mRNA, and by immunohistochemistry to its protein. We observed stained cell bodies for the receptor's mRNA, mainly in the midline region of the thoracic and in the abdominal ganglia, as well as diffuse staining in the brain ganglia. For the receptor's protein, we observed extensive punctate staining within the neuropil and on the membrane of specific groups of neurons in all ganglia throughout the CNS, including the brain, the midline region and neuropiles of the thoracic ganglia, and ventral part and neuropiles of the abdominal ganglia. The same pattern of stained cells was observed on the thoracic and abdominal ganglia in both in-situ hybridization and immunohistochemistry experiments. Diffuse staining observed with in-situ hybridization also coincides with punctate staining observed in brain, SEG, thoracic, and abdominal ganglia in immunohistochemical preparations. This work provides the first step towards characterizing the neural networks that mediate octopaminergic signaling in prawn.
Asunto(s)
Sistema Nervioso Central/metabolismo , Octopamina/metabolismo , Palaemonidae/anatomía & histología , Receptores de Amina Biogénica/metabolismo , Animales , Evolución Biológica , Sistema Nervioso Central/anatomía & histología , Clonación Molecular/métodos , Ganglios/metabolismo , Neurópilo/metabolismo , Octopamina/genética , Filogenia , ARN Mensajero/metabolismo , Receptores de Amina Biogénica/genéticaRESUMEN
Sleep is a fundamental process, but its regulation and function are still not well understood. The Drosophila model for sleep provides a powerful system to address the genetic and molecular mechanisms underlying sleep and wakefulness. Here we show that a Drosophila biogenic amine, octopamine, is a potent wake-promoting signal. Mutations in the octopamine biosynthesis pathway produced a phenotype of increased sleep, which was restored to wild-type levels by pharmacological treatment with octopamine. Moreover, electrical silencing of octopamine-producing cells decreased wakefulness, whereas excitation of these neurons promoted wakefulness. Because protein kinase A (PKA) is a putative target of octopamine signaling and is also implicated in Drosophila sleep, we investigated its role in the effects of octopamine on sleep. We found that decreased PKA activity in neurons rendered flies insensitive to the wake-promoting effects of octopamine. However, this effect of PKA was not exerted in the mushroom bodies, a site previously associated with PKA action on sleep. These studies identify a novel pathway that regulates sleep in Drosophila.
Asunto(s)
Encéfalo/metabolismo , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Drosophila/metabolismo , Octopamina/biosíntesis , Sueño/genética , Administración Oral , Animales , Encéfalo/citología , Drosophila/genética , Proteínas de Drosophila/metabolismo , Femenino , Proteínas Fluorescentes Verdes , Masculino , Actividad Motora/efectos de los fármacos , Actividad Motora/genética , Cuerpos Pedunculados/citología , Cuerpos Pedunculados/metabolismo , Mutación/genética , Neuronas/metabolismo , Norepinefrina/metabolismo , Octopamina/genética , Octopamina/farmacología , Sueño/efectos de los fármacos , Tiramina/metabolismo , Tirosina Descarboxilasa/metabolismo , Vigilia/efectos de los fármacos , Vigilia/genéticaRESUMEN
How do deficits in neuronal growth, aging or synaptic function affect the final, mechanical output of a single muscle twitch? We address this in vivo (indeed in situ) with a novel ergometer that records the output of a large specialised muscle, the Drosophila jump muscle. Here, we describe in detail the ergometer, its construction and use. We evaluated the ergometer by showing that adult fly jump muscle output varies little between 3 h and 7 days; but newly eclosed flies produce only 65%. In a mutant with little octopamine (Tbetah), jump muscle performance is reduced by 28%. The initial responses of synaptic growth mutants (highwire and spinster) do not differ from wild type, as expected on the homeostatic hypothesis. However, responses in highwire mutations gradually decline following repeated stimuli, suggesting physiological as well as anatomical abnormalities. We conclude that the assay is robust, sensitive and reliable with a good throughput.
Asunto(s)
Drosophila/fisiología , Ergometría , Músculos/fisiología , Mutación , Unión Neuromuscular/fisiología , Animales , Conducta Animal , Drosophila/genética , Ergometría/instrumentación , Ergometría/métodos , Octopamina/genéticaRESUMEN
Aggression is an innate behavior that is important for animal survival and evolution. We examined the molecular and cellular mechanisms underlying aggression in Drosophila. Reduction of the neurotransmitter octopamine, the insect equivalent of norepinephrine, decreased aggression in both males and females. Mutants lacking octopamine did not initiate fighting and did not fight other flies, although they still provoked other flies to fight themselves. Mutant males lost to the wild-type males in fighting and in competing for copulation with females. Enhanced octopaminergic signaling increased aggression in socially grouped flies, but not in socially isolated flies. We carried out genetic rescue experiments that revealed the functional importance of neuronal octopamine and identified a small subset of octopaminergic neurons in the suboesophageal ganglion as being important for aggression.
