Name: dii-labeling of drg neurons to study axonal branching in a whole mount preparation of mouse embryonic spinal cord
Uploaded: 2011-12-13T12:00:00
Description: Watch this Scientific Journal Video about DiI-Labeling of DRG Neurons to Study Axonal Branching in a Whole Mount Preparation of Mouse Embryonic Spinal Cord at JoVE.com

लेखकों के लिए उपयोगी टिप्पणी के लिए डॉ. एलिस्टेयर Garratt (अधिकतम Delbrück केंद्र, बर्लिन) धन्यवाद करना चाहते हैं. यह काम जर्मन अनुसंधान परिषद (DFG) के सहयोगात्मक अनुसंधान केंद्र (SFB665) द्वारा समर्थित किया गया.

1. विच्छेदन प्रक्रिया नोट: चूहों के प्रायोगिक उपयोग आधिकारिक तौर पर और प्रयोगशाला पशुओं की देखभाल और उपयोग के लिए दिशा निर्देशों का पालन करने के लिए अनुमोदित होना चाहिए. <ol><li> तैयारी पहले, ?...

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टकसाली प्रक्षेपण axonal शाखा गठन के दोनों प्रकार की तैयारी के DiI लेबलिंग के लिए तय ऊतक के उपयोग के साथ संयोजन में आसानी से एक साथ शामिल पैटर्न संलग्न DRG शाखाओं में बंटी axonal अध्ययन के लिए एक अनुकूल मॉडल के साथ भ?...

<table border="1"><tbody><tr><td> अभिकर्मक के नाम </td><td> कंपनी </td><td> सूचीपत्र संख्या </td><td> टिप्पणियां (वैकल्पिक) </td></tr><tr><td> Stereomicroscope STEMI DRC </td><td> Zeiss </td><td></td><td></td></tr><tr><td> फास्फेट - बफर समाधान (पीबीएस) </td><td> Biochrom एजी </td><td> L182 - 50 </td><td></td></tr><tr><td> Paraformaldehyde </td><td> मर्क </td><td> 8.18715.1000 </td><td></td></tr><tr><td> मानक शल्य कैंची </td><td> ललित विज्ञान उपकरण </td><td> 14001-13 </td><td></td></tr><tr><td> दांतेदार मानक संदंश </td><td> ललित विज्ञान उपकरण </td><td> 11021-14 </td><td></td></tr><tr><td> अतिरिक्त ठीक परितारिका कैंची </td><td> ललित विज्ञान उपकरण </td><td> 14088-10 </td><td></td></tr><tr><td> घुमावदार संदंश </td><td> ललित विज्ञान उपकरण </td><td> 11003-13 </td><td></td></tr><tr><td> Dumont No.5 ठीक सुझावों संदंश </td><td> ललित विज्ञान उपकरण </td><td> 11254-20 </td><td></td></tr><tr><td> Dumont No.5 दर्पण खत्म संदंश </td><td> ललित विज्ञान उपकरण </td><td> 11252-23 </td><td></td></tr><tr><td> Vannas - Tübingen वसंत कैंची </td><td> ललित विज्ञान उपकरण </td><td> 15008-08 </td><td></td></tr><tr><td> फिल्टर पेपर </td><td> फिशर साइंटिफिक </td><td> FB59041 </td><td></td></tr><tr><td> 184 Sylgard </td><td> विश्व Precission उपकरण </td><td> SYLG184 </td><td></td></tr><tr><td> 100 मिमी पेट्री डिश </td><td> Greiner </td><td> 663102 </td><td></td></tr><tr><td> 12 मिलीलीटर polypropylene ट्यूब </td><td> कार्ल रोथ GmbH </td><td> ECO3.1 &lt;/ P&gt; <td></td></tr><tr><td> 12 अच्छी तरह से संस्कृति की थाली </td><td> Becton Dickinson </td><td> 35-3043 </td><td></td></tr><tr><td> इथेनॉल </td><td> मर्क </td><td> 1.00983.2500 </td><td></td></tr><tr><td> / ब्राउन micropipette डांड़ी पी 97-ज्वलंत </td><td> Sutter साधन कं </td><td></td><td></td></tr><tr><td> Borosilicate ग्लास capillaries </td><td> हार्वर्ड उपकरण </td><td> 30-0066 </td><td></td></tr><tr><td> DiI (1,1 &#39;- Dioctadecyl-3, 3,3 3&#39;- tetramethyl, perchlorate indocarbocyanine) </td><td> सिग्मा Aldrich </td><td> 468495 </td><td></td></tr><tr><td> माइक्रोस्कोप SuperFrost प्लस स्लाइड </td><td> कार्ल रोथ GmbH </td><td> H867.1 </td><td></td></tr><tr><td> ग्लास कवर फिसल जाता है </td><td> कार्ल रोथ GmbH </td><td> 1870.2 </td><td></td&gt; </tr></tbody></table>

