These studies were supported by grants from the National Institutes of Health, National Institute for Alcoholism and Alcohol Abuse to M.G. (P20AA017828, R01AA020634, P50 AA022537). The authors thank Jill Bettinger for helpful discussions and Jacqueline DeLoyht for technical assistance.

1. Jour avant le test <ol><li> Recueillir les mouches dans des flacons de produits frais dans des groupes de 11 (même sexe) sous brève (1-5 min) CO 2. </li><li> Permettre aux mouches de récupérer O / N dans des flacons de produits alimentaires dans un espace à environnement contrôlé (généralement 25 ° C, 60% d&#39;humidité relative, 12 h cycle lumière / obscurité). </li><li> Préparer la solution (s) de l&#39;éthanol en diluant pur (100%) dans de l&#39;éthanol purifié (≥18 MQ) de l&#39;eau à la concentration finale (s) approprié à l&#39;essai prévu. Laisser la solution (s) pour revenir à RT O / N. Remarque: La dilution de l&#39;éthanol est exothermique. </li></ol> 2. jour de l&#39;analyse <ol><li> Pour chaque flacon de mouches à tester, préparer (i) un endroit propre, vide alimentaire flacon; ce est à dire, les tests flacon, (ii) une nouvelle fiche d&#39;acétate de cellulose, (iii) un bouchon en silicone et (iv) 1 ml de solution d&#39;éthanol (voir le tableau 3). </li><li> Préparer chambre d&#39;essai en ajustant la température à 20-25 ° C et une humidité relative de 55-65%. </li><li> Avoir un autre travailleurattribuer un code unique à chaque groupe de flacons et d&#39;enregistrer le code pour plus tard. Placez flacons codés avec des mouches dans la salle de test pour se acclimater pendant quelques minutes. </li><li> Étiqueter les flacons d&#39;essai vides pour correspondre codes sur les flacons de mouches de 2,3. </li><li> Construire un disque journal de test de copie en introduisant les codes dans les colonnes (une colonne / flacon) dans une feuille de calcul semblable au tableau 1. </li><li> Utilisation du journal de test comme un guide, organiser des flacons alimentaires codés avec des mouches et des flacons d&#39;essai vides en correspondant à des tableaux dans la salle d&#39;examen. Remarque: Un nombre maximum raisonnable de flacons à tester est 24 (ce est à dire, six séries de quatre flacons chacun). </li><li> Transfert d&#39;oiseau de flacons d&#39;aliments dans appariés flacons d&#39;essai vide / étiquetés et insérer immédiatement cellulose bouchons d&#39;acétate dans des flacons d&#39;essai jusqu&#39;à ce que la cellulose d&#39;acétate bouchons sont 2 cm en dessous de la cime des flacons. </li><li> Ci-après, traiter chaque rangée de quatre flacons comme un ensemble au quinconce des intervalles de 1 min. </li><li> Pour l&#39;évaluation du temps 0, saisir chaque flacon individuellement wième pouce et l&#39;index, appuyez doucement sur la table trois fois pour frapper les mouches au fond du flacon, attendez 30 secondes puis de compter le nombre de mouches qui sont immobiles / morts. Notez le nombre de mouches immobiles / morts pour chaque flacon au temps 0 min dans le