وأيد هذا العمل من قبل المعاهد الوطنية للصحة منح NS057195 ، UCSF راكوشبالوتاي الجائزة ، وجامعة كينت ستيت بدء صندوق لWH تشو.

<html> 1. أسلوب MCAO وتمت الموافقة على هذا البروتوكول من قبل رعاية الحيوان المؤسسية واللجان استخدم في سان فرانسيسكو وجامعة كينت الدولة ، وتلتزم المعاهد الوطنية للصحة المبادئ التوجيهية لاستخدام حيوانات التجارب. <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> قطع خياطة حيدة 5-0 (جهاز هارفارد ، Holliston ، MA) في 20 قطاعات ملم. جولة غيض من كل قطعة بتسخينه بالقرب من cauterizer (برينتري العلمية ، وشركة ، برينتري ، MA). قياس قطرها من طرف باستخدام ميكرومتر (صورة التطبيقية ، وشركة روتشستر ، نيويورك). نستخدم خياطة مع قطر الحافة النهائية لل0،21-0،22 مم للماوس مع وزن الجسم من 25-30 غرام. </li><li style=";text-align:right;direction:rtl"> تعقيم جميع الأدوات الجراحية بواسطة التعقيم (الحد الأدنى 121 درجة مئوية ، 15 PSI ، لمدة 15 دقيقة). تعقيم طاولة الجراحة والمعدات المرتبطة باستخدام الايثانول 70 ٪. </li><li style=";text-align:right;direction:rtl"> تخدير an 8-12 أسبوع من العمر الماوس (25-30 ز) مع 5 ٪ isoflurane (Aerrane ، باكستر ، ديرفيلد ، IL) في 30 ٪ O 2 / 70 ٪ N 2 O باستخدام نظام V - 10 التخدير (VetEquip ، المؤتمر الوطني العراقي . ، بليزانتون ، كاليفورنيا). بعد التخدير ، وخفض مستوى isoflurane والحفاظ عليه عند 1.5 ٪. </li><li style=";text-align:right;direction:rtl"> ضع الماوس في موقف ضعيف على وسادة التدفئة. ادخال مسبار المستقيم ، ورصد والمحافظة على درجة حرارة الجسم بين 36،5-37،5 درجة مئوية باستخدام نظام TR - 200 درجة حرارة homeothermic (العلوم أدوات الجميلة ، وشركة فوستر سيتي ، كاليفورنيا). </li><li style=";text-align:right;direction:rtl"> حلق الفراء على منطقة الرقبة بطني مع كليبرز الكهربائية (برينتري العلمية) لفضح الجلد. تطهير الموقع الجراحية باستخدام ثلاثة طلبات من الايثانول 70 ٪. </li><li style=";text-align:right;direction:rtl"> تحت المجهر ستيريو تشريح (نيكون ، اليابان) ، وجعل خط الوسط 1 سم شق طويل على الرقبة. استخدام الكامشات (برينتري العلمية) لفضح الجراحي الميداني والتعرف على حق الشريان السباتي المشترك (CCA) ، الشريان السباتي الخارجي (ECA) ، والشريان السباتي الداخلي (ICA). تشريح بعناية الشرايين خالية من الأعصاب المحيطة واللفافة. </li><li style=";text-align:right;direction:rtl"> تشريح اللجنة الاقتصادية لأفريقيا بشكل أقصى زيادة تخثر واللجنة الاقتصادية لأفريقيا في الغدة الدرقية والشريان متفوقة (STA) فرع باستخدام مخثار القطبين (هوارد الصك ، وشركة توسكالوسا ، AL). اللجنة الاقتصادية لأفريقيا وخفض STA في الجزء متخثر. </li><li style=";text-align:right;direction:rtl"> ربط خيوط الحرير two فضفاضة حول الجذع 8-0 اللجنة الاقتصادية لأفريقيا. تطبيق المشبك الأوعية الدموية (أدوات العلوم الجميلة) في التشعب من التقييم القطري المشترك في اللجنة الاقتصادية لأفريقيا والتحالف. </li><li style=";text-align:right;direction:rtl"> إجراء شق صغير في نهاية الجذع اللجنة الاقتصادية لأفريقيا مع مقص الربيع Vannas