We thank the Duke Physics Shop, especially Phil Lewis, for helping us build the behavioral apparatus and attachments and creating the drawing. This work is funded by the National Institutes of Health under award number R01GM100027.

1. Preparación de moscas a ensayar <ol><li> Cultura vuela de melaza estándar / media de harina de maíz en incubadora a 25 ° C y 65% ​​de humedad. Tenga cuidado de no saturar los viales. Por ejemplo, poner 8 mujeres y 6 hombres en un vial de alimentos estrecho. Nota: El &quot;estrecho vial comida&quot; usado aquí tiene un diámetro interior de 2,3 cm. Nosotros generalmente pueden obtenerse unos 10 ml de alimentos mosca en cada vial. La receta de comida mosca hemos utilizado se describe aquí: http://flystocks.bio.indiana.edu/Fly_Work/media-recipes/molassesfood.html. </li><li> 2 - 3 días después de la eclosión hembras, preparar viales con pasta de levadura fresca como se muestra en la Figura 1E. Hacer pasta de levadura fresca mediante la mezcla de 6 g de levadura activa con 10 ml de ácido propiónico 0,5%. Utilice una espátula para aplicar la pasta de levadura sobre la pared lateral de los viales de alimentos. Recoger 30 - 35 hembras eclosed junto con 20 - 25 machos en los viales. <ol><li> En general, recoger los machos de las mismas viales donde se recogen las hembras(Con el fin de ahorrar tiempo y esfuerzo). Sin embargo, si hay preocupación acerca de la fertilidad de los machos, machos WT utilizar en su lugar. Además, la pasta de levadura es importante para estimular la producción de huevos, por lo tanto prepararlo fresco todos los días. </li></ol></li><li> Mantener las hembras / machos recolectados a 25 ° C, 65% de humedad, a menos que las moscas son sensibles a la temperatura. </li><li> Después de ~ 4 - 5 días, consulte los viales para ver si las hembras están listas para experimentos que ponen huevos. Están listos cuando la superficie del medio de comida es mojado de larvas madriguera activamente en la comida (véase la Figura 1E - F). Nota: Las hembras normalmente retienen poner huevos una vez que la superficie del alimento se humedece y se arrastra con larvas. Este paso asegura que las mujeres estarían dispuestas a poner muchos huevos cuando se encuentran con sustratos con textura deseada (por ejemplo., ~ 1% de agarosa). </li></ol> 2. Cámara de la Construcción, montaje, instalación de Ensayo <ol><li> Tener un taller mecánico construir cámaras de la puesta de huevos de acrílico (Fifigura 1A - D). El dibujo de ingeniería de diferentes piezas se muestra en la Figura 1 complementario -. 3 imágenes de alta resolución también se pueden encontrar aquí (http://www.rebeccayang.org/pdf/chamber%20design.pdf). </li><li> Inserte láminas de plástico en la carga (arriba) pieza de la cámara como se muestra en la Figura 1C. Esto sirve como la superficie inferior de carga, mientras que vuela en los ámbitos individuales que ponen huevos. </li><li> Anestesiar a las hembras en un cojín de CO 2 y cargarlos individualmente en cada campo de la puesta de huevos. Permitir ~ 30 minutos para que las moscas se recuperan de las emisiones de CO 2 y se aclimate al nuevo entorno. </li><li> Preparar los sustratos de agarosa. <ol><li> Para mayor comodidad, tenga una botella de prefabricados de fundido de agarosa al 1% en un baño de agua a 55 ° C. </li><li> Añadir la cantidad deseada de solución de sacarosa (2 M) en un tubo cónico de 50 ml y se mezcla con la cantidad apropiada de agarosa. Por ejemplo, para preparar el 150 mM sustrato sacarosa, place 750 l de solución 2 M de sacarosa en el tubo y luego llenar el tubo con agarosa para la marca de 10 ml. </li><li> Preparar el sustrato liso de la misma manera pero añadir agua destilada en lugar de solución de sacarosa. Nota: La concentración final de agarosa en este protocolo es ligeramente menor que 1%. En nuestra experiencia, la concentración exacta de agarosa no importa con tal de que se controla para estar dentro de ~ 0,9 a 1,1% y que los dos sustratos son de la misma concentración de agarosa. </li></ol></li><li> Tomar la pieza de sustrato (parte inferior) de la cámara y la pipeta 1.000 l de sustrato de agarosa en cada cubeta como se ve en la Figura 1D. </li><li> Permitir agarosa para solidificar durante ~ 30 min. </li><li> Una vez que los sustratos de agarosa y las moscas están listos, el montaje de las tres piezas de la cámara de la puesta de huevos y luego sacar las láminas de plástico. </li><li> Colocar las cámaras en las incubadoras de moscas. Nota: La longitud de los experimentos que ponen huevos puede variar dependiendo de experimentales necesidades. normalmente se corre el experimento S / N (14 - 16 h). Además, no se observó efecto significativo de tiempo circadiano de las preferencias de la puesta de huevos. </li><li> Anestesiar a las hembras mediante la inyección de CO 2 en la cámara. Desmonte la cámara, desechar las moscas anestesiadas en la morgue mosca (es decir., Una lata vacía de café llena con un poco de aceite de maíz). Tome fotografías de los resultados de gestión de registros (véase la Figura 2). </li><li> Contar el número de huevos de forma manual y calcular los índices de preferencia para su análisis. Calcular el índice de preferencia como (N a - N b) / (N + un N b) donde N A y N b representan el número de huevos en el sitio de un sitio vs. b, respectivamente. </li></ol>

<ol>
	<li>Perisse, E., Burke, C., Huetteroth, W., Waddell, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Shocking+revelations+and+saccharin+sweetness+in+the+study+of+Drosophila+olfactory+memory.">Shocking revelations and saccharin sweetness in the study of Drosophila olfactory memory.</a> Curr. Biol. 23 (17), 752-763 (2013).</li><li>Yamamoto, N. C., Koganezawa, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genes+and+circuits+of+courtship+behaviour+in+Drosophila+males.">Genes and circuits of courtship behaviour in Drosophila males.</a> Nat. Rev. Neurosci. 14 (10), 681-692 (2013).</li><li>Zwarts, L., Versteven, M., Callaerts, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetics+and+neurobiology+of+aggression+in+Drosophila.">Genetics and neurobiology of aggression in Drosophila.</a> Fly. 6 (1), 35-48 (2012).</li><li>Joseph, R. M., Devineni, A. V., King, I. F., Heberlein, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oviposition+preference+for+and+positional+avoidance+of+acetic+acid+provide+a+model+for+competing+behavioral+drives+in+Drosophila.">Oviposition preference for and positional avoidance of acetic acid provide a model for competing behavioral drives in Drosophila.</a> Proc. Natl. Acad. Sci. 106 (27), 11352-11357 (2009).</li><li>Miller, P. M., Saltz, J. B., Cochrane, V. A., Marcinkowski, C. M., Mobin, R., Turner, T. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Natural+variation+in+decision-making+behavior+in+Drosophila+melanogaster.">Natural variation in decision-making behavior in Drosophila melanogaster.</a> PLoS One. 6 (1), 16436 (2011).</li><li>Schwartz, N. U., Zhong, L., Bellemer, A., Tracey, W. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Egg-laying+decisions+in+Drosophila+are+consistent+with+foraging+costs+of+larval+progeny.">Egg-laying decisions in Drosophila are consistent with foraging costs of larval progeny.</a> PloS One. 7 (5), 37910 (2012).</li><li>Yang, C. H., Belawat, P., Hafen, E., Jan, L. Y., Jan, Y. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+egg-laying+site+selection+a+system+to+study+simple+decision-making+processes.">Drosophila egg-laying site selection a system to study simple decision-making processes.</a> Science. 319 (5870), 1679-1683 (2008).</li><li>Kacsoh, B. Z., Lynch, Z. R., Mortimer, N. T., Schlenke, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fruit+flies+medicate+offspring+after+seeing+parasites.">Fruit flies medicate offspring after seeing parasites.</a> Science. 339 (6122), 947-950 (2013).</li><li>Wu, C. -. L., Fu, T. -. F., Chou, Y. -. Y., Yeh, S. -. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Single+Pair+of+Neurons+Modulates+Egg-Laying+Decisions+in+Drosophila.">A Single Pair of Neurons Modulates Egg-Laying Decisions in Drosophila.</a> PloS one. 10 (3), e0121335 (2015).</li><li>Yang, C. H., He, R., Stern, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavioral+and+circuit+basis+of+sucrose+rejection+by+Drosophila+females+in+a+simple+decision-making+task.">Behavioral and circuit basis of sucrose rejection by Drosophila females in a simple decision-making task.</a> J. Neurosci. 35 (4), 1396-1410 (2015).</li><li>Gou, B., Liu, Y., Guntur, A. R., Stern, U., Yang, C. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanosensitive+neurons+on+the+internal+reproductive+tract+contribute+to+egg-laying+induced+acetic+acid+attraction+in+Drosophila.">Mechanosensitive neurons on the internal reproductive tract contribute to egg-laying induced acetic acid attraction in Drosophila.</a> Cell Rep. 9 (2), 522-530 (2014).</li><li>Stern, U., Zhu, E. Y., He, R., Yang, C. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-duration+animal+tracking+in+difficult+lighting+conditions.">Long-duration animal tracking in difficult lighting conditions.</a> Sci. Rep. 5, 10432 (2015).</li><li>Zhu, E. Y., Guntur, A. R., He, R., Stern, U., Yang, C. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Egg-laying+demand+induces+aversion+of+UV+light+in+Drosophila+females.">Egg-laying demand induces aversion of UV light in Drosophila females.</a> Curr. Biol. 24 (23), (2014).</li><li>Branson, K., Robie, A. A., Bender, J., Perona, P., Dickinson, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-throughput+ethomics+in+large+groups+of+Drosophila.">High-throughput ethomics in large groups of Drosophila.</a> Nat. Methods. 6, 451-457 (2009).</li><li>Klatpoetke, N. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Independent+Optical+Excitation+of+Distinct+Neural+Populations.">Independent Optical Excitation of Distinct Neural Populations.</a> Nat. Methods. 11 (3), 338-346 (2014).</li></ol>