journal de test papier. </li><li> Commencez minuterie compter en continu au temps 0 et commencer immédiatement ajoutant 1 ml d&#39;éthanol à la cellulose d&#39;acétate bouchons dans les flacons pour la première ligne / ensemble de 4 flacons. Ajouter 1 ml d&#39;éthanol aux bougies d&#39;acétate de cellulose dans les flacons à intervalles de 5 secondes dans l&#39;ordre où elles seront testés. Ajouter de l&#39;éthanol pour les bouchons d&#39;acétate de cellulose dans un mouvement circulaire de sorte que l&#39;éthanol est absorbé uniformément dans les bouchons d&#39;acétate de cellulose. Lorsque l&#39;éthanol a été ajouté à tous les quatre flacons d&#39;essai dans le jeu, insérez un bouchon de silicone dans chaque flacon pour le sceller. </li><li> Parfois, 1, 2, 3, 4 et 5 min, ajouter 1 ml d&#39;éthanol à la seconde, troisième, quatrième, cinquième et ensembles de quatre flacons sixième, respectivement. Continuer introduction de bouchons en silicone aprèsajouter de l&#39;éthanol à chaque jeu de quatre flacons. </li><li> Au moment de 6 min, tester la première série de quatre flacons en saisissant chaque flacon avec le pouce et l&#39;index, tapotant doucement sur la table trois fois pour frapper les mouches au fond du flacon, en attendant 30 secondes, puis en comptant et en enregistrant le nombre total de mouches qui sont sous sédation. Score vole comme sédatif se ils (i) se tiennent sur le plancher de la fiole, mais ne marchent pas ou (ii) se trouvent sur le dos avec ou sans battant des ailes. </li><li> Tenez chaque flacon dans l&#39;ensemble à intervalles de 5 secondes en utilisant le calendrier dans le tableau 2. </li><li> Parfois, 7, 8, 9, 10 et 11 min, de tester les deuxième, troisième, quatrième, cinquième et sixième ensembles de fioles, respectivement, comme cela se fait pour le premier ensemble. </li><li> Au moment 12 min, tester la première série de quatre flacons à nouveau comme décrit dans 2.12 et continuer à tester les deuxième, troisième, quatrième, ensembles, cinquième et sixième de flacons à 13, 14, 15, 16 et 17 minutes, respectivement. </li><li> Continuer essai mouches comme décrit dans 2.12 jusqu&#39;à ce que toutes les mouches sont sedated. </li><li> Entrez le nombre total de mouches dans chaque flacon dans le disque journal de test de copie. Censurer mouches immobiles / morts au temps 0 à partir du nombre total de mouches. </li><li> Calculer le pourcentage mouches actifs à chaque évaluation point dans le temps et que les données des placettes% mouches actifs (axe des ordonnées) en fonction du temps (axe des x). Quantifier éthanol sédation par interpolation sédation Durée 50 valeurs (ST50, le temps de 50% la sédation) du troisième ordre courbe polynomiale ou sigmoïde se adapte ou calculer l&#39;aire sous la courbe. </li><li> Compiler les données décodées de flacons et effectuer des analyses statistiques (par exemple, une analyse de la variance avec Bonferroni test de comparaison multiple) en fonction de la conception expérimentale. </li></ol>

