Name: measuring and altering mating drive in male drosophila melanogaster
Uploaded: 2017-02-15T14:00:00
Description: Watch this Scientific Journal Video about Measuring and Altering Mating Drive in Male Drosophila melanogaster at JoVE.com

The authors thank Mike Crickmore, Dragana Rogulja, and Michelle Frank for comments on the manuscript. Pavel Gorelik provided technical support for manufacturing the behavioral arenas. This work was conducted in Mike Crickmore's lab and is also supported by the Whitehall Foundation (Principal Investigator: Dragana Rogulja). S.X.Z. is a Stuart H.Q. and Victoria Quan Fellow at Harvard Medical School.

BEMÆRK: Denne protokol beskriver fremstillingen (§§ 1 - 3), udførelse (afsnit 4), og analyse (afsnit 4) 2-D mæthed analyser. Derefter bruger dopaminerg stimulation som et eksempel, afsnit 5 viser, hvordan man kan kombinere thermogenetic stimulation med 2-D mæthed assays til at fremkalde hyperseksualitet. Afsnit 6 beskriver 3 måder at kontrollere resultaterne af 2-D mæthed analyser. Endelig, afsnit 7 viser hvordan man kan måle inddrivelse af parring drev i mandlige fluer. 1. Bearbejdni...

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Motiverende stater kan mæt, vedligeholdes, og genvundet 34. Vi præsenterer en 2-D mæthed assay, der hurtigt og håndfast måler alle disse aspekter af parring drev i fluen. Dette assay åbner mulighed for at anvende avancerede flyve genetiske manipulationer for at studere de molekylære og kredsløbskomponenter i en motiveret adfærd. Den mæthed analysen er afhængig af mandens evne til at retten og parre sig, og for at afslutte parringer på et passende tidspunkt...

Arbejde i Drosophila har givet dybe og banebrydende indsigt i mange biologiske fænomener, herunder arten af genet 1, principperne for fosterudvikling 2, døgnrytmen 3, samt udvikling og ledningsføring af nervesystemet 4, 5, 6. Motivation er stadig langt mindre godt forstået end disse fænomener, måske på grund af de begrænsninger på de systemer, der er blevet undersøgt hidtil. Motivation i gylp primært undersøgt i forbindelse med sult, som rummer mange udfordringer på grund af deres forsvindende lille fødeindtagelse pr fodring Bout og exoskelet der udelukker åbenlyse tegn på fedtaflejring. Der er derfor et behov for at udvide de anvendte systemer til at studere motivation i gylp. Vi beskriver en adfærdsmæssig ramme for studiet af parring drev iDrosophila. Dette system drager fordel af de neurogenetic værktøjer i gylp samt tilgængeligheden 7, 8, 9, 10, 11, 12 og den formodede connectome af sin seksuelt dimorfe kredsløb 8, 13. Desuden er meget af den medfødte 14, 15, 16, 17, 18, 19, 20, 21 og lærte 22, 23, 24 sensorisk-motorisk kredsløb kontrollerende frieri er udarbejdet i detaljer, hvilket giver en sjælden mulighedat finde den nøjagtige kredsløb node hvorpå motivation rammer. Vi har for nylig rapporteret, at i fluen, som hos mennesker, dopamin niveauer er centrale for parring drev 25, 26, 27. Vi har fået genetisk adgang til den relevante dopamin-producerende og modtage neuroner i fluen, lette detaljerede molekylære mekanisme og kredsløb niveau analyser af denne bevaret fænomen ved hjælp af de analyser vi beskriver her 25. Vi tilføjer til de adfærdsmæssige analyser i Zhang et al. 25 en ny flad adfærdsmæssige arena, der giver mulighed for video scoring, som vi kalder en 2-dimensional (2-D) mæthed assay, en vigtig forbedring i forhold til tidligere metoder. Følgelig den nye assay er mere skalerbar og målelig, og derfor mere egnet til genetiske skærme af gener og neuroner involveret i motivation. Vi bruger denne nye analyse, sammen med frieri assays og neurogetiske manipulationer, at vise, hvordan man kan måle og ændre parring drev i fluen.

