وأيد هذا العمل من قبل المعاهد الوطنية للصحة منح K08NS065157 (إلى TAF) وبالإضافة إلى ذلك، فإن مركز جامعة بنسلفانيا لاضطرابات العظام والعضلات، جائزة P30AR050950 عدد من المعهد الوطني لالتهاب المفاصل، أمراض العضلات والعظام ودعمت الجلد، وهذا العمل (TAF وستيفن شيرر S.). وفي الختام، أيد Shriners الأطفال البذور مركز تمويل البحوث (TAF) هذا العمل. نود أن نعترف الدكتور يونغ جين الابن لتظاهرهم في البداية كل إجراء جبل ايمي وكيم ألف لتقديم المساعدة لها في انتاج الرسومات من الشكل رقم 1.

<ol>
	<li>Pan, Y. A., Misgeld, T., Lichtman, J. W., Sanes, 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=Effects+of+neurotoxic+and+neuroprotective+agents+on+peripheral+nerve+regeneration+assayed+by+time-lapse+imaging+in+vivo.">Effects of neurotoxic and neuroprotective agents on peripheral nerve regeneration assayed by time-lapse imaging in vivo.</a> J. Neurosci. 23, 11479-11488 (2003).</li><li>Magill, C., Tong, A., Kawamura, D., Hayashi, A., Hunter, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reinnervation+of+the+tibialis+anterior+following+sciatic+nerve+crush+injury:+A+confocal+microscopic+study+in+transgenic+mice.">Reinnervation of the tibialis anterior following sciatic nerve crush injury: A confocal microscopic study in transgenic mice.</a> Exp. Neurol. 207, 64-74 (2007).</li><li>Amado, S., Sim&#241;es, M. J., Armada-da-Silva, P. A. S., Lu&#237;s, A. L., Shirosaki, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Use+of+hybrid+chitosan+membranes+and+N1E-115+cells+for+promoting+nerve+regeneration+in+an+axonotmesis+rat+model.">Use of hybrid chitosan membranes and N1E-115 cells for promoting nerve regeneration in an axonotmesis rat model.</a> Biomaterials. 29, 4409-4419 (2008).</li><li>Lu&#237;s, A. L., Rodrigues, J. M., Geuna, S., Amado, S., Sim&#240;es, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neural+cell+transplantation+effects+on+sciatic+nerve+regeneration+after+a+standardized+crush+injury+in+the+rat.">Neural cell transplantation effects on sciatic nerve regeneration after a standardized crush injury in the rat.</a> Microsurgery. 28, 458-470 (2008).</li><li>Lu&#237;s, A. L., Amado, S., Geuna, S., Rodrigues, J. M., Sim&#240;es, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-term+functional+and+morphological+assessment+of+a+standardized+rat+sciatic+nerve+crush+injury+with+a+non-serrated+clamp.">Long-term functional and morphological assessment of a standardized rat sciatic nerve crush injury with a non-serrated clamp.</a> J. Neurosci. Methods. 163, 92-104 (2007).</li><li>Baptista, A. F., de Souza Gomes, J. R., Oliveira, J. T., Santos, S. M. G., Vannier-Santos, M. 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+new+approach+to+assess+function+after+sciatic+nerve+lesion+in+the+mouse+-+adaptation+of+the+sciatic+static+index.">A new approach to assess function after sciatic nerve lesion in the mouse - adaptation of the sciatic static index.</a> J. Neurosci. Methods. 161, 259-264 (2007).</li><li>Ronchi, G., Raimondo, S., Varej&#227;o, A. S. P., Tos, P., Perroteau, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Standardized+crush+injury+of+the+mouse+median+nerve.">Standardized crush injury of the mouse median nerve.</a> J. Neurosci. Methods. 188, 71-75 (2010).</li><li>Berg, A., Zelano, J., Cullheim, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Netrin+G-2+ligand+mRNA+is+downregulated+in+spinal+motoneurons+after+sciatic+nerve+lesion.">Netrin G-2 ligand mRNA is downregulated in spinal motoneurons after sciatic nerve lesion.</a> Neuroreport. 21, 782-785 (2010).</li><li>Girolami, E. I., Bouhy, D., Haber, M., Johnson, H., David, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Differential+expression+and+potential+role+of+SOCS1+and+SOCS3+in+Wallerian+degeneration+in+injured+peripheral+nerve.">Differential expression and potential role of SOCS1 and SOCS3 in Wallerian degeneration in injured peripheral nerve.</a> Exp. Neurol. 223, 173-182 (2010).</li><li>Hossain-Ibrahim, M. K., Rezajooi, K., Stallcup, W. B., Lieberman, A. R., Anderson, P. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analysis+of+axonal+regeneration+in+the+central+and+peripheral+nervous+systems+of+the+NG2-deficient+mouse.">Analysis of axonal regeneration in the central and peripheral nervous systems of the NG2-deficient mouse.</a> BMC Neurosci. 8, 80-80 (2007).</li><li>Thornton, M. R., Mantovani, C., Birchall, M. A., Terenghi, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantification+of+N-CAM+and+N-cadherin+expression+in+axotomized+and+crushed+rat+sciatic+nerve.">Quantification of N-CAM and N-cadherin expression in axotomized and crushed rat sciatic nerve.</a> J. Anat. 206, 69-78 (2005).</li><li>Beer, G. M., Steurer, J., Meyer, V. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Standardizing+nerve+crushes+with+a+non-serrated+clamp.">Standardizing nerve crushes with a non-serrated clamp.</a> J. Reconstr. Microsurg. 17, 531-534 (2001).</li></ol>

