Name: paired nanoinjection and electrophysiology assay to screen for bioactivity of compounds using the drosophila melanogaster giant fiber system
Uploaded: 2012-04-15T14:00:00
Description: Watch this Scientific Journal Video about Paired Nanoinjection and Electrophysiology Assay to Screen for Bioactivity of Compounds using the Drosophila melanogaster Giant Fiber System at JoVE.com

Wij willen de leden van de Mari-lab en de Godenschwege lab, te erkennen in het bijzonder Aline Yonezawa, voor commentaar en hulp bij dit protocol. Dit werk werd gefinancierd door het Nationaal Instituut voor Neurologische Aandoeningen en Stroke subsidie ​​R21NS06637 naar FM-en TAG; AB werd gefinancierd door de National Science Foundation Award nummer 082925, URM: Integratieve Biologie voor toekomstige onderzoekers.

1. Elektrofysiologie Rig Set-up <ol><li> De benodigde apparatuur voor de elektrofysiologie rig set-up wordt in detail beschreven door Augustin et al.. In dit Journal 14. Raadpleeg dit artikel voor een uitgebreide uitleg van de benodigde elektrofysiologische apparaten. </li><li> Wijzig de eerder beschreven elektrofysiologie rig set-up 14 door het toevoegen van een zesde micromanipulator, die de nanoinjector houdt. Voor een gemakkelijke toegang tot de kop van het dier dient te worden gepla...

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De nanoinjection / elektrofysiologie bioassay hier gepresenteerde zorgt voor een snelle screening van verbindingen in het zenuwstelsel van de fruitvlieg. Dit is een nieuwe in vivo techniek dat kleine hoeveelheden van een verbinding vereist invloed op een verscheidenheid van eiwitten wekken in een goed gekarakteriseerd neuronale circuit. Deze methode kan worden gebruikt om de biologische activiteit van verschillende verbindingen testen van onbekende toxinen in de handel verkrijgbare farmacologische agentia. ...

<table border="1"><tbody><tr><td> Naam van het serum </td><td> Vennootschap </td><td> Catalogusnummer </td><td> Reacties </td></tr><tr><td> Opname glaselektroden: borosilicaatglas haarvaten </td><td> Wereld Precision Instruments </td><td> 1B100F-4 </td><td> 1,0 mm OD, 0.58mm ID </td></tr><tr><td> Stimulator </td><td> Grass Instrumenten </td><td> Model S48 </td><td></td></tr><tr><td> Versterker </td><td> Getting Instruments Inc </td><td> Model 5A </td><td></td></tr><tr><td> Data acquisitie Software: Digidata </td><td> Molecular Devices </td><td> Model 1440A </td><td></td></tr><tr><td> Het verzamelen van gegevens software: pCLAMP </td><td> Molecular Devices </td><td> Versie 10 </td><td></td></tr><tr><td> Stereomicroscoop wet glasvezel microscoop ringlicht </td><td> AmScope </td><td> SM-4T Model HL250-AR </td><td></td></tr><tr><td> Het ontleden van ruimte voor de montage </td><td> AmScope </td><td> SM-2TZ </td><td></td></tr><tr><td> Kite Manual micromanipulator &amp; kantelen Base </td><td> Wereld Precision Instruments </td><td> Model # M3301 Kite: Model # KITE-M3-L </td><td></td></tr><tr><td> Drosophila melanogaster Wild 10E genotype (wild-type stam) </td><td> Bloomington Stock centrum </td><td> Stock # 3892 </td><td></td></tr><tr><td> Verticale pipet trekker </td><td> David Kopf Instrumenten </td><td> Model 700C </td><td></td></tr><tr><td> Injectie glas micropipetten: borosilicaatglas haarvaten </td><td> Wereld Precision Instruments </td><td> Catalogus # 4878 </td&gt; <td> 1.14mm OD, 0,5 mm ID </td></tr><tr><td> Silicon olie </td><td> Visser </td><td> Catalogus # S159-500 </td><td></td></tr><tr><td> Beveler </td><td> Sutter Instrument Co </td><td> KT Brown Type Model # BV-10 </td><td></td></tr><tr><td> Nanoliter2000 </td><td> Wereld Precision Instruments </td><td> Catalogus # B203XVY </td><td></td></tr><tr><td> Blauwe kleurstof </td><td> McCormick </td><td> N / A </td><td> Ingrediënten: Water, Propylene Glycol, FD &amp; C Blue 1, en 0,1% Propylparaben (conserveermiddel). </td></tr><tr><td> Methyllycaconitine citraat (MLA) </td><td> Tocris bio- </td><td> Catalogus # 1029 </td><td></td></tr><tr><td> Plastic wax sticks </td><td> Hygiënisch Corporation (Akron Ohio USA) </td><td></td><td></td></tr></tbody&gt; </table>

