RESUMEN
In 1998, a special edition of Learning & Memory was published with a discrete focus of synthesizing the state of the field to provide an overview of the function of the insect mushroom body. While molecular neuroscience and optical imaging of larger brain areas were advancing, understanding the basic functioning of neuronal circuits, particularly in the context of the mushroom body, was rudimentary. In the past 25 years, technological innovations have allowed researchers to map and understand the in vivo function of the neuronal circuits of the mushroom body system, making it an ideal model for investigating the circuit basis of sensory encoding, memory formation, and behavioral decisions. Collaborative efforts within the community have played a crucial role, leading to an interactive connectome of the mushroom body and accessible genetic tools for studying mushroom body circuit function. Looking ahead, continued technological innovation and collaborative efforts are likely to further advance our understanding of the mushroom body and its role in behavior and cognition, providing insights that generalize to other brain structures and species.
Asunto(s)
Encéfalo , Insectos , Cuerpos Pedunculados , Cuerpos Pedunculados/fisiología , Animales , Insectos/fisiología , Encéfalo/fisiología , Historia del Siglo XXI , Historia del Siglo XXRESUMEN
Drosophila melanogaster is a powerful genetic model for investigating the mechanisms underlying ethanol-induced behaviors, metabolism, and preference. Ethanol-induced locomotor activity is especially useful for understanding the mechanisms by which ethanol acutely affects the brain and behavior. Ethanol-induced locomotor activity is characterized by hyperlocomotion and subsequent sedation with increased exposure duration or concentration. Locomotor activity is an efficient, easy, robust, and reproducible behavioral screening tool for identifying underlying genes and neuronal circuits as well as investigating genetic and molecular pathways. We introduce a detailed protocol for performing experiments investigating how volatilized ethanol affects locomotor activity using the fly Group Activity Monitor (flyGrAM). We introduce installation, implementation, data collection, and subsequent data-analysis methods for investigating how volatilized stimuli affect activity. We also introduce a procedure for how to optogenetically probe neuronal activity to identify the neural mechanisms underlying locomotor activity.
Asunto(s)
Drosophila , Etanol , Animales , Etanol/metabolismo , Drosophila melanogaster/genética , Conducta Animal/fisiología , Neuronas/metabolismoRESUMEN
Locomotion is a behavioral readout that can be used to understand responses to specific stimuli or perturbations. The fly Group Activity Monitor (flyGrAM) provides a high-throughput and high-content readout of the acute stimulatory and sedative effects of ethanol. The flyGrAM system is adaptable and seamlessly introduces thermogenetic or optogenetic stimulation to dissect neural circuits underlying behavior and tests responses to other volatilized stimuli (humidified air, odorants, anesthetics, vaporized drugs of abuse, etc.). The automated quantification and readout of activity provide users with a real-time representation of the group activity within each chamber throughout the experiment, helping users to quickly determine proper ethanol doses and duration, run behavioral screens, and plan follow-up experiments.
Asunto(s)
Drosophila , Etanol , Animales , Conducta Animal , Locomoción , OptogenéticaAsunto(s)
Etanol , Mitocondrias , Plasticidad Neuronal , Animales , Drosophila melanogaster , Etanol/toxicidad , Hipocampo/efectos de los fármacos , Hipocampo/metabolismo , Ratones , Mitocondrias/efectos de los fármacos , Plasticidad Neuronal/efectos de los fármacos , Plasticidad Neuronal/fisiología , ProteómicaRESUMEN
Opiate/opioid use disorder (OUD) is a chronic relapsing brain disorder that has increased in prevalence in the last two decades in the United States. Understanding the molecular correlates of OUD may provide key insights into the pathophysiology of this syndrome. Using publicly available RNA-sequencing data, our study investigated the possible role of alternative mRNA splicing in human brain tissue (dorsal-lateral prefrontal cortex (dlPFC), nucleus accumbens (NAc), and midbrain) of 90 individuals with OUD or matched controls. We found a total of 788 differentially spliced genes across brain regions. Alternative mRNA splicing demonstrated mostly tissue-specific effects, but a functionally characterized splicing change in the clathrin and AP-2-binding (CLAP) domain of the Bridging Integrator 1 (BIN1) gene was significantly linked to OUD across all brain regions. We investigated two hypotheses that may underlie differential splicing in OUD. First, we tested whether spliceosome genes were disrupted in the brains of individuals with OUD. Pathway enrichment analyses indicated spliceosome perturbations in OUD across brain regions. Second, we tested whether alternative mRNA splicing regions were linked to genetic predisposition. Using a genome-wide association study (GWAS) of OUD, we found no evidence that DNA variants within or surrounding differentially spliced genes were implicated in the heritability of OUD. Altogether, our study contributes to the understanding of OUD pathophysiology by providing evidence of a possible role of alternative mRNA splicing in OUD.