ليس لدينا ما يكشف.

قدمنا ​​وسيلة للحصول على كامل موثوق سحق العصب الوركي مع الدقيق وسم من موقع التدافع. كما ذكر سابقا، الوركي سحق العصب هو نموذج شائع للإصابة الأعصاب الطرفية في الفئران والجرذان. على الرغم من أن كل طريقة من سحق له مزاياه وعيوبه، وجدنا هذه الطريقة تنتج سحق الكاملة التي ت?...

<table border="1" style=";text-align:right;direction:rtl"><tbody><tr><td> اسم (وصف / كمية) </td><td> المزود </td><td> فهرس العدد </td></tr><tr><td> مصغرة مع المقص رقم 0000 بليد </td><td> Roboz </td><td> RC-5903 </td></tr><tr><td> نير مزيل الشعر (9 أونصة.) </td><td> الكنيسة وشركاه دوايت، المؤتمر الوطني العراقي. </td><td> N / A </td></tr><tr><td> فرك Betadine الجراحية (1 جالون) </td><td> فيشر العلمية </td><td> 19066452 </td></tr><tr><td> مرهم للعين (1 أوقية). </td><td> فيشر العلمية </td><td> 19082795 </td></tr></tbody></table> أدوات جراحية: <table border="1" style=";text-align:right;direction:rtl"><tbody><tr><td> اسم (وصف / كمية) </td><td> المزود </td><td> فهرس العدد </td></tr><tr><td> FST 250 حار حبة معقم </td><td> أدوات العلم غرامة </td><td> 18000-45 </td> &lt;/ TR&gt; <tr><td> قزحية مقص (11 سنتيمترا) </td><td> الآلات الدقيقة العالم </td><td> 500216 </td></tr><tr><td> بوتس سميث الملقط (مستقيم؛ 18cm؛ 1X2 الأسنان) </td><td> أدوات العلم غرامة </td><td> 11024-18 </td></tr><tr><td> مكفرسون، Vannas مقص (ريش ملم 5) </td><td> الآلات الدقيقة العالم </td><td> 14124-G </td></tr><tr><td> دومون # 5 الملقط - Dumoxel ستاندرد تلميح </td><td> أدوات العلم غرامة </td><td> 11252-30 </td></tr><tr><td> دومون # 5/45 الملقط - Dumoxel ستاندرد تلميح </td><td> أدوات العلم غرامة </td><td> 11251-35 </td></tr><tr><td> المرقأة الجميلة جدا (مستقيم؛ السطوح الملساء في الداخل) </td><td> أدوات العلم غرامة </td><td> 13020-12 </td></tr><tr><td> مسحوق الكربون المنشط (500 غرام) </td><td> فيشر العلمية </td><td> C272-500 </td></tr><tr><td> حجم 6-0 خيوط مع C-1 الإبرة (العقيمة، الحرير، أسود، مزين،غير قابل للامتصاص، 18 &quot;طول؛ صندوق من 36) </td><td> Roboz </td><td> SUT، 1073-1011 </td></tr><tr><td> منعكس مطبق كليب (لمدة 9 مقاطع مم) </td><td> الآلات الدقيقة العالم </td><td> 500345 </td></tr><tr><td> 9MM الفولاذ المقاوم للصدأ لقطات رد الفعل (100/box) </td><td> الآلات الدقيقة العالم </td><td> 500346 </td></tr></tbody></table> رعاية الحيوان: <table border="1" style=";text-align:right;direction:rtl"><tbody><tr><td> اسم (وصف / كمية) </td><td> المزود </td><td> فهرس العدد </td></tr><tr><td> 0.9٪ كلوريد الصوديوم حقن (مجاني الحافظة، 20 مل) </td><td> هوسبيرا </td><td> 0409-4888-20 </td></tr><tr><td> نظام كامل مع التحقيق غطاء Homeothermic مرنة (متوسطة، و 115 فولت، 60 هرتز) </td><td> هارفارد جهاز </td><td> 507222F </td></tr></tbody></table> الجراحية المنهاجم والكامشات: (تم شراؤها في متجر لاجهزة الكمبيوتر المحلي) <table border="1" style=";text-align:right;direction:rtl"><tbody><tr><td> اسم (وصف /) </td><td> كمية </td></tr><tr><td> الفولاذ المقاوم للصدأ منهاج (ناعم، ~ 12 &quot;X12&quot; X1 / 8 &quot;) </td><td> 1 </td></tr><tr><td> مغناطيسات زر (مثال: الكسوف E825) </td><td> 3 </td></tr><tr><td> مسامير الصلب غير القابل للصدأ (3 &quot;لونغ، القطر وتصميما من قبل مغناطيسات زر) </td><td> 3 </td></tr><tr><td> غير القابل للصدأ البندق الصلب (الحجم لاحتواء مسامير) </td><td> 9 </td></tr><tr><td> الأربطة المطاطية (التوتر الخفيفة) </td><td> 3 </td></tr><tr><td> حشرة الدبابيس (كلاهما ينتهي بينت إلى نموذج خطاف) </td><td> 3 </td></tr></tbody></table> شبه يخفف: <table border="1" style=";text-align:right;direction:rtl"><tbody><tr><td> اسم (وصف / كمية) </td><td> المورد &lt;/ قوي&gt; </td><td> فهرس العدد </td></tr><tr><td> لامتصاص العرق (1 كلغ) </td><td> سيغما الدريخ </td><td> P6148-1KG </td></tr><tr><td> ثنائي القاعدة فوسفات الصوديوم اللامائية (500 غرام، وتستخدم لإعداد العازلة الفوسفات) </td><td> فيشر العلمية </td><td> S375-500 </td></tr><tr><td> صوديوم فوسفات أحادى اللامائى (1 كغم؛ تستخدم لإعداد العازلة الفوسفات) </td><td> فيشر العلمية </td><td> AC38987-0010 </td></tr><tr><td> غلوتارالدهيد (50٪، 10 × 10 مل) </td><td> تيد بيلا، شركة </td><td> 18431 </td></tr><tr><td> رباعي أكسيد الأوزميوم (مائي 4٪، 10 × 10 مل) </td><td> تيد بيلا، شركة </td><td> 18465 </td></tr><tr><td> أكسيد البروبيلين (450 مل) </td><td> تيد بيلا، شركة </td><td> 18601 </td></tr><tr><td> تضمين 812 (كيت، لكتل ​​الصلب / عالية التباين صورة) </td><td> علوم المجهر الإلكتروني </td><td> 14120 </td></tr><tr><td> Toluidiشمال شرق الزرقاء (25 ز) </td><td> تيد بيلا، شركة </td><td> 19451 </td></tr></tbody></table> كامل تحضير العضلات وجبل المناعية: <table border="1" style=";text-align:right;direction:rtl"><tbody><tr><td> اسم (وصف / كمية) </td><td> المزود </td><td> فهرس العدد </td></tr><tr><td> لامتصاص العرق (1 كلغ) </td><td> سيغما الدريخ </td><td> P6148-1KG </td></tr><tr><td> ثنائي القاعدة فوسفات الصوديوم اللامائية (500 غرام، وتستخدم لإعداد العازلة الفوسفات) </td><td> فيشر العلمية </td><td> S375-500 </td></tr><tr><td> صوديوم فوسفات أحادى اللامائى (1 كغم؛ تستخدم لإعداد العازلة الفوسفات) </td><td> فيشر العلمية </td><td> AC38987-0010 </td></tr><tr><td> جيش صرب البوسنة 10٪ مخفف / الحل حجب (200 مل) </td><td> Kirkegaard بيري والمختبرات، وشركة </td><td> 50-61-00 </td></tr><tr><td> تريتون X-100 (100 مل) </td&gt; <td> نقطة العلمية للإعلام </td><td> 9002-93-1 </td></tr><tr><td> جليكاين، 98٪ (1 كلغ) </td><td> فيشر العلمية </td><td> AC12007-0010 </td></tr><tr><td> الأنسجة تيك البرد أكتوبر مجمع (حالة من 12، 4 زجاجات أونصة) </td><td> فيشر العلمية </td><td> 14-373-65 </td></tr><tr><td> Sylgard DOW 170 (2 كيت بالرطل) </td><td> فيشر العلمية </td><td> NC9492579 </td></tr><tr><td> غير القابل للصدأ الدبابيس الحشرات الصلب، حجم 1 (100/pkg) </td><td> أدوات العلم غرامة </td><td> 26001-40 </td></tr><tr><td> Tetramethylrhodamine-A-بنغاروتوكسين (0.5 ملغ) </td><td> سيغما الدريخ </td><td> T0195-.5MG </td></tr><tr><td> جسم الفأر وحيدة النسيلة ضد SMI-312 (0.1 مل) </td><td> كوفانس </td><td> SMI-312R </td></tr><tr><td> جسم الفأر وحيدة النسيلة ضد SV2 (0.1 مل) </td><td> الدراسات التنموية ورم هجين بنك (DSHB) </td><td> SV2 </td></tr><tr><td> أرنب بولينسيلي الأجسام المضادة ضد 43-GAP </td><td> نوفوس البيولوجية </td><td> NB300-143 </td></tr><tr><td> Flourescein مترافق عنزة المضادة للماوس مفتش الحكومة، Fcγ الفئة الفرعية 1 محددة </td><td> جاكسون ImmunoResearch </td><td> <a href="http://www.jacksonimmuno.com/MERCHANT2/merchant.mv?Screen=BASK&amp;Store_Code=JI&amp;Action=ADPR&amp;Product_Code=115-095-205&amp;Attributes=Yes&amp;Quantity=1">11 5-095-205</a> </td></tr><tr><td> DyLight 649 مترافق حمار المضادة للأرنب مفتش </td><td> جاكسون ImmunoResearch </td><td> <a href="http://www.jacksonimmuno.com/MERCHANT2/merchant.mv?Screen=BASK&amp;Store_Code=JI&amp;Action=ADPR&amp;Product_Code=711-495-152&amp;Attributes=Yes&amp;Quantity=1">711-495-152</a> </td></tr><tr><td> 4 &#39;،6-diamidino-2-phenylindole (دابي، dilactate، و 10 ملغ) </td><td> إينفيتروجن </td><td> D3571 </TD&gt; </tr><tr><td> Vectashield التركيب متوسطة (10 مل) </td><td> ناقلات مختبرات </td><td> H-1000 </td></tr><tr><td> Superfrost زائد شرائح المجهر (الأبيض، الحجم: 75 × 25 مم، حزمة من 144) </td><td> فيشر العلمية </td><td> 12-550-15 </td></tr><tr><td> Fisherfinest النظارات غطاء بريميوم (الحجم: 40 × 22 مم؛ حزمة من 1 أوقية.) </td><td> فيشر العلمية </td><td> 12-548-5C </td></tr></tbody></table>