The authors have nothing to disclose.

Las cámaras y los protocolos descritos aquí tienen varias mejoras con respecto a los ensayos de la puesta de huevos anteriores. En primer lugar, que aumentan el rendimiento del ensayo de las preferencias de los animales individuales de manera significativa. Cada cámara puede ensayar 30 hembras individuales y se tarda menos de una hora de configurar. En segundo lugar, aumentan la consistencia de las preferencias de la puesta de huevos respecto a los métodos anteriores. La estandarización de las dimensiones de la are...

Drosophila melanogaster is a powerful genetic model organism to study the neural basis of behaviors. The rapid developments of genetic tools to manipulate neurons in a targeted manner and the emergence of sophisticated behavioral analysis tools have significantly improved our ability to dissect the circuit mechanisms that underlie the sensory-motor transformation processes of several innate and learned behaviors1-3.
Drosophila egg-laying is a suitable model to study the neural basis of simple decision making processes. In particular, Drosophila females have been shown to possess the ability to compare and rank their options before "committing" to depositing an egg onto a given option4-8. For example, when given only a plain (sucrose-free) substrate or only a sucrose-containing substrate, females readily accept either option for egg-laying. However, when presented with both options, females robust reject the sucrose substrate in some contexts7,9,10. Relatively little is known about the neural mechanisms that allow females to "choose the greater of two goods", however. A major obstacle has been the lack of an efficient method to assay egg-laying preferences such that one can use a systematic genetic screening approach to study this problem.
In this report, we describe the protocol we developed that allows egg-laying preferences of females to be assayed at single-animal resolution and with substantially improved throughput and consistency over previous methods. Specifically, we provide the blueprints for constructing the chambers we designed, the protocol for preparing the females so that each is primed to lay many eggs, and the protocol for using the chambers.