Asunto(s)
Agresión , Drosophila/fisiología , Neuronas/clasificación , Neuronas/metabolismo , Octopamina/metabolismo , Análisis de Varianza , Animales , Animales Modificados Genéticamente , Conducta Animal , Clorfenamidina/farmacología , Discriminación en Psicología/fisiología , Relación Dosis-Respuesta a Droga , Drosophila/anatomía & histología , Proteínas de Drosophila/genética , Femenino , Regulación de la Expresión Génica/fisiología , Proteínas Fluorescentes Verdes/genética , Calor , Locomoción/efectos de los fármacos , Locomoción/genética , Masculino , Oxigenasas de Función Mixta/genética , Modelos Biológicos , Inhibidores de la Monoaminooxidasa/farmacología , Mutación/genética , Sistema Nervioso/citología , Neuronas/efectos de los fármacos , Octopamina/genética , Tiempo de Reacción/genética , Conducta Sexual Animal/efectos de los fármacos , Conducta Sexual Animal/fisiología , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología , Conducta SocialRESUMEN
Insect flight is one of the fastest, most intense and most energy-demanding motor behaviors. It is modulated on multiple levels by the biogenic amine octopamine. Within the CNS, octopamine acts directly on the flight central pattern generator, and it affects motivational states. In the periphery, octopamine sensitizes sensory receptors, alters muscle contraction kinetics, and enhances flight muscle glycolysis. This study addresses the roles for octopamine and its precursor tyramine in flight behavior by genetic and pharmacological manipulation in Drosophila. Octopamine is not the natural signal for flight initiation because flies lacking octopamine [tyramine-beta-hydroxylase (TbetaH) null mutants] can fly. However, they show profound differences with respect to flight initiation and flight maintenance compared with wild-type controls. The morphology, kinematics, and development of the flight machinery are not impaired in TbetaH mutants because wing-beat frequencies and amplitudes, flight muscle structure, and overall dendritic structure of flight motoneurons are unaffected in TbetaH mutants. Accordingly, the flight behavior phenotypes can be rescued acutely in adult flies. Flight deficits are rescued by substituting octopamine but also by blocking the receptors for tyramine, which is enriched in TbetaH mutants. Conversely, ablating all neurons containing octopamine or tyramine phenocopies TbetaH mutants. Therefore, both octopamine and tyramine systems are simultaneously involved in regulating flight initiation and maintenance. Different sets of rescue experiments indicate different sites of action for both amines. These findings are consistent with a complex system of multiple amines orchestrating the control of motor behaviors on multiple levels rather than single amines eliciting single behaviors.
Asunto(s)
Aminas Biogénicas/antagonistas & inhibidores , Aminas Biogénicas/metabolismo , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Vuelo Animal/fisiología , Animales , Aminas Biogénicas/fisiología , Drosophila melanogaster/enzimología , Drosophila melanogaster/fisiología , Masculino , Oxigenasas de Función Mixta/deficiencia , Oxigenasas de Función Mixta/genética , Actividad Motora/genética , Mutación , Octopamina/antagonistas & inhibidores , Octopamina/genética , Octopamina/fisiologíaRESUMEN
The nervous system plays a critical role in adaptation to a new environment. In Caenorhabditis elegans, reduced access to food requires both changes in behavior as well as metabolic adaptation for survival, which is postulated to involve the bioamine octopamine. The transcription factor cAMP response element-binding protein (CREB) is generally activated by G-protein-coupled receptors (GPCRs) that activate G alpha(s) and is known to play an important role in long-term changes, including synaptic plasticity. We show that, in C. elegans, the CREB ortholog CRH-1 (CREB homolog family member 1) activates in vivo a cAMP response element-green fluorescent protein fusion reporter in a subset of neurons during starvation. This starvation response is mediated by octopamine via the GPCR SER-3 (serotonin/octopamine receptor family member 3) and is fully dependent on the subsequent activation of the G alpha(q) ortholog EGL-30 (egg-laying defective family member 30). The signaling cascade is only partially dependent on the phospholipase C beta (EGL-8) and is negatively regulated by G alpha(o) [GOA-1 (G-protein, O, alpha subunit family member 1)] and calcium/calmodulin-dependent kinase [UNC-43 (uncoordinated family member 43)]. Nonstarved animals in a liquid environment mediate a similar response that is octopamine independent. The results show that the endogenous octopamine system in C. elegans is activated by starvation and that different environmental stimuli can activate CREB through G alpha(q).