reproducible mouse sciatic nerve crush and subsequent assessment of regeneration by whole mount muscle analysis

تدرس على نطاق واسع في تجديد النظام العصبي الطرفي (المحيطي) على حد سواء لأهميتها بالنسبة لمرض الإنسان وفهم ردا قويا على التجدد التي تنفذها الخلايا العصبية الجهاز العصبي المحيطي وبالتالي إلقاء الضوء ربما فشل تجديد الجهاز العصبي المركزي 1. الوركي سحق العصبية (انهتاك المحوار) هو واحد من النماذج الأكثر شيوعا لإصابة الأعصاب الطرفية في القوارض 2. سحق المقاطعات هي الحفاظ على جميع المحاور لكن شوان الصفائح الخلايا القاعدية بحيث التجديد هو الأمثل 3،4. هذا يسمح للمحقق على وجه التحديد لدراسة قدرة محور عصبي متزايد للتفاعل مع كل من خلايا شوان والصفيحة القاعدية 4. وكانت الفئران عموما على نماذج حيوانية المفضل لسحق العصب التجريبية. وهي متاحة على نطاق واسع وأعصابهم الوركي lesioned يوفر تقريب معقول من الآفات العصبية الإنسان 5،4. على الرغم من أصغر في الحجم من العصب الفئران، العصب الماوس لديه صفات كثيرة مماثلة. إلا أن الأهم، مذكرات تفاهمالنماذج الإلكترونية هي ذات قيمة متزايدة بسبب توافر واسعة من خطوط المعدلة وراثيا يسمح الآن لتشريح مفصل للجزيئات الفردية حاسمة لتجديد العصبية 6 و 7. وقد استخدم المحققون قبل أساليب متعددة لإنتاج سحق العصب أو إصابة بما في ذلك ملقط الزاوية بسيط، ملقط المبردة، ملقط مرقئ، والمشابك الأوعية الدموية، والمشابك محقق المصممة 8،9،10،11،12. وقد استخدم الباحثون أيضا أساليب مختلفة لوضع العلامات على موقع الإصابة بما في ذلك خياطة الجروح، وجزيئات الكربون والخرز فلوري 13،14،1. نحن لدينا وصف طريقة للحصول على كامل بتكاثر سحق العصب الوركي مع دقيق ومستمر من وضع علامات للموقع سحق باستخدام ملقط غرامة مرقئ والكربون لاحق سحق الموقع وسم. كجزء من الوصف الذي نعيش فيه من إجراء العصب الوركي سحق ادرجنا أيضا طريقة بسيطة نسبيا من كل عضلة جبل نستخدمها لتحديد وقت لاحق تجديد.