<table><tbody><tr><td>UltraPure Agarose&amp;nbsp;</td><td>Invitrogen</td><td>16500-500</td><td></td></tr><tr><td>Sucrose</td><td>Sigma</td><td>S0389</td><td></td></tr><tr><td>Water bath&amp;nbsp;</td><td>Fisher</td><td>15-462-6Q</td><td></td></tr><tr><td>LifeCam Cinema webcam</td><td>Microsoft</td><td>H5D-00013</td><td></td></tr><tr><td>Red LEDs</td><td>Cree</td><td>C503B-RAN-CA0B0AA1</td><td></td></tr><tr><td>Egg-laying chambers</td><td>Custom Built</td><td></td><td></td></tr><tr><td>Camera holders</td><td>Custom Built</td><td></td><td></td></tr><tr><td>LED holders</td><td>Custom Built</td><td></td><td></td></tr><tr><td>Fly vials (narrow)</td><td>Genesee</td><td>32-116BC</td><td></td></tr></tbody></table>

high throughput assay to examine egg-laying preferences of individual drosophila melanogaster

Recientemente, la preferencia de la puesta de huevos de Drosophila se ha convertido en un modelo manejable de forma genética para estudiar la base neural de los procesos de toma de decisiones simples. Cuando la selección de sitios para depositar sus huevos, las moscas hembras son capaces de clasificar el atractivo relativo de sus opciones y elegir el &quot;mayor de dos bienes.&quot; Sin embargo, la mayoría de los ensayos de preferencia que ponen huevos no son prácticos si se quiere adoptar un enfoque sistemático para la detección genética buscar la base de circuito que subyace a este sencillo proceso de toma de decisiones, ya que son laboriosos y configurar basado en la población. Para aumentar el rendimiento de estudiar las preferencias puesta de huevos de las hembras individuales, desarrollamos cámaras personalizadas que cada uno puede al mismo tiempo de ensayo preferencias de la puesta de huevos de hasta treinta individuo moscas, así como un protocolo que asegura cada mujer tiene una alta tasa de puesta de huevos (de modo que su preferencia es fácilmente discernible y más convincente). Nuestro enfoque es sencillo de ejecutary produce resultados muy coherentes. Además, estas cámaras pueden equiparse con diferentes accesorios para permitir la grabación de vídeo a los animales ovíparos y para suministrar luz para los estudios de la optogenética. Este artículo proporciona los planos para la fabricación de estas cámaras y el procedimiento para la preparación de las moscas para ser ensayadas en estas cámaras.

Las cámaras de la puesta de huevos se componen de varias piezas: una pieza de sustrato (parte inferior), una pieza divisora ​​(centro), una pieza de carga (parte superior), y 2 puertas correderas (Figura 1 A - D). Estas piezas se utilizan para moscas de forma independiente de configuración y los sustratos antes de los experimentos que ponen huevos. Figura 1F muestra cómo viales deben buscar cuando las moscas hembras están listas para la puesta de...

Watch this Scientific Journal Video about High Throughput Assay to Examine Egg-Laying Preferences of Individual Drosophila melanogaster at JoVE.com

High Throughput Assay to Examine Egg-Laying Preferences of Individual Drosophila melanogaster

Ensayo de Alto Rendimiento para el Examen de Preferencias de la puesta de huevos individual<em&gt; Drosophila melanogaster</em

This protocol describes a high throughput assay for testing egg-laying preferences of Drosophila melanogaster at single-animal resolution. This assay provides a simple, efficient, and scalable platform to identify genes and circuit components that control a simple decision-making process.

Recently, egg-laying preference of Drosophila has emerged as a genetically tractable model to study the neural basis of simple decision-making processes. ...

high-throughput-assay-to-examine-egg-laying-preferences-individual

Research

JoVE Journal

Neuroscience

High Throughput Assay to Examine Egg-Laying Preferences of Individual Drosophila melanogaster

7.9K vistas.  Duke University. This protocol describes a high throughput assay for testing egg-laying preferences of Drosophila melanogaster at single-animal resolution. This assay provides a simple, efficient, and scalable platform to identify genes and circuit components that control a simple decision-making process.