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The authors have nothing to disclose.

Tests simples qui quantifient de manière reproductible phénotypes significatives sont d&#39;une grande valeur pour l&#39;analyse du comportement. Les travaux décrits ici traite de plusieurs aspects pratiques d&#39;un test pour mesurer la sensibilité de l&#39;éthanol de sédation et la tolérance rapide chez la drosophile. Bien que pas un objectif de ce travail, des analyses comportementales sont facilitées par le maintien de l&#39;environnement et génétique constante de fond pour les sujets de test dans...

trouble de consommation d&#39;alcool (AUD) est un problème de santé dans le monde énorme (examiné en 1). Bien que les mécanismes favorisant le développement de AUD sont complexes, ces troubles ont une composante génétique importante (par exemple, 2). La grande héritabilité de l&#39;AUD et les réponses comportementales conservés à l&#39;éthanol dans de nombreuses espèces (3,4) examinés en ont généré un vif intérêt dans l&#39;utilisation des organismes modèles génétiques pour enquêter sur l&#39;implication de gènes spécifiques dans les comportements liés à l&#39;éthanol vers une meilleure compréhension de la base moléculaire de AUD. La mouche des fruits, Drosophila melanogaster, a émergé comme un organisme modèle leader pour explorer les mécanismes moléculaires génétique de comportements liés à l&#39;éthanol (examinés dans 3,4). Des études ont mis en évidence chez les mouches rôles pendant plusieurs voies de signalisation dans les réponses comportementales à l&#39;éthanol (examinés dans cinq). Curieusement, certains des gènes et les voies qui influencent responsab comportementaleses à l&#39;éthanol dans les mouches ont également été impliqués dans les comportements liés à l&#39;éthanol rongeurs et / ou AUD humain (par exemple, 6-14). La conservation des mécanismes de conduire les comportements liés à l&#39;éthanol à travers les espèces, couplés avec la suite d&#39;outils génétiques disponibles dans le système de modèle de drosophile, soulignent l&#39;utilité du modèle de la mouche des fruits pour étudier la génétique des réponses comportementales à l&#39;éthanol. Sensibilité 15,16 et la tolérance (revue dans 17) à l&#39;éthanol chez l&#39;homme est lié au développement de AUD. Ces deux réponses comportementales à l&#39;éthanol peut être modélisée en vol via une variété de tests de laboratoire (revue dans 3,4). Tous les tests de mouches connues des auteurs sont fondés sur des dépendant du temps induite par l&#39;éthanol sédation / incoordination ou la récupération en fonction du temps à partir d&#39;éthanol sédation. Dans un précédent article de notre groupe sur la génétique de la sensibilité à l&#39;éthanol et rla tolérance de apid chez la drosophile, un test de comportement à base d&#39;éthanol sédation induite vapeur de mouches a été utilisé 18. Les tests dans ce dosage a été initié en transférant adulte vivant vole sans anesthésie pour vider des flacons de produits alimentaires, le piégeage des mouches dans les flacons avec un bouchon en acétate de cellulose, l&#39;addition d&#39;éthanol à la partie supérieure (c.-à-face non volantes) de la bougie d&#39;acétate de cellulose, et sceller le flacon contenant les mouches, bouchon en acétate de cellulose et de l&#39;éthanol avec un bouchon de silicone (voir schéma de la figure S3, référence 18). Plusieurs flacons représentant différents groupes de mouches ont été évalués en parallèle, augmenter le débit de cet essai. Les flacons ont été donnés un code anonyme et expérimentateurs ont été aveuglés au groupe de traitement pour prévenir biais involontaire dans l&#39;évaluation de la sédation. Dans une expérience standard, les mouches dans des flacons ont été frappa doucement à intervalles de 6 min et, après une récupération de 30 secondes, le nombre de mouches sous sédation dans chaque flacon a été compté et converted pour cent vol actives. Les mouches absorbés vapeur d&#39;éthanol à partir de la prise d&#39;acétate de cellulose de façon dépendante du temps, entraînant des augmentations progressives dans l&#39;éthanol interne 18 et la sédation (cf de référence 18 et la figure 1A et 1B dans ce rapport). Sédation dans cet essai a été opérationnel défini comme mouches (i) permanent en l&#39;absence de la marche ou (ii) couché sur le dos avec ou sans battant des ailes. Ici, ce dosage de la sédation de l&#39;éthanol est décrite en détail, outre l&#39;optimisation opérationnelle pertinente à l&#39;utilisation, il est prévu, et le test est utilisé pour traiter la contribution d&#39;options de supplémentation alimentaire sur la sensibilité mouche de sédation.

<table border="0" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td>food vials</td>
			<td>VWR</td>
			<td>89092-772</td>
			<td>narrow</td>
		</tr>
		<tr>
			<td>Flugs</td>
			<td>Genesee/flystuff.com</td>
			<td>49-102</td>
			<td>narrow</td>
		</tr>
		<tr>
			<td>silicone stopper</td>
			<td>Fisher Scientific</td>
			<td>09-704-1l</td>
			<td>#4</td>
		</tr>
		<tr>
			<td>ethanol</td>
			<td>Pharmaco-Aaper</td>
			<td>111000200</td>
			<td>200 proof</td>
		</tr>
	</tbody>
</table>

an inexpensive, scalable behavioral assay for measuring ethanol sedation sensitivity and rapid tolerance in drosophila