Watch this Scientific Journal Video about Reproducible Mouse Sciatic Nerve Crush and Subsequent Assessment of Regeneration by Whole Mount Muscle Analysis at JoVE.com

Reproducible Mouse Sciatic Nerve Crush and Subsequent Assessment of Regeneration by Whole Mount Muscle Analysis

نحن في هذا التقرير وصفا لطريقة لسحق العصب الوركي الماوس. هذا الأسلوب يستخدم الملقط مرقئ متاحة بسهولة ويسر، وبتكاثر تنتج كاملة سحق العصب الوركي. وبالإضافة إلى ذلك، ونحن تصف طريقة لإعداد يتصاعد العضلات كلها مناسبة للتحليل من تجديد العصب بعد سحق العصب الوركي.

استنساخه سحق الفأر العصب الوركي وتقييم لاحق للتجديد من قبل الجامعة جبل تحليل العضلات

Regeneration in the peripheral nervous system (PNS) is widely studied both for its relevance to human disease and to understand the robust regenerative ...

reproducible-mouse-sciatic-nerve-crush-subsequent-assessment

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33.4K Views.  Temple University. نحن في هذا التقرير وصفا لطريقة لسحق العصب الوركي الماوس. هذا الأسلوب يستخدم الملقط مرقئ متاحة بسهولة ويسر، وبتكاثر تنتج كاملة سحق العصب الوركي. وبالإضافة إلى ذلك، ونحن تصف طريقة لإعداد يتصاعد العضلات كلها مناسبة للتحليل من تجديد العصب بعد سحق العصب الوركي.

Video: Reproducible Mouse Sciatic Nerve Crush and Subsequent Assessment of Regeneration by Whole Mount Muscle Analysis

Axoplasm Isolation from Rat Sciatic Nerve

Isolation of pure axonal cytoplasm (axoplasm) from peripheral nerve is crucial for biochemical studies of many biological processes. In this article, we demonstrate and describe a protocol for axoplasm isolation from adult rat sciatic nerve based on the following steps: (1) dissection of nerve fascicles and separation of connective tissue; (2) incubation of short segments of nerve fascicles in hypotonic medium to release myelin and lyse non-axonal structures; and (3) extraction of the remaining axon-enriched material. Proteomic and biochemical characterization of this preparation has confirmed a high degree of enrichment for axonal components.

We demonstrate a protocol for axoplasm isolation from adult rat sciatic nerve based on dissection of nerve fascicles and incubation in hypotonic medium to release myelin and lyse non-axonal structures, followed by extraction of the remaining axon-enriched material.

جبلة المحوار من عزل العصب الوركي الجرذ

Isolation of pure axonal cytoplasm (axoplasm) from peripheral nerve is crucial for biochemical studies of many biological processes. In this article, we ...

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

To analyze the axonal and dendritic morphology of neurons, it is essential to obtain accurate labeling of neuronal structures. Preparing well labeled samples with little to no tissue damage enables us to analyze cell morphology and to compare individual samples to each other, hence allowing the identification of mutant anomalies.
In the demonstrated dissection method the nervous system remains mostly inside the adult fly. Through a dorsal incision, the abdomen and thorax are opened and most of the internal organs are removed. Only the dorsal side of the ventral nerve cord (VNC) and the cervical connective 
(CvC) containing the big axons of the giant fibers (GFs)1 are exposed, while the brain containing the GF cell body and dendrites remains2 in the 
intact head. In this preparation most nerves of the VNC should remain attached to their muscles. 
Following the dissection, the intracellular filling of the giant fiber (GF) with a fluorescent dye is demonstrated. In the CvC the GF axons are located at the dorsal surface and thus can be easily visualized under a microscope with differential interference contrast (DIC) optics. This allows the injection of the GF axons with dye at this site to label the entire GF including the axons and their terminals in the VNC. This method results in reliable and strong staining of the GFs allowing the neurons to be imaged immediately after filling with an epifluorescent microscope. Alternatively, the fluorescent signal can be enhanced using standard immunohistochemistry procedures3 suitable for high resolution confocal microscopy.