Video: High Throughput Assay to Examine Egg-Laying Preferences of Individual Drosophila melanogaster

Methods to Assay Drosophila Behavior

Drosophila melanogaster, the fruit fly, has been used to study molecular mechanisms of a wide range of human diseases such as cancer, cardiovascular disease and various neurological diseases1. We have optimized simple and robust behavioral assays for determining larval locomotion, adult climbing ability (RING assay), and courtship behaviors of Drosophila. These behavioral assays are widely applicable for studying the role of genetic and environmental factors on fly behavior. Larval crawling ability can be reliably used for determining early stage changes in the crawling abilities of Drosophila larvae and also for examining effect of drugs or human disease genes (in transgenic flies) on their locomotion. The larval crawling assay becomes more applicable if expression or abolition of a gene causes lethality in pupal or adult stages, as these flies do not survive to adulthood where they otherwise could be assessed. This basic assay can also be used in conjunction with bright light or stress to examine additional behavioral responses in Drosophila larvae. Courtship behavior has been widely used to investigate genetic basis of sexual behavior, and can also be used to examine activity and coordination, as well as learning and memory. Drosophila courtship behavior involves the exchange of various sensory stimuli including visual, auditory, and chemosensory signals between males and females that lead to a complex series of well characterized motor behaviors culminating in successful copulation. Traditional adult climbing assays (negative geotaxis) are tedious, labor intensive, and time consuming, with significant variation between different trials2-4. The rapid iterative negative geotaxis (RING) assay5 has many advantages over more widely employed protocols, providing a reproducible, sensitive, and high throughput approach to quantify adult locomotor and negative geotaxis behaviors. In the RING assay, several genotypes or drug treatments can be tested simultaneously using large number of animals, with the high-throughput approach making it more amenable for screening experiments.

Methods to Assay Drosophila Behavior

Drosophila melanogaster is a genetically and behaviorally tractable model system that has been used to understand the molecular and cellular basis of many important biological processes for over a century 1. Drosophila has been well exploited to gain insights into the genetic basis of fly behavior.

Métodos de ensayo a Drosophila Comportamiento

Drosophila melanogaster, the fruit fly, has been used to study molecular mechanisms of a wide range of human diseases such as cancer, cardiovascular ...

Investigación

Neurociencias

Light Preference Assay to Study Innate and Circadian Regulated Photobehavior in Drosophila Larvae

Light acts as environmental signal to control animal behavior at various levels. The Drosophila larval nervous system is used as a unique model to answer basic questions on how light information is processed and shared between rapid and circadian behaviors. Drosophila larvae display a stereotypical avoidance behavior when exposed to light. To investigate light dependent behaviors comparably simple light-dark preference tests can be applied. In vertebrates and arthropods the neural pathways involved in sensing and processing visual inputs partially overlap with those processing photic circadian information. The fascinating question of how the light sensing system and the circadian system interact to keep behavioral outputs coordinated remains largely unexplored. Drosophila is an impacting biological model to approach these questions, due to a small number of neurons in the brain and the availability of genetic tools for neuronal manipulation. The presented light-dark preference assay allows the investigation of a range of visual behaviors including circadian control of phototaxis.

Light Preference Assay to Study Innate and Circadian Regulated Photobehavior in Drosophila Larvae

Here we describe a light-dark preference test for Drosophila larva. This assay provides information about innate and circadian regulation of light sensing and processing photobehavior.

Ensayo Preferencia luz para estudiar innata y circadiano Photobehavior Regulado de<em&gt; Drosophila</em&gt; Las larvas

Light acts as environmental signal to control animal behavior at various levels. The Drosophila larval nervous system is used as a unique model to answer ...