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			<td height="21" style="height:21px;width:216px;">1/16 inch clear acrylic</td>
			<td style="width:119px;">McMaster-Carr</td>
			<td style="width:119px;">8589K12</td>
			<td style="width:743px;">Used to make arenas; see Supplemental Material 1 for designs.</td>
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			<td height="21" style="height:21px;">1/8 inch clear acrylic</td>
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			<td height="21" style="height:21px;">3/16 inch clear acrylic</td>
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			<td>Used to make arenas; see Supplemental Material 1 for designs.</td>
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			<td height="21" style="height:21px;">1/32 inch black delrin</td>
			<td style="width:119px;">McMaster-Carr</td>
			<td>8575K132</td>
			<td>Used to make arenas; see Supplemental Material 1 for designs.</td>
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			<td height="21" style="height:21px;">Hex screws, 1 inch long (50x)</td>
			<td style="width:119px;">McMaster-Carr</td>
			<td>92314A115&#160;</td>
			<td>Used to make arenas. Can be replaced by 3/4 inch screws (92314A113, McMaster-Carr) for 32-chamber arenas.</td>
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			<td height="21" style="height:21px;">Camcorder</td>
			<td style="width:119px;">Canon</td>
			<td>Vixia HF R700</td>
			<td>Can be replaced by any consumer comcorder.</td>
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measuring and altering mating drive in male drosophila melanogaster

Trods årtiers efterforskning, de neuronale og molekylære grundlag for motiverende stater forbliver mystisk. Vi har for nylig udviklet en ny, reduktionistisk og skalerbare system til dybtgående undersøgelse af motivation ved hjælp af parring drevet af mandlige Drosophila melanogaster (Drosophila), de metoder, som vi detaljer her. Den adfærdsmæssige paradigme centre på den konstatering, at mandlige parring drev falder sammen fertiliteten i løbet af gentagne parringer og genopretter løbet ~ 3 d. I dette system, de kraftfulde neurogenetic værktøjer til rådighed i fluen konvergerer med den genetiske tilgængelighed og formodede ledningsdiagram til rådighed for seksuel adfærd. Denne konvergens muliggør hurtig isolation og forhør af små neuronale populationer med specifikke motiverende funktioner. Here We detalje projektering og udførelse af mæthed assay, der bruges til at måle og ændre frieri motivation i den mandlige flue. Brug af denneassay viser vi også, at lave mandlige parring drev kan overvindes ved at stimulere dopaminerge neuroner. Den mæthedsfornemmelse analysen er enkel og billig, og robust til påvirkninger af genetisk baggrund. Vi forventer, at mæthed assay til at generere mange nye indsigter i neurobiologi af motiverende stater.

For at karakterisere Drosophila parring drev, 3 dage gammel, WT Canton-S hanner blev testet i en 2-D mæthed assay. I løbet af assayet (4,5 timer), hanner parrer et gennemsnit på 4,8 ± 0,3 (middelværdi ± standardfejl på middelværdi, SEM) gange. Parringer indlede meste i de første 2 timer (78%) (figur 6a, 6b) og blive mindre hyppige som assayet skrider (figur 6A, 6B). Dette fald skyldes ikke mangel på parring partnere (74% kvinder forbli...

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Measuring and Altering Mating Drive in Male Drosophila melanogaster

Denne artikel beskriver en adfærdsmæssig analyse der bruger mandlige parring drev i Drosophila melanogaste r at studere motivation. Ved hjælp af denne metode, kan forskerne udnytte avancerede flyve neurogenetic teknikker til at afdække de genetiske, molekylære og cellulære mekanismer der ligger til grund denne motivation.

Måling og Ændre Parring Drive i 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, ...