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

An in vivo dissection of the adult Drosophila ventral nerve cord (VNC) is demonstrated. This particular dissection method causes little damage to the VNC allowing the subsequent labeling of the giant fiber neurons with fluorescent dye for high resolution imaging.

كله من جبل إعداد الكبار ذبابة الفاكهة الحبل الاعصاب الاماميه للحقن صبغ الألياف العملاق

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

Dissection of the Transversus Abdominis Muscle for Whole-mount Neuromuscular Junction Analysis

Analysis of neuromuscular junction morphology can give important insight into the physiological status of a given motor neuron. Analysis of thin flat muscles can offer significant advantage over traditionally used thicker muscles, such as those from the hind limb (e.g. gastrocnemius). Thin muscles allow for comprehensive overview of the entire innervation pattern for a given muscle, which in turn permits identification of selectively vulnerable pools of motor neurons. These muscles also allow analysis of parameters such as motor unit size, axonal branching, and terminal/nodal sprouting. A common obstacle in using such muscles is gaining the technical expertise to dissect them. In this video, we detail the protocol for dissecting the transversus abdominis (TVA) muscle from young mice and performing immunofluorescence to visualize axons and neuromuscular junctions (NMJs). We demonstrate that this technique gives a complete overview of the innervation pattern of the TVA muscle&#160;and can be used to investigate NMJ pathology in a mouse model of the childhood motor neuron disease, spinal muscular atrophy.

Dissection of the Transversus Abdominis Muscle for Whole-mount Neuromuscular Junction Analysis

In this video we demonstrate a protocol for dissection of the transversus abdominis muscle of the mouse and use immunofluorescence and microscopy to visualize neuromuscular junctions.

تشريح العضلات Abdominis المستعرض لتحليل تقاطع العصبية والعضلية جبل كامل

Analysis of neuromuscular junction morphology can give important insight into the physiological status of a given motor neuron. Analysis of thin flat ...

Whole-mount Imaging of Mouse Embryo Sensory Axon Projections

The visualization of full-length neuronal projections in embryos is essential to gain an understanding of how mammalian neuronal networks develop. Here we describe a method to label in situ a subset of dorsal root ganglion (DRG) axon projections to assess their phenotypic characteristics using several genetically manipulated mouse lines. The TrkA-positive neurons are nociceptor neurons, dedicated to the transmission of pain signals. We utilize a TrkAtaulacZ mouse line to label the trajectories of all TrkA-positive peripheral axons in the intact mouse embryo. We further breed the TrkAtaulacZ line onto a Bax null background, which essentially abolishes neuronal apoptosis, in order to assess growth-related questions independently of possible effects of genetic manipulations on neuronal survival. Subsequently, genetically modified mice of interest are bred with the TrkAtaulacZ/Bax null line and are then ready for study using the techniques described herein. This presentation includes detailed information on mouse breeding plans, genotyping at the time of dissection, tissue preparation, staining and clearing to allow for visualization of full-length axonal trajectories in whole-mount preparation.

We present here an optimized protocol to genotype, stain and prepare fetal mice for the imaging of peripheral nociceptor axon projections in the whole animal, as an effective method to assess sensory axon growth phenotypes in developing genetically engineered mice.

كله جبل التصوير من جنين فأر التوقعات الحسية أكسون

The visualization of full-length neuronal projections in embryos is essential to gain an understanding of how mammalian neuronal networks develop. Here we ...

Whole Mount Dissection and Immunofluorescence of the Adult Mouse Cochlea

The organ of Corti, housed in the cochlea of the inner ear, contains mechanosensory hair cells and surrounding supporting cells which are organized in a spiral shape and have a tonotopic gradient for sound detection. The mouse cochlea is approximately 6 mm long and often divided into three turns (apex, middle, and base) for analysis. To investigate cell loss, cell division, or mosaic gene expression, the whole mount or surface preparation of the cochlea is useful. This dissection method allows visualization of all cells within the organ of Corti when combined with immunostaining and confocal microscopy to image cells at different planes in the z-axis. Multiple optical cross-sections can also be obtained from these z-stack images. In addition, the whole mount dissection method can be used for scanning electron microscopy, although a different fixation method is needed. Here, we present a method to isolate the organ of Corti as three intact cochlear turns (apex, middle, and base). This method can be used for mice ranging from one week of age through adulthood and differs from the technique used for neonatal samples where calcification of the cochlea is incomplete. A slightly modified version can be used for dissection of the rat cochlea. We also demonstrate a procedure for immunostaining with fluorescently tagged antibodies.

