Name: isolation and culture of post-natal mouse cerebellar granule neuron progenitor cells and neurons
Uploaded: 2009-01-16T00:00:00
Description: Watch this Scientific Journal Video about Isolation and Culture of Post-Natal Mouse Cerebellar Granule Neuron Progenitor Cells and Neurons at JoVE.com

我々は、貴重な提案をバーバラカルレッティとアンナマリーケニーに感謝します。 、DK07328：HYLはトレーニンググラント&quot;生化学分子生物学ホルモン&quot;によってサポートされています。 NIHの助成金5R01 NS16951（CAM）によって部分的にサポート。

パート1：（ビデオで示されていません）（解剖前に1〜2日）の設定<ol><li>培養液とメディアを準備： <ul><li> 4X CMF - PBS - EDTA（パーコールの希釈のためのカルシウムとマグネシウムのフリーリン酸緩衝生理食塩水- EDTA）：リットルは、32グラムのNaCl、1.2グラムのKCl、8 gのグルコース、2グラムのNaH 2 PO 4、1gのKH 2 POを追加する4、8 mLの2％のNaHCO 3の株式、脱?...

<ol>
	<li>Altman, J., Bayer, S. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Development of the cerebellar system: in relation to its evolution, structure, and functions. , (1997).</li><li>Hatten, M. E., Heintz, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Annual Review of Neuroscience. 18, 385-285 (1995).</li><li>Hatten, M. E., Gao, W. -. Q., Morrison, M. E., Banker, G., Goslin, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> in Culturing Nerve Cells. , 419-419 (1998).</li><li>Hatten, M. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The Journal of Cell Biology. 100 (2), 384-384 (1985).</li><li>Carletti, B., Rossi, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Neuroscientist. 14 (1), 91-91 (2008).</li><li>Knoepfler, P. S., Kenney, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Cell Cycle. 5 (1), 47-47 (2006).</li><li>Manzini, M. C., Joseph, D. J., MacDermott, A. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Molecular and Cellular Neuroscience. 35 (2), 328-328 (2007).</li><li>Manzini, M. C., Ward, M. S., Zhang, Q. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Journal of Neuroscience. 26 (22), 6040-6040 (2006).</li><li>Baptista, C. A., Hatten, M. E., Blazeski, 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> Neuron. 12 (2), 243-243 (1994).</li><li>Palay, S. L., Chan-Palay, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Cerebellar Cortex: Cytology and Organization. , (1974).</li><li>Messer, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Brain Research. 130 (1), 1-1 (1977).</li><li>Ruiz I Altaba, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Development. 126, 3205-3205 (1999).</li><li>Wallace, V. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Current Biology. 9 (8), 445-445 (1999).</li><li>Wechsler-Reya, R. J., Scott, M. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Neuron. 22 (1), 103-103 (1999).</li><li>Solecki, D. J., Liu, X. L., Tomoda, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Neuron. 31 (4), 557-557 (2001).</li><li>Powell, S. K., Williams, C. C., Nomizu, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Journal of Neuroscience Research. 54 (2), 233-233 (1998).</li><li>Lander, A. D., Fujii, D. K., Reichardt, L. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Proceedings of the National Academy of Sciences of the United States of America. 82 (7), 2183-2183 (1985).</li><li>Kenney, A. M., Rowitch, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Molecular and Cellular Biology. 20 (23), 9055-9055 (2000).</li><li>Kenney, A. M., Cole, M. D., Rowitch, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Development. 130 (1), 15-15 (2003).</li><li>Pons, S., Trejo, J. L., Martinez-Morales, J. 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> Development. 128, 1481-1481 (2001).</li></ol>

このプロトコルは、過去3,7,11に記載されている手順の変更に基づいています。以下に述べるように注意することは、いくつかの重要な点があります。 顆粒ニューロンおよび前駆細胞は、メッキの2時間以内に従っている。健康な細胞は位相差顕微鏡の4の下にある丸い形態を持っている。メッキ後24時間以内に、健康な細胞はカバースリップまたはプラスチック製のよく周りに均等に広が...