trouble de consommation d&#39;alcool (AUD) est un grave problème de santé. Malgré une forte composante héréditaire à AUD, quelques gènes ont été clairement mis en cause dans leur étiologie. La mouche des fruits, Drosophila melanogaster, est un modèle puissant pour explorer les mécanismes moléculaires sous-jacents génétique des comportements liés à l&#39;alcool et détient donc une grande promesse pour l&#39;identification et la compréhension de la fonction des gènes qui influencent AUD. L&#39;utilisation du modèle de Drosophila pour ces types d&#39;études dépend de la disponibilité de dosages qui mesurent de façon fiable les réponses comportementales de l&#39;éthanol. Ce rapport décrit un dosage approprié pour évaluer la sensibilité de l&#39;éthanol et de la tolérance rapide de mouches. la sensibilité à l&#39;éthanol mesurée dans ce dosage est influencé par le volume et la concentration d&#39;éthanol utilisé, une variété de manipulations génétiques signalés précédemment, et également la durée de vol sont logés sans nourriture immédiatement avant l&#39;essai. En revanche, l&#39;éthanol sensitivity mesurée dans ce dosage ne est pas affectée par la manipulation de la vigueur à la mouche, sexe du vol, et de suppléments de milieu de croissance avec des antibiotiques ou des levures vivantes. Trois méthodes différentes pour quantifier la sensibilité de l&#39;éthanol sont décrits, tous conduisant à des résultats de sensibilité d&#39;éthanol essentiellement indiscernables. La nature évolutive de ce test, combiné avec sa simplicité globale de set-up et relativement faible coût, le rendent approprié pour petits et grands analyse génétique à grande échelle de la sensibilité de l&#39;éthanol et de la tolérance rapide chez la drosophile.

Les données brutes à partir de ce dosage de l&#39;éthanol sédation est le nombre de mouches qui sont sédation en fonction de la vapeur d&#39;éthanol temps d&#39;exposition. Les données brutes sont converties au pourcentage vol actif en fonction du temps (données primaires, les figures 1A, B, D - F). Sensibilité à la sédation éthanol à partir des données primaires peut être quantifiée comme sédation Temps 50 (ST50), le temps nécessaire pour 50% des mouches pour devenir ...

Watch this Scientific Journal Video about An Inexpensive, Scalable Behavioral Assay for Measuring Ethanol Sedation Sensitivity and Rapid Tolerance in Drosophila at JoVE.com

An Inexpensive, Scalable Behavioral Assay for Measuring Ethanol Sedation Sensitivity and Rapid Tolerance in Drosophila

Straightforward assays for measuring ethanol sensitivity and rapid tolerance in Drosophila facilitate the use of this model organism for investigating these important ethanol-related behaviors. Here, a relatively simple, scalable assay for measuring ethanol sensitivity and rapid tolerance in flies is described.

Un, test de comportement évolutives et économiques pour mesurer l&#39;éthanol sédation sensibilité et la tolérance rapide dans<em&gt; Drosophila</em

Alcohol use disorder (AUD) is a serious health challenge. Despite a large hereditary component to AUD, few genes have been unambiguously implicated in ...

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Research

JoVE Journal

Neuroscience

An Inexpensive, Scalable Behavioral Assay for Measuring Ethanol Sedation Sensitivity and Rapid Tolerance in Drosophila

10.2K vues.  Virginia Commonwealth University. Straightforward assays for measuring ethanol sensitivity and rapid tolerance in Drosophila facilitate the use of this model organism for investigating these important ethanol-related behaviors. Here, a relatively simple, scalable assay for measuring ethanol sensitivity and rapid tolerance in flies is described.

Vidéo: An Inexpensive, Scalable Behavioral Assay for Measuring Ethanol Sedation Sensitivity and Rapid Tolerance in Drosophila

Combining Behavioral Endocrinology and Experimental Economics: Testosterone and Social Decision Making

Behavioral endocrinological research in humans as well as in animals suggests that testosterone plays a key role in social interactions. Studies in rodents have shown a direct link between testosterone and aggressive behavior1 and folk wisdom adapts these findings to humans, suggesting that testosterone induces antisocial, egoistic or even aggressive behavior2. However, many researchers doubt a direct testosterone-aggression link in humans, arguing instead that testosterone is primarily involved in status-related behavior3,4. As a high status can also be achieved by aggressive and antisocial means it can be difficult to distinguish between anti-social and status seeking behavior.
We therefore set up an experimental environment, in which status can only be achieved by prosocial means. In a double-blind and placebo-controlled experiment, we administered a single sublingual dose of 0.5 mg of testosterone (with a hydroxypropyl-&beta;-cyclodextrin carrier) to 121 women and investigated their social interaction behavior in an economic bargaining paradigm. Real monetary incentives are at stake in this paradigm; every player A receives a certain amount of money and has to make an offer to another player B on how to share the money. If B accepts, she gets what was offered and player A keeps the rest. If B refuses the offer, nobody gets anything. A status seeking player A is expected to avoid being rejected by behaving in a prosocial way, i.e. by making higher offers.
The results show that if expectations about the hormone are controlled for, testosterone administration leads to a significant increase in fair bargaining offers compared to placebo. The role of expectations is reflected in the fact that subjects who report that they believe to have received testosterone make lower offers than those who say they believe that they were treated with a placebo. These findings suggest that the experimental economics approach is sensitive for detecting neurobiological effects as subtle as those achieved by administration of hormones. Moreover, the findings point towards the importance of both psychosocial as well as neuroendocrine factors in determining the influence of testosterone on human social behavior.