Asunto(s)
Estudio de Asociación del Genoma Completo , Trastornos Relacionados con Opioides , Analgésicos Opioides , ADN Recombinante , Humanos , Trastornos Relacionados con Opioides/epidemiología , Trastornos Relacionados con Opioides/genética , ARN Mensajero/genética , Recompensa , Estados UnidosRESUMEN
The genetic basis of alcohol use disorder (AUD) is complex. Understanding how natural genetic variation contributes to alcohol phenotypes can help us identify and understand the genetic basis of AUD. Recently, a single nucleotide polymorphism in the human foraging (for) gene ortholog, Protein Kinase cGMP-Dependent 1 (PRKG1), was found to be associated with stress-induced risk for alcohol abuse. However, the mechanistic role that PRKG1 plays in AUD is not well understood. We use natural variation in the Drosophila for gene to describe how variation of cGMP-dependent protein kinase (PKG) activity modifies ethanol-induced phenotypes. We found that variation in for affects ethanol-induced increases in locomotion and memory of the appetitive properties of ethanol intoxication. Further, these differences may stem from the ability to metabolize ethanol. Together, this data suggests that natural variation in PKG modulates cue reactivity for alcohol, and thus could influence alcohol cravings by differentially modulating metabolic and behavioral sensitivities to alcohol.
Asunto(s)
Alcoholismo/genética , Depresores del Sistema Nervioso Central/metabolismo , Proteínas Quinasas Dependientes de GMP Cíclico/genética , Proteínas de Drosophila/genética , Etanol/metabolismo , Memoria/fisiología , Animales , Depresores del Sistema Nervioso Central/farmacología , Drosophila melanogaster , Etanol/farmacología , Locomoción/efectos de los fármacosRESUMEN
The mushroom body (MB) is a well-characterized associative memory structure within the Drosophila brain. Analyzing MB connectivity using multiple approaches is critical for understanding the functional implications of this structure. Using the genetic anterograde transsynaptic tracing tool, trans-Tango, we identified divergent projections across the brain and convergent downstream targets of the MB output neurons (MBONs). Our analysis revealed at least three separate targets that receive convergent input from MBONs: other MBONs, the fan-shaped body (FSB), and the lateral accessory lobe (LAL). We describe, both anatomically and functionally, a multilayer circuit in which inhibitory and excitatory MBONs converge on the same genetic subset of FSB and LAL neurons. This circuit architecture enables the brain to update and integrate information with previous experience before executing appropriate behavioral responses. Our use of trans-Tango provides a genetically accessible anatomical framework for investigating the functional relevance of components within these complex and interconnected circuits.
Asunto(s)
Drosophila melanogaster/fisiología , Cuerpos Pedunculados/fisiología , Neuronas/fisiología , Animales , Femenino , MasculinoRESUMEN
A powerful feature of adaptive memory is its inherent flexibility. Alcohol and other addictive substances can remold neural circuits important for memory to reduce this flexibility. However, the mechanism through which pertinent circuits are selected and shaped remains unclear. We show that circuits required for alcohol-associated preference shift from population level dopaminergic activation to select dopamine neurons that predict behavioral choice in Drosophila melanogaster. During memory expression, subsets of dopamine neurons directly and indirectly modulate the activity of interconnected glutamatergic and cholinergic mushroom body output neurons (MBON). Transsynaptic tracing of neurons important for memory expression revealed a convergent center of memory consolidation within the mushroom body (MB) implicated in arousal, and a structure outside the MB implicated in integration of naïve and learned responses. These findings provide a circuit framework through which dopamine neuronal activation shifts from reward delivery to cue onset, and provide insight into the maladaptive nature of memory.