A Single-fly Assay for Foraging Behavior in Drosophila

For many animals, hunger promotes changes in the olfactory system in a manner that facilitates the search for appropriate food sources. In this video article, we describe an automated assay to measure the effect of hunger or satiety on olfactory dependent food search behavior in the adult fruit fly Drosophila melanogaster. In a light-tight box illuminated by red light that is invisible to fruit flies, a camera linked to custom data acquisition software monitors the position of six flies simultaneously. Each fly is confined to walk in individual arenas containing a food odor at the center. The testing arenas rest on a porous floor that functions to prevent odor accumulation. Latency to locate the odor source, a metric that reflects olfactory sensitivity under different physiological states, is determined by software analysis. Here, we discuss the critical mechanics of running this behavioral paradigm and cover specific issues regarding fly loading, odor contamination, assay temperature, data quality, and statistical analysis.

A Single-fly Assay for Foraging Behavior in Drosophila

In this video article, we describe an automated assay to measure the effect of hunger or satiety on olfactory dependent food search behavior in the adult fruit fly Drosophila melanogaster.

Un ensayo de una sola mosca por el comportamiento de forrajeo en<em&gt; Drosophila</em

For many animals, hunger promotes changes in the olfactory system in a manner that facilitates the search for appropriate food sources. In this video ...

Assay for Blood-brain Barrier Integrity in Drosophila melanogaster

Proper nervous system development includes the formation of the blood-brain barrier, the diffusion barrier that tightly regulates access to the nervous system and protects neural tissue from toxins and pathogens. Defects in the formation of this barrier have been correlated with neuropathies, and the breakdown of this barrier has been observed in many neurodegenerative diseases. Therefore, it is critical to identify the genes that regulate the formation and maintenance of the blood-brain barrier to identify potential therapeutic targets. In order to understand the exact roles these genes play in neural development, it is necessary to assay the effects of altered gene expression on the integrity of the blood-brain barrier. Many of the molecules that function in the establishment of the blood-brain barrier have been found to be conserved across eukaryotic species, including the fruit fly, Drosophila melanogaster. Fruit flies have proven to be an excellent model system for examining the molecular mechanisms regulating nervous system development and function. This protocol describes a step-by-step procedure to assay for blood-brain barrier integrity during the embryonic and larval stages of D. melanogaster development.

Assay for Blood-brain Barrier Integrity in Drosophila melanogaster

Blood-brain barrier integrity is critical for nervous system function. In Drosophila melanogaster, the blood-brain barrier is formed by glial cells during late embryogenesis. This protocol describes methods to assay for blood-brain barrier formation and maintenance in D. melanogaster embryos and third instar larvae.

Ensayo para la integridad de la barrera hematoencefálica en Drosophila melanogaster

Proper nervous system development includes the formation of the blood-brain barrier, the diffusion barrier that tightly regulates access to the nervous ...

Biología del Desarrollo

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.

La medición y la modificación de acoplamiento de unidad en 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, ...

A Temperature Gradient Assay to Determine Thermal Preferences of Drosophila Larvae

Many animals, including the fruit fly, Drosophila melanogaster, are capable of discriminating minute differences in&#160;environmental temperature, which enables them to seek out their preferred thermal landscape. To define the temperature preferences of larvae over a defined linear range, we developed an assay using a temperature gradient. To establish a single-directional gradient, two aluminum blocks are connected to independent water baths, each of which controls the temperature of individual blocks. The two blocks set the lower and upper limits of the gradient. The temperature gradient is established by placing an agarose-coated aluminum plate over the two water-controlled blocks so that the plate spans the distance between them. The ends of the aluminum plate that is set on the top of the water blocks defines the minimum and maximum temperatures, and the regions in-between the two blocks form a linear temperature gradient. The gradient assay can be applied to&#160;larvae of different ages and can be used to identify mutants that exhibit phenotypes, such as those with mutations affecting genes encoding TRP channels and opsins, which are required for temperature discrimination.

A Temperature Gradient Assay to Determine Thermal Preferences of Drosophila Larvae

Here, we present a protocol to determine the preferred environmental temperature of Drosophila larvae using a continuous thermal gradient.

Un análisis de gradiente de temperatura para determinar las preferencias térmicas de larvas de Drosophila

Many animals, including the fruit fly, Drosophila melanogaster, are capable of discriminating minute differences in&#160;environmental temperature, which ...

