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생물학

संचयन एवं तैयारी ड्रोसोफिला भ्रूण electrophysiological रिकॉर्डिंग और अन्य प्रक्रियाओं के लिए

Published: May 20, 2009
doi:
10.3791/1347
, ,
1Department of Biological Sciences,University of Illinois, 2Department of Biological Sciences,Vanderbilt University

Summary

इस तकनीक immunohistochemistry या electrophysiological रिकॉर्डिंग के लिए ड्रोसोफिला भ्रूण neuromusculature उजागर. यह neuromuscular विकास में जल्दी घटनाओं का अध्ययन या म्यूटेंट है कि नहीं हैच कर सकते हैं इलेक्ट्रोफिजियोलॉजी प्रदर्शन के लिए उपयोगी है.

Abstract

ड्रोसोफिला दोनों भ्रूण के विकास और कार्यात्मक तंत्रिका विज्ञान के अध्ययन के लिए एक प्रमुख आनुवंशिक मॉडल है . परंपरागत रूप से, इन क्षेत्रों एक दूसरे से काफी अलग कर रहे हैं काफी हद तक स्वतंत्र इतिहास और वैज्ञानिक समुदाय के साथ. हालांकि, इन आमतौर पर असमान क्षेत्रों के बीच इंटरफेस विकासात्मक कार्यक्रमों कार्यात्मक बिजली के संकेत गुण और तंत्रिका सर्किट के गठन के अंतिम चरणों के दौरान कार्यात्मक रासायनिक synapses के भेदभाव के अधिग्रहण अंतर्निहित है. इस अंतरफलक को जांच के लिए एक महत्वपूर्ण क्षेत्र है. ड्रोसोफिला में, कार्यात्मक विकास के इन चरणों <8 घंटे की एक embryogenesis के आखिरी तीसरे के दौरान (25 डिग्री सेल्सियस पर) अवधि के दौरान पाए जाते हैं . यह देर विकासात्मक अवधि लंबी जांच एक कठिन, अभेद्य epidermal छल्ली के बयान के कारण असभ्य माना जाता था. एक सफलता अग्रिम पानी polymerizing शल्य चिकित्सा गोंद के अनुप्रयोग है कि स्थानीय स्तर पर छल्ली के लिए लागू किया जा सकता है देर चरण भ्रूण की नियंत्रित विच्छेदन सक्षम था. एक पृष्ठीय अनुदैर्ध्य चीरा के साथ, भ्रूण फ्लैट रखी जा सकता है, प्रयोगात्मक जांच करने के लिए उदर तंत्रिका की हड्डी और शरीर दीवार मांसलता को उजागर. इस प्रणाली के भारी कर दिया गया है को अलग और आनुवंशिक म्यूटेंट कि भ्रूण synapse गठन ख़राब है, और इस प्रकार आणविक तंत्र विनिर्देश और synapse कनेक्शन और कार्यात्मक synaptic संकेत गुणों के भेदभाव को नियंत्रित प्रकट विशेषताएँ इस्तेमाल किया.

Protocol

भाग 1: उपकरण और आपूर्ति एक अच्छा विच्छेदन खुर्दबीन भ्रूण dissections के लिए आवश्यक है, 40x बढ़ाई, 25x-आंख टुकड़ों के साथ सुझाव दिया है करने के लिए ज़्यादा से ज़्यादा बढ़ाई वृद्धि. ठीक संदंश (संख्या 5) भ्रूण और ?…

Discussion

भ्रूण की सटीक मचान कारण सिर्फ कई घंटे के समय के पाठ्यक्रम पर कार्यात्मक संपत्तियों का तेजी से परिपक्वता के लिए महत्वपूर्ण है. कई मुद्दों पर इस मचान जटिल. सबसे पहले, सबसे शोधकर्ताओं समय अंडे चरण भ्रूण क?…

Acknowledgements

KB NIH अनुदान GM54544 द्वारा समर्थित है.

Materials

Material Name Type Company Catalogue Number Comment
Embryo Collection Cages   Genesee Scientific (www.flystuff.com) 59-100 (for 60 mm dish; other sizes available) Cages can also be home made using punctured tri-pour beakers, as shown in video
Sylgard 184   Dow Corning Available from various companies Surgical glue adheres better to sylgard-coated coverslips
Fine glass tubing, outer diameter 1.0-1.5 mm   various   For pulling into fine glass needles for dissection and tubes for glue delivery
Plastic tubing, to attach to glass pipette for mouth suction and glue application   Tygon   Tubing inner diameter needs to match glass outer diameter.
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Tags

DrosophilaEmbryosElectrophysiological RecordingProceduresGenetic ModelEmbryonic DevelopmentFunctional NeuroscienceDevelopmental ProgramsElectrical Signaling PropertiesChemical SynapsesNeural Circuit FormationInvestigationLate Developmental PeriodEpidermal CuticleSurgical GlueControlled DissectionDorsal Longitudinal IncisionVentral Nerve CordBody Wall MusculatureGenetic MutantsSynapse FormationMolecular Mechanisms
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Featherstone, D. E., Chen, K., Broadie, K. Harvesting and Preparing Drosophila Embryos for Electrophysiological Recording and Other Procedures. J. Vis. Exp. (27), e1347, doi:10.3791/1347 (2009).
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