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Neuroscience

Measuring and Altering Mating Drive in Male Drosophila melanogaster

Single Sensillum Recordings in the Insects Drosophila melanogaster and Anopheles gambiae

Single Sensillum Recordings in the Insects Drosophila melanogaster and Anopheles gambiae

The sense of smell is essential for insects to find foods, mates, predators, and oviposition sites3. Insect olfactory sensory neurons (OSNs) are enclosed ...

Optogenetic Stimulation of Escape Behavior in Drosophila melanogaster

Optogenetic Stimulation of Escape Behavior in Drosophila melanogaster

Optogenetic Stimulering af flugt Opførsel i<em&gt; Drosophila melanogaster</em

A  growing number of genetically encoded tools are becoming available that allow non-invasive  manipulation of the neural activity of specific neurons in ...

Determination of the Spontaneous Locomotor Activity in Drosophila melanogaster

Determination of the Spontaneous Locomotor Activity in Drosophila melanogaster

Bestemmelse af Spontan motorisk aktivitet i<em&gt; Drosophila melanogaster</em

Drosophila melanogaster has been used as an excellent model organism to study environmental and genetic manipulations that affect behavior. One such ...

Straightforward Assay for Quantification of Social Avoidance in Drosophila melanogaster

Straightforward Assay for Quantification of Social Avoidance in Drosophila melanogaster

Ligetil Assay til kvantificering af Social undgåelse i<em&gt; Drosophila melanogaster</em

Drosophila melanogaster is an emerging model to study different aspects of social interactions. For example, flies avoid areas previously occupied by ...

Dyeing Insects for Behavioral Assays: the Mating Behavior of Anesthetized Drosophila

Dyeing Insects for Behavioral Assays: the Mating Behavior of Anesthetized Drosophila

Farvning Insekter for Behavioral Analyser: den parringsadfærd af anæsteseret<em&gt; Drosophila</em

Mating experiments using Drosophila have contributed greatly to the understanding of sexual selection and behavior. Experiments often require simple, easy ...

Conditions Affecting Social Space in Drosophila melanogaster

Conditions Affecting Social Space in Drosophila melanogaster

Forhold, der påvirker sociale rum i<em&gt; Drosophila melanogaster</em

The social space assay described here can be used to quantify social interactions of Drosophila melanogaster &#8212; or other small insects &#8212; in a ...

Foraging Path-length Protocol for Drosophila melanogaster Larvae

Foraging Path-length Protocol for Drosophila melanogaster Larvae

Fouragering Sti-længde Protokol for<em&gt; Drosophila melanogaster</em&gt; Larver

The Drosophila melanogaster larval path-length phenotype is an established measure used to study the genetic and environmental contributions to behavioral ...

The CApillary FEeder Assay Measures Food Intake in Drosophila melanogaster

The CApillary FEeder Assay Measures Food Intake in Drosophila melanogaster

Den kapillære Feeder-analysen Måler fødeindtagelse i<em&gt; Drosophila melanogaster</em

For most animals, feeding is an essential behavior for securing survival, and it influences development, locomotion, health and reproduction. Ingestion of ...

Metabolic Analysis of Drosophila melanogaster Larval and Adult Brains

Metabolic Analysis of Drosophila melanogaster Larval and Adult Brains

Metaboliske analyse af Drosophila melanogaster Larval og voksen hjerne

This protocol describes a method for measuring the metabolism in Drosophila melanogaster larval and adult brains. Quantifying metabolism in whole organs ...

Morphological and Functional Evaluation of Axons and their Synapses during Axon Death in Drosophila melanogaster

Morphological and Functional Evaluation of Axons and their Synapses during Axon Death in Drosophila melanogaster

Morfologisk og funktionel vurdering af Axoner og deres synapser under Axon Death i Drosophila melanogaster

Axon degeneration is a shared feature in neurodegenerative disease and when nervous systems are challenged by mechanical or chemical forces. However, our ...