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		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Cell strainer with 70&mu;m mesh pore</td>
<td>&nbsp;</td>
<td>BD Biosciences</td>
<td>352350</td>
<td>&nbsp;</td>
<tr>
<td>Spinal Needle</td>
<td>&nbsp;</td>
<td>BD</td>
<td>405182</td>
<td>20G x 3-1/2 inches</td>
</tr>
<tr>
<td>Poly-D-Lysine mol wt>300,000</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>P1024</td>
<td>Each new batch of Poly-D-Lysine must be tested. 
</td>
</tr>
<tr>
<td>Percoll</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>P-1644</td>
<td>&nbsp;</td>
<tr>
<td>Penicillin-Streptomycin (100X)</td>
<td>&nbsp;</td>
<td>Sigma</td>
<td>P4333</td>
<td>&nbsp;</td>
<tr>
<td>HBSS</td>
<td>&nbsp;</td>
<td>Invitrogen</td>
<td>14175-103</td>
<td>Must be calcium and magnesium free.</td>
</tr>
<tr>
<td>Neurobasal A-Medium</td>
<td>&nbsp;</td>
<td>Invitrogen</td>
<td>10888-022</td>
<td>&nbsp;</td>
<tr>
<td>Glutamax I supplement</td>
<td>&nbsp;</td>
<td>Invitrogen</td>
<td>35050-061</td>
<td>&nbsp;</td>
<tr>
<td>B-27 Serum Free Supplement (50x)</td>
<td>&nbsp;</td>
<td>Invitrogen</td>
<td>17504-044</td>
<td>&nbsp;</td>
<tr>
<td>12-mm circle coverslips </td>
<td>&nbsp;</td>
<td>Carolina Biological Supply</td>
<td>63-3029</td>
<td>These are made in Germany by Deckgluder.</td>
</tr>
<tr>
<td>25-mm circle coverslips</td>
<td>&nbsp;</td>
<td>Carolina Biological Supply</td>
<td>63-3037</td>
<td>These are made in Germany by Deckgluder.</td>
</tr>
<tr>
<td>Papain Dissociation System </td>
<td>&nbsp;</td>
<td>Worthington Biochemical Corporation</td>
<td>LK 003150</td>
<td>Kit is good for five isolations.</td>
</tr>
<tr>
<td>Permaset scissors</td>
<td>&nbsp;</td>
<td>Roboz</td>
<td>RS6782</td>
<td>&nbsp;</td>
<tr>
<td>Dumont #5 (Dumostar)</td>
<td>&nbsp;</td>
<td>Roboz</td>
<td>RS4978</td>
<td>&nbsp;</td>
<tr>
<td>4-well culture dish</td>
<td>&nbsp;</td>
<td>Nunc</td>
<td>176740</td>
<td>&nbsp;</td>
<tr>
<td>6-well culture dish</td>
<td>&nbsp;</td>
<td>Nunc</td>
<td>140685</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

isolation and culture of post-natal mouse cerebellar granule neuron progenitor cells and neurons 

小脳皮質は神経細胞の特性と開発の1,2を研究するためのユニークな機会を提供するだけでなく説明する構造です。小脳ニューロンタイプ（顆粒細胞、プルキンエ細胞と抑制性介在）のうち、顆粒ニューロンは、これまでで最も多数あり、哺乳類の脳における神経細胞の中で最も豊富なタイプです。げっ歯類では、小脳顆粒ニューロンは小脳皮質、外顆粒層（EGL）の最外層の前駆細胞からの最初の二つの出生後の数週間の間に生成されます。ここで紹介するプロトコールは、豊かにする技術と文化の顆粒ニューロンと出生後のマウスの小脳からそれらの前駆細胞を説明します。我々は、顆粒ニューロン5,6に前駆細胞の増殖分化を研究するために使用することができます増やす純度3,4の文化を、取得する手順を説明します。一度前駆細胞が分化、文化はまた、シナプス形成、グルタミン酸受容体の機能7、他の精製された小脳細胞8,9または細胞死7との相互作用などの現象の実験的操作と特性評価のための顆粒神経細胞の均質な集団を提供しています。