paired nanoinjection and electrophysiology assay to screen for bioactivity of compounds using the drosophila melanogaster giant fiber system

Screening verbindingen in vivo activiteit kan worden gebruikt als eerste stap kandidaten die kunnen worden ontwikkeld tot farmacologische agentia 1,2 identificeren. We ontwikkelden een nieuwe nanoinjection / elektrofysiologische test die de detectie van bioactieve modulerende effecten van stoffen op de functie van een neuronale circuit dat bemiddelt de ontsnapping respons bij Drosophila melanogaster 3,4 mogelijk maakt. De in vivo assay gebruikt Drosophila Giant vezelsysteem (GFS, figuur 1) kan screening van verschillende verbindingen, zoals kleine moleculen of peptiden en vereist slechts minimale hoeveelheden te wekken effect. Daarnaast is de Drosophila GFS biedt een grote verscheidenheid van mogelijke moleculaire doelwitten op neuronen en spieren. De Giant Fibers (GFS) synaps elektrisch (gap junctions) als chemisch (cholinerge) op een Perifere Synapsing Interneuron (PSI) en de Tergo Trochanteral Muscle neuron (TTMn) 5 </sup&gt;. De PSI naar DLMn (Dorsale Longitudinale Muscle neuron) aansluiting is afhankelijk van Dα7 nicotine acetylcholine receptoren (nAChRs) 6. Ten slotte is de neuromusculaire knooppunten (NMJ) van de TTMn en de DLMn met de sprong (TTM) en vliegspieren (DLM) zijn glutamaat 7-12. Hier hebben we laten zien hoe je nanoliter hoeveelheden van een stof te injecteren, terwijl het verkrijgen van elektrofysiologische intracellulaire opnames van de Giant Fiber System 13 en hoe de effecten van de verbinding op de functie van dit circuit te controleren. We tonen specificiteit van de test met methyllycaconitine citraat (MLA), een nAChR antagonist, die de PSI om de verbinding DLMn maar niet de GF om verbinding of de functie van de NMJ op de sprong of vlucht spieren TTMn verstoort. Alvorens te beginnen met deze video is het belangrijk dat je zorgvuldig in de gaten en vertrouwd raken met de Jove video met de titel &quot;elektrofysiologische registraties van de Giant Fiber Pathway van D. Melanogaster &quot;van Augustin et al. 7 de video hier gepresenteerde is bedoeld als een uitbreiding van deze bestaande techniek. Hier elektrofysiologische opnames methode en focus gebruiken in bijzonderheden over de toevoeging van de gepaarde nanoinjections en meettechniek.

Watch this Scientific Journal Video about Paired Nanoinjection and Electrophysiology Assay to Screen for Bioactivity of Compounds using the Drosophila melanogaster Giant Fiber System at JoVE.com

Paired Nanoinjection and Electrophysiology Assay to Screen for Bioactivity of Compounds using the Drosophila melanogaster Giant Fiber System

Een snelle<em&gt; In vivo</em&gt; Test om te testen op neuromodulatory verbindingen met behulp van de Giant Fiber System (GFS) van<em&gt; Drosophila melanogaster</em&gt; Beschreven. Nanoinjections in de kop van het dier, samen met elektrofysiologische opnames van de SBO kan onthullen bioactiviteit van verbindingen op neuronen en spieren.