Asunto(s)
Dopamina/metabolismo , Neuronas Dopaminérgicas , Etanol , Memoria , Animales , Neuronas Dopaminérgicas/citología , Neuronas Dopaminérgicas/fisiología , Drosophila melanogaster/fisiología , Etanol/metabolismo , Etanol/farmacología , Femenino , Masculino , Memoria/efectos de los fármacos , Memoria/fisiología , Cuerpos Pedunculados/citología , Cuerpos Pedunculados/fisiología , Red Nerviosa/fisiología , Recompensa , Sinapsis/fisiologíaRESUMEN
Repeated alcohol experiences can produce long-lasting memories for sensory cues associated with intoxication. These memories can problematically trigger relapse in individuals recovering from alcohol use disorder (AUD). The molecular mechanisms by which ethanol changes memories to become long-lasting and inflexible remain unclear. New methods to analyze gene expression within precise neuronal cell types can provide further insight toward AUD prevention and treatment. Here, we used genetic tools in Drosophila melanogaster to investigate the lasting consequences of ethanol on transcription in memory-encoding neurons. Drosophila rely on mushroom body (MB) neurons to make associative memories, including memories of ethanol-associated sensory cues. Differential expression analyses revealed that distinct transcripts, but not genes, in the MB were associated with experiencing ethanol alone compared to forming a memory of an odor cue associated with ethanol. Adult MB-specific knockdown of spliceosome-associated proteins demonstrated the necessity of RNA-processing in ethanol memory formation. These findings highlight the dynamic, context-specific regulation of transcription in cue-encoding neurons, and the lasting effect of ethanol on transcript usage during memory formation.
Asunto(s)
Depresores del Sistema Nervioso Central/farmacología , Etanol/farmacología , Cuerpos Pedunculados/efectos de los fármacos , Células Receptoras Sensoriales/efectos de los fármacos , Transcriptoma , Animales , Drosophila melanogaster , Memoria , Cuerpos Pedunculados/citología , Cuerpos Pedunculados/fisiología , Células Receptoras Sensoriales/metabolismo , Empalmosomas/genética , Empalmosomas/metabolismoRESUMEN
Recent advances in neurogenetics have highlighted Drosophila melanogaster as an exciting model to study neural circuit dynamics and complex behavior. Automated tracking methods have facilitated the study of complex behaviors via high throughput behavioral screening. Here we describe a newly developed low-cost assay capable of real-time monitoring and quantifying Drosophila group activity. This platform offers reliable real-time quantification with open source software and a user-friendly interface for data acquisition and analysis. We demonstrate the utility of this platform by characterizing ethanol-induced locomotor activity in a dose-dependent manner as well as the effects of thermo and optogenetic manipulation of ellipsoid body neurons important for ethanol-induced locomotor activity. As expected, low doses of ethanol induced an initial startle and slow ramping of group activity, whereas high doses of ethanol induced sustained group activity followed by sedation. Advanced offline processing revealed discrete behavioral features characteristic of intoxication. Thermogenetic inactivation of ellipsoid body ring neurons reduced group activity whereas optogenetic activation increased activity. Together, these data establish the fly Group Activity Monitor (flyGrAM) platform as a robust means of obtaining an online read out of group activity in response to manipulations to the environment or neural activity, with an opportunity for more advanced post-processing offline.