A Rapid Food-Preference Assay in Drosophila

To select food with nutritional value while avoiding the consumption of harmful agents, animals need a sophisticated and robust taste system to evaluate their food environment. The fruit fly, Drosophila melanogaster, is a genetically tractable model organism that is widely used to decipher the molecular, cellular, and neural underpinnings of food preference. To analyze fly food preference, a robust feeding method is needed. Described here is a two-choice feeding assay, which is rigorous, cost-saving, and fast. The assay is Petri-dish-based and involves the addition of two different foods supplemented with blue or red dye to the two halves of the dish. Then, ~70 prestarved, 2-4-day-old flies are placed in the dish and&#160;allowed to choose between blue and red foods in the dark for about 90 min. Examination of the abdomen of each fly is followed by the calculation of the preference index. In contrast to multiwell plates, each Petri dish takes only ~20 s to fill and saves time and effort. This feeding assay can be employed to quickly determine whether flies like or dislike a particular food.

A Rapid Food-Preference Assay in Drosophila

We present a protocol for a two-choice feeding assay for flies. This feeding assay is fast and easy to run and is suitable not only for small-scale laboratory research, but also for high-throughput behavioral screens in flies.

Un ensayo rápido de preferencia alimentaria en Drosophila

To select food with nutritional value while avoiding the consumption of harmful agents, animals need a sophisticated and robust taste system to evaluate ...

Fly Climbing Assay: A Method to Test General Locomotion in Drosophila melanogaster

Source: Gevedon, O., et al. <a href="https://www.jove.com/video/59720/in-vivo-forward-genetic-screen-to-identify-novel-neuroprotective">In Vivo Forward Genetic Screen to Identify Novel Neuroprotective Genes in Drosophila melanogaster</a>. J. Vis. Exp. (2019).
This video describes a simple method to quantify fly locomotion and shows the climbing assay performed.&#160; The fly climbing assay is a general test of motor function which relies on a negative geotaxis response.

Ensayo de escalada con mosca: un método para probar la locomoción general en Drosophila melanogaster

Source: Gevedon, O., et al. In Vivo Forward Genetic Screen to Identify Novel Neuroprotective Genes in Drosophila melanogaster. J. Vis. Exp. (2019).
This ...

JoVE Encyclopedia of Experiments

Biología

Drosophila melanogaster (mosca de la fruta)

Adulto

Egg Laying Assay: A Method to Quantify the Egg-Laying Behavior of C. elegans

Source: Lee, S. Y. and Kang, K. <a href="https://www.jove.com/video/60419/measuring-effects-bacteria-chemicals-on-intestinal-permeability">Measuring the Effect of Chemicals on the Growth and Reproduction of&#160;Caenorhabditis elegans</a>.&#160;J. Vis. Exp. (2017).
C. elegans egg-laying behavior can provide important information in developmental, reproductive, and anatomical studies.&#160; This video introduces the important steps of an egg laying assay and shows a sample experiment in which a drug is tested for reproductive toxicity.

Ensayo de puesta de huevos: un método para cuantificar el comportamiento de puesta de huevos de C. elegans

Source: Lee, S. Y. and Kang, K. Measuring the Effect of Chemicals on the Growth and Reproduction of&#160;Caenorhabditis elegans.&#160;J. Vis. Exp. 
...

Caenorhabditis elegans (gusano)

Métodos básicos

Island Assay: A High-Throughput Method to Evaluate Drosophila Locomotor Behavior

Source: Eidhof, I., et al. <a href="https://www.jove.com/video/55892/" target="_blank">High-throughput Analysis of Locomotor Behavior in the Drosophila Island Assay</a>. J. Vis. Exp. (2017).
This video describes the island assay, a behavioral method used to study Drosophila locomotion. The sample protocol features a setup for the assay compatible with semi-automated image analysis.

Ensayo de isla: un método de alto rendimiento para evaluar el comportamiento locomotor de Drosophila

Source: Eidhof, I., et al. High-throughput Analysis of Locomotor Behavior in the Drosophila Island Assay. J. Vis. Exp. (2017).
This video describes the ...

Ensayo de Alto Rendimiento para el Examen de Preferencias de la puesta de huevos individual

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