The procedure described in this protocol shows that testosterone administration and folk beliefs about testosterone may be associated with directly opposed social behaviors.

Combinant endocrinologie du comportement et l&#39;économie expérimentale: Décision de la testostérone et social Rendre

Behavioral endocrinological research in humans as well as in animals suggests that testosterone plays a key role in social interactions. Studies in ...

Recherche

Neurosciences

A Simple Way to Measure Ethanol Sensitivity in Flies

Low doses of ethanol cause flies to become hyperactive, while high doses are sedating. The sensitivity to ethanol-induced sedation of a given fly strain is correlated with that strain s ethanol preference, and therefore sedation is a highly relevant measure to study the genetics of alcohol responses and drinking. We demonstrate a simple way to expose flies to ethanol and measure its intoxicating effects. The assay we describe can determine acute sensitivity, as well as ethanol tolerance induced by repeat exposure. It does not require a technically involved setup, and can therefore be applied in any laboratory with basic fly culture tools.

A simple assay to measure the sedating effects of ethanol on Drosophila flies, based on the loss of righting reflex, is described.

Une façon simple de mesurer la sensibilité d&#39;éthanol dans les mouches

Low doses of ethanol cause flies to become hyperactive, while high doses are sedating. The sensitivity to ethanol-induced sedation of a given fly strain ...

A General Method for Evaluating Incubation of Sucrose Craving in Rats

For someone on a food-restricted diet, food craving in response to food-paired cues may serve as a key behavioral transition point between abstinence and relapse to food taking 1. Food craving conceptualized in this way is akin to drug craving in response to drug-paired cues. A rich literature has been developed around understanding the behavioral and neurobiological determinants of drug craving; we and others have been focusing recently on translating techniques from basic addiction research to better understand addiction-like behaviors related to food 2-4.
As done in previous studies of drug craving, we examine sucrose craving behavior by utilizing a rat model of relapse. In this model, rats self-administer either drug or food in sessions over several days. In a session, lever responding delivers the reward along with a tone+light stimulus. Craving behavior is then operationally defined as responding in a subsequent session where the reward is not available. Rats will reliably respond for the tone+light stimulus, likely due to its acquired conditioned reinforcing properties 5. This behavior is sometimes referred to as sucrose seeking or cue reactivity. In the present discussion we will use the term "sucrose craving" to subsume both of these constructs.
In the past decade, we have focused on how the length of time following reward self-administration influences reward craving. Interestingly, rats increase responding for the reward-paired cue over the course of several weeks of a period of forced-abstinence. This "incubation of craving" is observed in rats that have self-administered either food or drugs of abuse 4,6. This time-dependent increase in craving we have identified in the animal model may have great potential relevance to human drug and food addiction behaviors.
Here we present a protocol for assessing incubation of sucrose craving in rats. Variants of the procedure will be indicated where craving is assessed as responding for a discrete sucrose-paired cue following extinction of lever pressing within the sucrose self-administration context (Extinction without cues) or as responding for sucrose-paired cues in a general extinction context (Extinction with cues).

Responding for food or drug-paired cues increases over the course of a period of abstinence and this may relate to an increased susceptibility to relapse behaviors. Here we detail a procedure for evaluating this "incubation of craving" in rats that have self-administered sucrose.