Watch this Scientific Journal Video about Isolation and Culture of Post-Natal Mouse Cerebellar Granule Neuron Progenitor Cells and Neurons at JoVE.com

Isolation and Culture of Post-Natal Mouse Cerebellar Granule Neuron Progenitor Cells and Neurons

ここでは、出生後のマウスから小脳顆粒ニューロン前駆細胞や小脳顆粒ニューロンを分離して培養する方法を提示する。

ポストナタールマウス小脳顆粒ニューロン前駆細胞と神経細胞の単離と培養

The cerebellar cortex is a well described structure that provides unique opportunities for studying neuronal properties and development1,2. Of the ...

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Biology

Isolation and Culture of Post-Natal Mouse Cerebellar Granule Neuron Progenitor Cells and Neurons 

Isolation and Culture of Avian Embryonic Valvular Progenitor Cells

鳥類胚弁膜前駆細胞の単離と培養

Proper formation and function of embryonic heart valves is critical for developmental progression. The early embryonic heart is a U-shaped tube of ...

Isolation and Culture of Cells from the Nephrogenic Zone of the Embryonic Mouse Kidney

マウス胎児の腎臓の腎性帯からの細胞の単離と培養

Embryonic development of the kidney has been extensively studied both as a model for epithelial-mesenchymal interaction in organogenesis and  to gain ...

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ラット肝細胞の単離及び初代培養

Primary hepatocyte culture is a valuable tool that has been extensively used in basic research of liver function, disease, pathophysiology, pharmacology ...

Serial Enrichment of Spermatogonial Stem and Progenitor Cells (SSCs) in Culture for Derivation of Long-term Adult Mouse SSC Lines

Serial Enrichment of Spermatogonial Stem and Progenitor Cells SSCs in Culture for Derivation of Long-term Adult Mouse SSC Lines

長期成体マウスのSSCラインの導出のために文化の精原幹細胞や前駆細胞の濃縮シリアル（SSCS）

Spermatogonial stem and progenitor cells (SSCs) of the testis represent a classic example of adult mammalian stem cells and preserve fertility for nearly ...

Isolation and Culture of Neonatal Mouse Cardiomyocytes

新生仔マウス心筋細胞の単離および培養

Cultured neonatal cardiomyocytes have long been used to study  myofibrillogenesis and myofibrillar functions. Cultured cardiomyocytes allow  for easy ...

Isolation, Culture, and Functional Characterization of Adult Mouse Cardiomyoctyes

アイソレーション、文化、および成体マウスCardiomyoctyesの機能解析

The use of primary cardiomyocytes (CMs) in culture has provided a powerful complement to murine models of heart disease in advancing our understanding of ...

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マウスプライマリ膵臓腺房細胞の単離と培養

This protocol permits rapid isolation (in less than 1 hr) of murine pancreatic acini, making it possible to maintain them in culture for more than one ...

Isolation, Culture, and Transplantation of Muscle Satellite Cells

筋衛星細胞の単離、培養、および移植

Muscle satellite cells are a stem cell population required for postnatal skeletal muscle development and regeneration, accounting for 2-5% of sublaminal ...

Isolation and Culture of Dental Epithelial Stem Cells from the Adult Mouse Incisor

成体マウス切歯から歯科上皮幹細胞の単離および培養

Understanding the cellular and molecular mechanisms that underlie tooth regeneration and renewal has become a topic of great interest1-4, and the mouse ...

Isolation and Primary Culture of Mouse Aortic Endothelial Cells

マウス大動脈内皮細胞の単離および初代培養

The vascular endothelium is essential to normal vascular homeostasis. Its dysfunction participates in various cardiovascular disorders. The mouse is an ...