dii-labeling of drg neurons to study axonal branching in a whole mount preparation of mouse embryonic spinal cord

Here we present a technique to label the trajectories of small groups of DRG neurons into the embryonic spinal cord by diffusive staining using the lipophilic tracer 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI)1. The comparison of axonal pathways of wild-type with those of mouse lines in which genes are mutated allows testing for a functional role of candidate proteins in the control of axonal branching which is an essential mechanism in the wiring of the nervous system. Axonal branching enables an individual neuron to connect with multiple targets, thereby providing the physical basis for the parallel processing of information. Ramifications at intermediate target regions of axonal growth may be distinguished from terminal arborization. Furthermore, different modes of axonal branch formation may be classified depending on whether branching results from the activities of the growth cone (splitting or delayed branching) or from the budding of collaterals from the axon shaft in a process called interstitial branching2 (Fig. 1).
The central projections of neurons from the DRG offer a useful experimental system to study both types of axonal branching: when their afferent axons reach the dorsal root entry zone (DREZ) of the spinal cord between embryonic days 10 to 13 (E10 - E13) they display a stereotyped pattern of T- or Y-shaped bifurcation. The two resulting daughter axons then proceed in rostral or caudal directions, respectively, at the dorsolateral margin of the cord and only after a waiting period collaterals sprout from these stem axons to penetrate the gray matter (interstitial branching) and project to relay neurons in specific laminae of the spinal cord where they further arborize (terminal branching)3. DiI tracings have revealed growth cones at the dorsal root entry zone of the spinal cord that appeared to be in the process of splitting suggesting that bifurcation is caused by splitting of the growth cone itself4 (Fig. 2), however, other options have been discussed as well5.
This video demonstrates first how to dissect the spinal cord of E12.5 mice leaving the DRG attached. Following fixation of the specimen tiny amounts of DiI are applied to DRG using glass needles pulled from capillary tubes. After an incubation step, the labeled spinal cord is mounted as an inverted open-book preparation to analyze individual axons using fluorescence microscopy.

Watch this Scientific Journal Video about DiI-Labeling of DRG Neurons to Study Axonal Branching in a Whole Mount Preparation of Mouse Embryonic Spinal Cord at JoVE.com

DiI-Labeling of DRG Neurons to Study Axonal Branching in a Whole Mount Preparation of Mouse Embryonic Spinal Cord

कृंतक रीढ़ की हड्डी में संवेदी afferents के स्टिरियोटाइप अनुमानों एक आसानी से सुलभ प्रयोगात्मक प्रणाली एकल axons की अनुरेखण के माध्यम से axonal शाखाओं में अध्ययन की पेशकश करते हैं.

DRG न्यूरॉन्स की DiI लेबल axonal अध्ययन माउस भ्रूणीय स्पाइनल कॉर्ड के एक साबुत माउंट तैयारी में शाखाओं में बंटी

Here we  present a technique to label the trajectories of small groups of DRG neurons  into the embryonic spinal cord by diffusive staining using the ...

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