Une méthode générale pour l&#39;évaluation d&#39;incubation du désir saccharose chez les rats

For someone on a food-restricted diet, food craving in response to food-paired cues may serve as a key behavioral transition point between abstinence and ...

Measuring Changes in Tactile Sensitivity in the Hind Paw of Mice Using an Electronic von Frey Apparatus

Measuring inflammation-induced changes in thresholds of hind paw withdrawal from mechanical pressure is a useful technique to assess changes in pain perception in rodents. Withdrawal thresholds can be measured first at baseline and then following drug, venom, injury, allergen,&#160;or otherwise evoked inflammation by applying an accurate force on very specific areas of the skin. An electronic von Frey apparatus allows precise assessment of mouse hind paw withdrawal thresholds that are not limited by the available filament sizes in contrast to classical von Frey measurements. The ease and rapidity of measurements allow for incorporation of assessment of tactile sensitivity outcomes in diverse models of rapid-onset inflammatory and neuropathic pain as multiple measurements can be taken within a short time period. Experimental measurements for individual rodent subjects can be internally controlled against individual baseline responses and exclusion criteria easily established to standardize baseline responses within and across experimental groups. Thus, measurements using an electronic von Frey apparatus represent a useful modification of the well-established classical von Frey filament-based assays for rodent mechanical allodynia that may also be applied to other nonhuman mammalian models.

Electronic von Frey measurements are an effective and improved alternative to classical von Frey filaments, as they allow rapid and precise measurement of changes in rodent mechanical withdrawal thresholds to continuously applied pressure stimuli before and after an inflammatory event.

Measuring inflammation-induced changes in thresholds of hind paw withdrawal from mechanical pressure is a useful technique to assess changes in pain ...

A Simple and Inexpensive Method for Determining Cold Sensitivity and Adaptation in Mice

Cold hypersensitivity is a serious clinical problem, affecting a broad subset of patients and causing significant decreases in quality of life. The cold plantar assay allows the objective and inexpensive assessment of cold sensitivity in mice, and can quantify both analgesia and hypersensitivity. Mice are acclimated on a glass plate, and a compressed dry ice pellet is held against the glass surface underneath the hindpaw. The latency to withdrawal from the cooling glass is used as a measure of cold sensitivity.
Cold sensation is also important for survival in regions with seasonal temperature shifts, and in order to maintain sensitivity animals must be able to adjust their thermal response thresholds to match the ambient temperature. The Cold Plantar Assay (CPA) also allows the study of adaptation to changes in ambient temperature by testing the cold sensitivity of mice at temperatures ranging from 30 &#176;C to 5 &#176;C. Mice are acclimated as described above, but the glass plate is cooled to the desired starting temperature using aluminum boxes (or aluminum foil packets) filled with hot water, wet ice, or dry ice. The temperature of the plate is measured at the center using a filament T-type thermocouple probe. Once the plate has reached the desired starting temperature, the animals are tested as described above.
This assay allows testing of mice at temperatures ranging from innocuous to noxious. The CPA yields unambiguous and consistent behavioral responses in uninjured mice and can be used to quantify both hypersensitivity and analgesia. This protocol describes how to use the CPA to measure cold hypersensitivity, analgesia, and adaptation in mice.

The Cold Plantar Assay (CPA) measures cold responsiveness between 30 &#176;C and 5 &#176;C, and can also measure cold adaptation. This protocol describes how to use the CPA to measure cold hypersensitivity, analgesia, and adaptation in mice.

Une méthode simple et peu coûteuse pour déterminer Sensibilité au froid et adaptation chez la souris

Cold hypersensitivity is a serious clinical problem, affecting a broad subset of patients and causing significant decreases in quality of life.  The cold ...