Asunto(s)
Conducta Animal/efectos de los fármacos , Drosophila melanogaster/fisiología , Etanol/farmacología , Locomoción/efectos de los fármacos , Neuronas/fisiología , Optogenética , Termogénesis , Animales , Antiinfecciosos Locales/farmacología , Drosophila melanogaster/efectos de los fármacos , Femenino , Masculino , Neuronas/efectos de los fármacosRESUMEN
Our understanding of alcohol use disorder (AUD), particularly alcohol's effects on the nervous system, has unquestionably benefited from the use of model systems such as Drosophila melanogaster. Here, we briefly introduce the use of flies in alcohol research, and highlight the genetic accessibility and neurobiological contribution that flies have made to our understanding of AUD. Future fly research offers unique opportunities for addressing unresolved questions in the alcohol field, such as the neuromolecular and circuit basis for cravings and alcohol-induced neuroimmune dysfunction. This review strongly advocates for interdisciplinary approaches and translational collaborations with the united goal of confronting the major health problems associated with alcohol abuse and addiction.
Asunto(s)
Alcoholismo/genética , Drosophila melanogaster/efectos de los fármacos , Drosophila melanogaster/genética , Animales , Drosophila melanogaster/fisiología , Estudio de Asociación del Genoma CompletoRESUMEN
Drugs of abuse, like alcohol, modulate gene expression in reward circuits and consequently alter behavior. However, the in vivo cellular mechanisms through which alcohol induces lasting transcriptional changes are unclear. We show that Drosophila Notch/Su(H) signaling and the secreted fibrinogen-related protein Scabrous in mushroom body (MB) memory circuitry are important for the enduring preference of cues associated with alcohol's rewarding properties. Alcohol exposure affects Notch responsivity in the adult MB and alters Su(H) targeting at the dopamine-2-like receptor (Dop2R). Alcohol cue training also caused lasting changes to the MB nuclear transcriptome, including changes in the alternative splicing of Dop2R and newly implicated transcripts like Stat92E. Together, our data suggest that alcohol-induced activation of the highly conserved Notch pathway and accompanying transcriptional responses in memory circuitry contribute to addiction. Ultimately, this provides mechanistic insight into the etiology and pathophysiology of alcohol use disorder.
Asunto(s)
Proteínas de Drosophila/metabolismo , Etanol/administración & dosificación , Glicoproteínas/metabolismo , Memoria/efectos de los fármacos , Cuerpos Pedunculados/metabolismo , Receptores Notch/metabolismo , Proteínas Represoras/metabolismo , Animales , Señales (Psicología) , Masculino , Cuerpos Pedunculados/efectos de los fármacos , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Odorantes , Isoformas de Proteínas/metabolismo , Receptores de Dopamina D2/metabolismo , TranscriptomaRESUMEN
Alcohol use disorder generates devastating social, medical and economic burdens, making it a major global health issue. The persistent nature of memories associated with intoxication experiences often induces cravings and triggers relapse in recovering individuals. Despite recent advances, the neural and molecular mechanisms underlying these memories are complex and not well understood. This makes finding effective pharmacological targets challenging. The investigation of persistent alcohol-associated memories in the fruit fly, Drosophila melanogaster, presents a unique opportunity to gain a comprehensive understanding of the memories for ethanol reward at the level of genes, molecules, neurons and circuits. Here we characterize the dose-dependent nature of ethanol on the expression of memory for an intoxication experience. We report that the concentration of ethanol, number of ethanol exposures, length of ethanol exposures, and timing between ethanol exposures are critical in determining whether ethanol is perceived as aversive or appetitive, and in how long the memory for the intoxicating properties of ethanol last. Our study highlights that fruit flies display both acute and persistent memories for ethanol-conditioned odor cues, and that a combination of parameters that determine the intoxication state of the fly influence the seemingly complex retention and expression of memories associated with intoxication. Our thorough behavioral characterization provides the opportunity to interrogate the biological underpinnings of these observed preference differences in future studies.
RESUMEN
Dopamine is associated with a variety of conserved responses across species including locomotion, sleep, food consumption, aggression, courtship, addiction and several forms of appetitive and aversive memory. Historically, dopamine has been most prominently associated with dynamics underlying reward, punishment, or salience. Recent emerging evidence from Drosophila supports a role in all of these functions, as well as additional roles in the interplay between external sensation and internal states and forgetting of the very memories dopamine helped encode. We discuss how cell-specific resolution and manipulation are elucidating the rules of dopamine's involvement in encoding valence and memory.