We present a method to isolate the adult organ of Corti as three intact cochlear turns (apex, middle, and base). We also demonstrate a procedure for immunostaining with fluorescently tagged antibodies. Together these techniques allow the study of hair cells, supporting cells, and other cell types found in the cochlea.

جبل كله تشريح والمناعي من الكبار ماوس القوقعة

The organ of Corti, housed in the cochlea of the inner ear, contains mechanosensory hair cells and surrounding supporting cells which are organized in a ...

In Vivo Gene Transfer to Schwann Cells in the Rodent Sciatic Nerve by Electroporation

The formation of the myelin sheath by Schwann cells (SCs) is essential for rapid conduction of nerve impulses along axons in the peripheral nervous system. SC-selective genetic manipulation in living animals is a powerful technique for studying the molecular and cellular mechanisms of SC myelination and demyelination in vivo. While knockout/knockin and transgenic mice are powerful tools for studying SC biology, these methods are costly and time consuming. Viral vector-mediated transgene introduction into the sciatic nerve is a simpler and less laborious method. However, viral methods have limitations, such as toxicity, transgene size constraints, and infectivity restricted to certain developmental stages. Here, we describe a new method that allows selective transfection of myelinating SCs in the rodent sciatic nerve using electroporation. By applying electric pulses to the sciatic nerve at the site of plasmid DNA injection, genes of interest can be easily silenced or overexpressed in SCs in both neonatal and more mature animals. Furthermore, this in vivo electroporation method allows for highly efficient simultaneous expression of multiple transgenes. Our novel technique should enable researchers to efficiently manipulate SC gene expression, and facilitate studies on SC development and function.

In Vivo Gene Transfer to Schwann Cells in the Rodent Sciatic Nerve by Electroporation

Here, we present an in vivo technique for gene transfer to Schwann cells (SCs) in the rodent sciatic nerve. This simple technique is useful for investigating signaling mechanisms involved in the development and maintenance of myelinating SCs.

في نقل فيفو جين لخلايا شوان في العصب القوارض وركي من Electroporation لل

The formation of the myelin sheath by Schwann cells (SCs) is essential for rapid conduction of nerve impulses along axons in the peripheral nervous ...

3D Kinematic Analysis for the Functional Evaluation in the Rat Model of Sciatic Nerve Crush Injury

Compared to the Sciatic Functional Index (SFI), kinematic analysis is a more reliable and sensitive method for performing functional evaluations of sciatic nerve injury rodent models. In this protocol, we describe a novel kinematic analysis method that uses a three-dimensional (3D) motion capture apparatus for functional evaluations using a rat sciatic nerve crush injury model. First, the rat is familiarized with treadmill walking. Markers are then attached to the designated bone landmarks and the rat is made to walk on the treadmill at the desired speed. Meanwhile, the posterior limb movements of the rat are recorded using four cameras. Depending on the software used, marker tracings are created using both automatic and manual modes and the desired data are produced after subtle adjustments. This method of kinematic analysis, which uses a 3D motion capture apparatus, offers numerous advantages, including superior precision and accuracy. Many more parameters can be investigated during the comprehensive functional evaluations. This method has several shortcomings that require consideration: The system is expensive, can be complicated to operate, and may produce data deviations due to skin shifting. Nevertheless, kinematic analysis using a 3D motion capture apparatus is useful for performing functional anterior and posterior limb evaluations. In the future, this method may become increasingly useful for generating accurate assessments of various traumas and diseases.

We introduce a kinematic analysis method that uses a three-dimensional motion capture apparatus containing four cameras and data processing software for performing functional evaluations during fundamental research involving rodent models.

3D تحليل الحركية للتقييم الوظيفي في نموذج الفئران من عرق النسا الأعصاب سحق الإصابة

Compared to the Sciatic Functional Index (SFI), kinematic analysis is a more reliable and sensitive method for performing functional evaluations of ...