Gekoppelde Nanoinjection en Elektrofysiologie test voor het screenen op bioactiviteit van verbindingen met behulp van de Drosophila melanogaster Giant Fiber System

Screening compounds for in vivo activity can be used as a first step to identify candidates that may be developed into pharmacological agents1,2. We ...

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Neuroscience

Paired Nanoinjection and Electrophysiology Assay to Screen for Bioactivity of Compounds using the Drosophila melanogaster Giant Fiber System

Electrophysiological Recordings from the Giant Fiber Pathway of D. melanogaster

Electrophysiological Recordings from the Giant Fiber Pathway of D. melanogaster

Elektrofysiologische opnames van de Giant Fiber Pathway van D. melanogaster</em

When startled adult D. melanogaster react by jumping into the air and flying away. In many invertebrate species, including D. melanogaster, the "escape" ...

Whole Mount Preparation of the Adult Drosophila Ventral Nerve Cord for Giant Fiber Dye Injection

Whole Mount Preparation of the Adult Drosophila Ventral Nerve Cord for Giant Fiber Dye Injection

Hele Mount Voorbereiding van de Adult Drosophila Ventraal Nerve Fiber Koord voor Giant Dye Injectie

To analyze the axonal and dendritic morphology of neurons, it is essential to obtain accurate labeling of neuronal structures. Preparing well labeled ...

Labeling of Single Cells in the Central Nervous System of Drosophila melanogaster

Labeling of Single Cells in the Central Nervous System of Drosophila melanogaster

Labeling van afzonderlijke cellen in het centrale zenuwstelsel van<em&gt; Drosophila melanogaster</em

In this  article we describe how to individually label neurons in the embryonic CNS of Drosophila melanogaster by juxtacellular injection of the ...

Determination of the Spontaneous Locomotor Activity in Drosophila melanogaster

Determination of the Spontaneous Locomotor Activity in Drosophila melanogaster

Bepaling van de Spontane bewegingsactiviteit in<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

Ongecompliceerd Assay voor de kwantificering van Sociale Avoidance in<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 ...

High-resolution Quantification of Odor-guided Behavior in Drosophila melanogaster Using the Flywalk Paradigm

High-resolution Quantification of Odor-guided Behavior in Drosophila melanogaster Using the Flywalk Paradigm

Hoge-resolutie Kwantificering van Geur-begeleide Gedrag in<em&gt; Drosophila melanogaster</em&gt; Met behulp van de<em&gt; Flywalk</em&gt; Paradigm

In their natural environment, insects such as the vinegar fly Drosophila melanogaster are bombarded with a huge amount of chemically distinct odorants. To ...

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

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

Een Behavioral Assay voor Mechanosensation van MARCM-gebaseerde klonen in<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 ...

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

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

High Throughput Assay to-eierleggende Voorkeuren van de individuele onderzoeken<em&gt; Drosophila melanogaster</em

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

A Simple Way to Measure Alterations in Reward-seeking Behavior Using Drosophila melanogaster

A Simple Way to Measure Alterations in Reward-seeking Behavior Using Drosophila melanogaster

Een eenvoudige manier om Veranderingen Meet in Reward-zoekend gedrag Met behulp<em&gt; Drosophila melanogaster</em

We describe a protocol for measuring ethanol self-administration in fruit flies (Drosophila melanogaster) as a proxy for changes in reward states. We ...

The CApillary FEeder Assay Measures Food Intake in Drosophila melanogaster

The CApillary FEeder Assay Measures Food Intake in Drosophila melanogaster

De capillaire Feeder test meet voedselinname bij<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 ...