An Assay for Measuring the Effects of Ethanol on the Locomotion Speed of Caenorhabditis elegans

Alcohol use disorders are a significant public health concern, for which there are few effective treatment strategies. One difficulty that has delayed the development of more effective treatments is the relative lack of understanding of the molecular underpinnings of the effects of ethanol on behavior. The nematode, Caenorhabditis elegans (C.&#160;elegans), provides a useful model in which to generate and test hypotheses about the molecular effects of ethanol. Here, we describe an assay that has been developed and used to examine the roles of particular genes and environmental factors in behavioral responses to ethanol, in which locomotion is the behavioral output. Ethanol dose-dependently causes an acute depression of crawling on an agar surface. The effects are dynamic; animals exposed to a high concentration demonstrate an initial strong depression of crawling, referred to here as initial sensitivity, and then partially recover locomotion speed despite the continued presence of the drug. This ethanol-induced behavioral plasticity is referred to here as the development of acute functional tolerance. This assay has been used to demonstrate that these two phenotypes are distinct and genetically separable. The straightforward locomotion assay described here is suitable for examining the effects of both genetic and environmental manipulations on these acute behavioral responses to ethanol in C.&#160;elegans.

An Assay for Measuring the Effects of Ethanol on the Locomotion Speed of Caenorhabditis elegans

C. elegans is a useful model for studying the effects of ethanol on behavior. We present a behavioral assay that quantifies the effects of ethanol on the locomotion speed of crawling worms; both initial sensitivity and the development of acute functional tolerance to ethanol can be measured with this assay.

Un test pour mesurer les effets de l&#39;éthanol sur la vitesse de Locomotion<em&gt; Caenorhabditis elegans</em

Alcohol use disorders are a significant public health concern, for which there are few effective treatment strategies. One difficulty that has delayed the ...

A Behavioral Assay for Mechanosensation of MARCM-based Clones in Drosophila melanogaster

Because of the structural and functional homology to the hair cells of the mammalian inner ear, the neurons that innervate the Drosophila external sense organs provide an excellent model system for the study of mechanosensation. This protocol describes a simple touch behavior in fruit flies which can be used to identify mutations that interfere with mechanosensation. The tactile stimulation of a macrochaete bristle on the thorax of flies elicits a grooming reflex from either the first or third leg. Mutations that interfere with mechanotransduction (such as NOMPC), or with other aspects of the reflex arc, can inhibit the grooming response. A traditional screen of adult behaviors would have missed mutants that have essential roles during development. Instead, this protocol combines the touch screen with mosaic analysis with a repressible cell marker (MARCM) to allow for only limited regions of homozygous mutant cells to be generated and marked by the expression of green fluorescent protein (GFP). By testing MARCM clones for abnormal behavioral responses, it is possible to screen a collection of lethal p-element mutations to search for new genes involved in mechanosensation that would have been missed by more traditional methods.

A Behavioral Assay for Mechanosensation of MARCM-based Clones in Drosophila melanogaster

In order to identify novel mutations affecting mechanosensation, we designed an assay that measures the behavioral response to tactile stimulation of fly bristles in mutant clones generated by the MARCM method. The combination of techniques allows for the identification of mechanosensitive mutations that would otherwise be lethal.

A Behavioral Essai pour mécanosensibilité de marcm-fondé clones dans<em&gt; Drosophila melanogaster</em

Because of the structural and functional homology to the hair cells of the mammalian inner ear, the neurons that innervate the Drosophila external sense ...

Electrophysiological Method for Recording Intracellular Voltage Responses of Drosophila Photoreceptors and Interneurons to Light Stimuli In Vivo

Voltage responses of insect photoreceptors and visual interneurons can be accurately recorded with conventional sharp microelectrodes. The method described here enables the investigator to measure long-lasting (from minutes to hours) high-quality intracellular responses from single Drosophila R1-R6 photoreceptors and Large Monopolar Cells (LMCs) to light stimuli. Because the recording system has low noise, it can be used to study variability among individual cells in the fly eye, and how their outputs reflect the physical properties of the visual environment. We outline all key steps in performing this technique. The basic steps in constructing an appropriate electrophysiology set-up for recording, such as design and selection of the experimental equipment are described. We also explain how to prepare for recording by making appropriate (sharp) recording and (blunt) reference electrodes. Details are given on how to fix an intact fly in a bespoke fly-holder, prepare a small window in its eye and insert a recording electrode through this hole with minimal damage. We explain how to localize the center of a cell's receptive field, dark- or light-adapt the studied cell, and to record its voltage responses to dynamic light stimuli. Finally, we describe the criteria for stable normal recordings, show characteristic high-quality voltage responses of individual cells to different light stimuli, and briefly define how to quantify their signaling performance. Many aspects of the method are technically challenging and require practice and patience to master. But once learned and optimized for the investigator's experimental objectives, it grants outstanding in vivo neurophysiological data.