Whole-Mount Staining, Visualization, and Analysis of Fungiform, Circumvallate, and Palate Taste Buds

Taste buds are collections of taste-transducing cells specialized to detect subsets of chemical stimuli in the oral cavity. These transducing cells communicate with nerve fibers that carry this information to the brain. Because taste-transducing cells continuously die and are replaced throughout adulthood, the taste-bud environment is both complex and dynamic, requiring detailed analyses of its cell types, their locations, and any physical relationships between them. Detailed analyses have been limited by tongue-tissue heterogeneity and density that have significantly reduced antibody permeability. These obstacles require sectioning protocols that result in splitting taste buds across sections so that measurements are only approximated, and cell relationships are lost. To overcome these challenges, the methods described herein involve collecting, imaging, and analyzing whole taste buds and individual terminal arbors from three taste regions: fungiform papillae, circumvallate papillae, and the palate. Collecting whole taste buds reduces bias and technical variability and can be used to report absolute numbers for features including taste-bud volume, total taste-bud innervation, transducing-cell counts, and the morphology of individual terminal arbors. To demonstrate the advantages of this method, this paper provides comparisons of taste bud and innervation volumes between fungiform and circumvallate taste buds using a general taste-bud marker and a label for all taste fibers. A workflow for the use of sparse-cell genetic labeling of taste neurons (with labeled subsets of taste-transducing cells) is also provided. This workflow analyzes the structures of individual taste-nerve arbors, cell type numbers, and the physical relationships between cells using image analysis software. Together, these workflows provide a novel approach for tissue preparation and analysis of both whole taste buds and the complete morphology of their innervating arbors.

This paper describes methods for tissue preparation, staining, and analysis of whole fungiform, circumvallate, and palate taste buds that consistently yield whole and intact taste buds (including the nerve fibers that innervate them) and maintain the relationships between structures within taste buds and the surrounding papilla.

كامل جبل تلطيخ، والتصور، وتحليل الفطريات، Circumvallate، وبراعم طعم الحنك

Taste buds are collections of taste-transducing cells specialized to detect subsets of chemical stimuli in the oral cavity. These transducing cells ...

Multifactorial Assessment of Motor Behavior in Rats after Unilateral Sciatic Nerve Crush Injury

The induction of a peripheral nerve injury is a widely used method in neuroscience for the assessment of repair and pain mechanisms among others. In addition, in the research field of movement disorders, sciatic crush injury has been employed to trigger a dystonia-like phenotype in genetically predisposed DYT-TOR1A rodent models of dystonia. To achieve consistent, reproducible and comparable results after a sciatic nerve crush injury, a standardized method for inducing the nerve crush is essential, in addition to a standardized phenotypical characterization. Attention must be paid not only to the specific assortment of behavioral tests, but also to the technical requirements, the correct execution and consecutive data analysis. This protocol describes in detail how to perform a sciatic nerve crush injury and provides a behavioral test battery for the assessment of motor deficits in rats that includes the open field test, the CatWalk XT gait analysis, the beam walking task, and the ladder rung walking task.

We provide a protocol for the assessment of motor behavior via a behavioral test battery in rats after sciatic nerve crush injury.

تقييم متعدد العوامل للسلوك الحركي في الفئران بعد إصابة سحق العصب الوركي من جانب واحد

The induction of a peripheral nerve injury is a widely used method in neuroscience for the assessment of repair and pain mechanisms among others. In ...

Dissection of Single Skeletal Muscle Fibers for Immunofluorescent and Morphometric Analyses of Whole-Mount Neuromuscular Junctions

The neuromuscular junction (NMJ) is a specialized point of contact between the motor nerve and the skeletal muscle. This peripheral synapse exhibits high morphological and functional plasticity. In numerous nervous system disorders, NMJ is an early pathological target resulting in neurotransmission failure, weakness, atrophy, and even in muscle fiber death. Due to its relevance, the possibility to quantitatively assess certain aspects of the relationship between NMJ components can help to understand the processes associated with its assembly/disassembly. The first obstacle when working with muscles is to gain the technical expertise to quickly identify and dissect without damaging their fibers. The second challenge is to utilize high-quality detection methods to obtain NMJ images that can be used to perform quantitative analysis. This article presents a step-by-step protocol for dissecting extensor digitorum longus and soleus muscles from rats. It also explains the use of immunofluorescence to visualize pre and postsynaptic elements of whole-mount NMJs. Results obtained demonstrate that this technique can be used to establish the microscopic anatomy of the synapsis and identify subtle changes in the status of some of its components under physiological or pathological conditions.

The ability to accurately detect neuromuscular junction components is crucial in evaluating modifications in its architecture because of pathological or developmental processes. Here we present a complete description of a straightforward method to obtain high-quality images of whole-mount neuromuscular junctions that can be used to perform quantitative measurements.

تشريح ألياف العضلات الهيكل العظمي واحد للتحليلات Immunofluorescent ومورفومتري من تقاطعات العضلات العصبية جبل كامل

The neuromuscular junction (NMJ) is a specialized point of contact between the motor nerve and the skeletal muscle. This peripheral synapse exhibits high ...