Asunto(s)
Dopamina/metabolismo , Drosophila/fisiología , Memoria/fisiología , Animales , Castigo , RecompensaRESUMEN
In a complex environment, animals learn from their responses to stimuli and events. Appropriate response to reward and punishment can promote survival, reproduction and increase evolutionary fitness. Interestingly, the neural processes underlying these responses are remarkably similar across phyla. In all species, dopamine is central to encoding reward and directing motivated behaviors, however, a comprehensive understanding of how circuits encode reward and direct motivated behaviors is still lacking. In part, this is a result of the sheer diversity of neurons, the heterogeneity of their responses and the complexity of neural circuits within which they are found. We argue that general features of reward circuitry are common across model organisms, and thus principles learned from invertebrate model organisms can inform research across species. In particular, we discuss circuit motifs that appear to be functionally equivalent from flies to primates. We argue that a comparative approach to studying and understanding reward circuit function provides a more comprehensive understanding of reward circuitry, and informs disorders that affect the brain's reward circuitry.
Asunto(s)
Encéfalo/fisiología , Motivación/fisiología , Recompensa , Animales , Conducta AnimalRESUMEN
UNLABELLED: Acute ethanol inebriation causes neuroadaptive changes in behavior that favor increased intake. Ethanol-induced alterations in gene expression, through epigenetic and other means, are likely to change cellular and neural circuit function. Ethanol markedly changes histone acetylation, and the sirtuin Sir2/SIRT1 that deacetylates histones and transcription factors is essential for the rewarding effects of long-term drug use. The molecular transformations leading from short-term to long-term ethanol responses mostly remain to be discovered. We find that Sir2 in the mushroom bodies of the fruit fly Drosophila promotes short-term ethanol-induced behavioral plasticity by allowing changes in the expression of presynaptic molecules. Acute inebriation strongly reduces Sir2 levels and increases histone H3 acetylation in the brain. Flies lacking Sir2 globally, in the adult nervous system, or specifically in the mushroom body α/ß-lobes show reduced ethanol sensitivity and tolerance. Sir2-dependent ethanol reward is also localized to the mushroom bodies, and Sir2 mutants prefer ethanol even without a priming ethanol pre-exposure. Transcriptomic analysis reveals that specific presynaptic molecules, including the synaptic vesicle pool regulator Synapsin, depend on Sir2 to be regulated by ethanol. Synapsin is required for ethanol sensitivity and tolerance. We propose that the regulation of Sir2/SIRT1 by acute inebriation forms part of a transcriptional program in mushroom body neurons to alter presynaptic properties and neural responses to favor the development of ethanol tolerance, preference, and reward. SIGNIFICANCE STATEMENT: We identify a mechanism by which acute ethanol inebriation leads to changes in nervous system function that may be an important basis for increasing ethanol intake and addiction liability. The findings are significant because they identify ethanol-driven transcriptional events that target presynaptic properties and direct behavioral plasticity. They also demonstrate that multiple forms of ethanol behavioral plasticity that are relevant to alcoholism are initiated by a shared mechanism. Finally, they link these events to the Drosophila brain region that associates context with innate approach and avoidance responses to code for reward and other higher-order behavior, similar in aspects to the role of the vertebrate mesolimbic system.