Electrophysiological Method for Recording Intracellular Voltage Responses of Drosophila Photoreceptors and Interneurons to Light Stimuli In Vivo

Sharp microelectrodes enable accurate electrophysiological characterization of photoreceptor and visual interneuron output in living Drosophila. Here we show how to use this method to record high-quality voltage responses of individual cells to controlled light stimulation. This method is ideal for studying neural information processing in insect compound eyes.

Méthode électrophysiologique de réponses d&#39;enregistrement intracellulaire tension de<em&gt; Drosophila</em&gt; Photorécepteurs et Interneurones à Light Stimuli<em&gt; In Vivo</em

Voltage responses of insect photoreceptors and visual interneurons can be accurately recorded with conventional sharp microelectrodes. The method ...

Measuring and Altering Mating Drive in Male Drosophila melanogaster

Despite decades of investigation, the neuronal and molecular bases of motivational states remain mysterious. We have recently developed a novel, reductionist, and scalable system for in-depth investigation of motivation using the mating drive of male Drosophila&#160;melanogaster (Drosophila), the methods for which we detail here. The behavioral paradigm centers on the finding that male mating drive decreases alongside fertility over the course of repeated copulations and recovers over ~3 d. In this system, the powerful neurogenetic tools available in the fly converge with the genetic accessibility and putative wiring diagram available for sexual behavior. This convergence allows rapid isolation and interrogation of small neuronal populations with specific motivational functions. Here we detail the design and execution of the satiety assay that is used to measure and alter courtship motivation in the male fly. Using this assay, we also demonstrate that low male mating drive can be overcome by stimulating dopaminergic neurons. The satiety assay is simple, affordable, and robust to influences of genetic background. We expect the satiety assay to generate many new insights into the neurobiology of motivational states.

Measuring and Altering Mating Drive in Male Drosophila melanogaster

This article describes a behavioral assay that uses male mating drive in Drosophila melanogaster to study motivation. Using this method, researchers can utilize advanced fly neurogenetic techniques to uncover the genetic, molecular, and cellular mechanisms that underlie this motivation.

Mesure et Altération Mating Drive à Male<em&gt; Drosophila melanogaster</em

Despite decades of investigation, the neuronal and molecular bases of motivational states remain mysterious. We have recently developed a novel, ...

Identification of Dopamine D1-Alpha Receptor Within Rodent Nucleus Accumbens by an Innovative RNA In Situ Detection Technology

In the central nervous system, the D1-alpha subtype receptor (Drd1&#945;) is the most abundant dopamine (DA) receptor, which plays a vital role in regulating neuronal growth and development. However, the mechanisms underlying Drd1&#945; receptor abnormalities mediating behavioral responses and modulating working memory function are still unclear. Using a novel RNA in situ hybridization assay, the current study identified dopamine Drd1&#945; receptor and tyrosine hydroxylase (TH) RNA expression from DA-related circuitry in the nucleus accumbens (NAc) area and substantia nigra region (SNR), respectively. Drd1&#945; expression in the NAc shows a &#34;discrete dot&#34; staining pattern. Clear sex differences in Drd1&#945; expression were observed. In contrast, TH shows a &#34;clustered&#34; staining pattern. Regarding TH expression, female rats displayed a higher signal expression per cell relative to male animals. The methods presented here provide a novel in situ hybridization technique for investigating changes in dopamine system dysfunction during the progression of central nervous system diseases.

Identification of Dopamine D1-Alpha Receptor Within Rodent Nucleus Accumbens by an Innovative RNA In Situ Detection Technology

Identification of dopamine D1-alpha receptor in the nucleus accumbens is critical for clarifying D1 receptor dysfunction during a central nervous system disease. We performed a novel RNA in situ hybridization assay to visualize single RNA molecules in a specific brain area.

Identification de la Dopamine D1-Alpha du récepteur au sein de rongeurs noyau Accumbens par une ARN innovantes In Situ la technologie de détection