Asunto(s)
Intoxicación Alcohólica/metabolismo , Alcoholismo/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila/metabolismo , Histona Desacetilasas/metabolismo , Terminales Presinápticos/metabolismo , Recompensa , Sirtuinas/metabolismo , Intoxicación Alcohólica/genética , Alcoholismo/genética , Animales , Drosophila/genética , Drosophila/fisiología , Proteínas de Drosophila/genética , Histona Desacetilasas/genética , Histonas/metabolismo , Cuerpos Pedunculados/metabolismo , Terminales Presinápticos/fisiología , Sirtuinas/genética , Sinapsinas/genética , Sinapsinas/metabolismo , TranscriptomaRESUMEN
Hunger is a complex motivational state that drives multiple behaviors. The sensation of hunger is caused by an imbalance between energy intake and expenditure. One immediate response to hunger is increased food consumption. Hunger also modulates behaviors related to food seeking such as increased locomotion and enhanced sensory sensitivity in both insects and vertebrates. In addition, hunger can promote the expression of food-associated memory. Although progress is being made, how hunger is represented in the brain and how it coordinates these behavioral responses is not fully understood in any system. Here, we use Drosophila melanogaster to identify neurons encoding hunger. We found a small group of neurons that, when activated, induced a fed fly to eat as though it were starved, suggesting that these neurons are downstream of the metabolic regulation of hunger. Artificially activating these neurons also promotes appetitive memory performance in sated flies, indicating that these neurons are not simply feeding command neurons but likely play a more general role in encoding hunger. We determined that the neurons relevant for the feeding effect are serotonergic and project broadly within the brain, suggesting a possible mechanism for how various responses to hunger are coordinated. These findings extend our understanding of the neural circuitry that drives feeding and enable future exploration of how state influences neural activity within this circuit.
Asunto(s)
Drosophila melanogaster/fisiología , Hambre , Animales , Proteínas de Drosophila/metabolismo , Conducta Alimentaria , Femenino , Privación de Alimentos , Canales Iónicos , Masculino , Memoria , Motivación , Neuronas Serotoninérgicas/fisiología , Canal Catiónico TRPA1 , Canales Catiónicos TRPC/metabolismoRESUMEN
Animals discriminate stimuli, learn their predictive value and use this knowledge to modify their behavior. In Drosophila, the mushroom body (MB) plays a key role in these processes. Sensory stimuli are sparsely represented by â¼2000 Kenyon cells, which converge onto 34 output neurons (MBONs) of 21 types. We studied the role of MBONs in several associative learning tasks and in sleep regulation, revealing the extent to which information flow is segregated into distinct channels and suggesting possible roles for the multi-layered MBON network. We also show that optogenetic activation of MBONs can, depending on cell type, induce repulsion or attraction in flies. The behavioral effects of MBON perturbation are combinatorial, suggesting that the MBON ensemble collectively represents valence. We propose that local, stimulus-specific dopaminergic modulation selectively alters the balance within the MBON network for those stimuli. Our results suggest that valence encoded by the MBON ensemble biases memory-based action selection.
Asunto(s)
Conducta de Elección , Drosophila melanogaster/citología , Drosophila melanogaster/fisiología , Memoria , Cuerpos Pedunculados/citología , Cuerpos Pedunculados/inervación , Neuronas/fisiología , Animales , Conducta Apetitiva/efectos de la radiación , Aprendizaje por Asociación/efectos de la radiación , Reacción de Prevención/efectos de la radiación , Conducta Animal/efectos de la radiación , Conducta de Elección/efectos de la radiación , Luz , Memoria/efectos de la radiación , Modelos Neurológicos , Cuerpos Pedunculados/efectos de la radiación , Neuronas/efectos de la radiación , Odorantes , Sueño/efectos de la radiación , Factores de Tiempo , Visión OcularRESUMEN
Epidermal Growth Factor Receptor (EGFR) signaling has a conserved role in ethanol-induced behavior in flies and mice, affecting ethanol-induced sedation in both species. However it is not known what other effects EGFR signaling may have on ethanol-induced behavior, or what roles other Receptor Tyrosine Kinase (RTK) pathways may play in ethanol induced behaviors. We examined the effects of both the EGFR and Fibroblast Growth Factor Receptor (FGFR) RTK signaling pathways on ethanol-induced enhancement of locomotion, a behavior distinct from sedation that may be associated with the rewarding effects of ethanol. We find that both EGFR and FGFR genes influence ethanol-induced locomotion, though their effects are opposite - EGFR signaling suppresses this behavior, while FGFR signaling promotes it. EGFR signaling affects development of the Drosophila mushroom bodies in conjunction with the JNK MAP kinase basket (bsk), and with the Ste20 kinase tao, and we hypothesize that the EGFR pathway affects ethanol-induced locomotion through its effects on neuronal development. We find, however, that FGFR signaling most likely affects ethanol-induced behavior through a different mechanism, possibly through acute action in adult neurons.