على غرار (أدوات العلوم الجميلة). وسجل قياس طول الخيط حيدة 5-0 مدورة على الحافة. تضاف الى خياطة الفتحة وتقدم إلى المشبك. تشديد خيوط الحرير two حول تجويف ما يكفي لضمان الحفاظ على بعد التنقل من الدرز حيدة داخل المسكن. </li><li style=";text-align:right;direction:rtl"> إزالة المشبك من التشعب. تقدم بلطف خياطة حيدة من التجويف للجنة الاقتصادية لأفريقيا في ICA لمسافة 9-10 ملم وراء انشعاب CCA لتسد أصل MCA. مدة الجراحة حوالي 30-45 دقيقة. </li><li style=";text-align:right;direction:rtl"> خياطة الفتحة على الرقبة ووضع الماوس في مربع 35 درجة مئوية التمريض للتعافي من التخدير ، وإعادته إلى القفص. يستغرق عادة 50-10 دقيقة للفئران للتعافي من التخدير. لأداء MCAO عابرة ، يمكن للباحث إعادة تخدير الماوس وسحب الخيط مرة أخرى إلى اللجنة الاقتصادية لأفريقيا جدعة بعد فترة من الزمن ، عادة ما بين 0،5-2 ساعة. </li><li style=";text-align:right;direction:rtl"> أربع وعشرين ساعة بعد تحريض MCAO ، تخدير الفأر مع 5 ٪ isoflurane والموت ببطء من قبل خلع عنق الرحم. قطع رأس الفأر وجمع الدماغ. شريحة الدماغ coronally إلى شرائح 2 ملم الأربع مع مصفوفة الدماغ (برينتري العلمية) على الجليد. احتضان شرائح الدماغ في 2 ٪ 2،3،5 - triphenyltetrazolium كلوريد (TTC) (سيغما الدريخ) في برنامج تلفزيوني لمدة 20 دقيقة 1X في درجة حرارة الغرفة لتحديد حجم ومدى احتشاء. إصلاح شرائح المخ في 10 ٪ محايدة مخزنة حل الفورمالين (سيغما الدريخ) في 4 درجات مئوية حتى التصوير. ويمكن قياس مدى احتشاء كما هو موضح في البروتوكول إن الرب 955 ( <a href="http://www.jove.com/index/Details.stp?ID=955">http://www.jove.com/index/Details.stp؟ID=955</a> ) 9. </li></ol> 2. ممثل النتائج وينظر للعوائق التي تولدها MCAO في المخطط والقشرة ظهراني. والمخطط هو أكثر حساسية لنقص تروية من القشرة الدماغية. وثلاثين دقيقة من إنتاج MCAO احتشاء فقط في المخطط ، بينما أكثر من ساعة واحدة من MCAO سيضر كل من المخطط والقشرة. بعد 24 ساعة MCAO دائمة ، فإن النسبة المئوية الإجمالية احتشاء ~ 40 ± 5 ٪ من الكرة الأرضية ومعدل وفيات بعد الجراحة ~ 10 ٪. استثنينا الفئران من إجراء المزيد من الدراسات إذا يحدث النزيف أثناء الجراحة ، ووقت تشغيل يتجاوز 90 دقيقة ، والفئران فشل في التعافي من التخدير في غضون 15 دقيقة ، أو وجدت نزيف في الدماغ أو شرائح في القاعدة من دائرة ويليس خلال الفحص بعد الوفاة . 61/2761fig1.jpg &quot;بديل =&quot; الشكل 1 &quot;/&gt; صور الممثل الشكل 1. شرائح الدماغ TTC الملطخة (المستوى الإكليلي 1-4) بعد 24 ساعة من MCAO دائمة. في المعيشة ينخفض ​​إنزيمي الأنسجة TTC بواسطة dehydrogenases لtriphenylformazan - 1،3،5 (TPF) ، وهو أحمر اللون ، بينما في المناطق نخرية يبقى الأبيض بسبب عدم وجود مثل هذا النشاط الأنزيمي. ولذلك ، يمكن تحديد المنطقة التي من احتشاء لونه أبيض بسبب عدم وجود لتحويل TTC ​​TPF. ملاحظة : TTC الحرارة والضوء إلى حد ما غير مستقر حتى المقاطع ملطخة حماية من الحرارة والضوء قدر الإمكان.

<ol>
	<li>Howard, G., Howard, V. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Distribution of stroke: hetrogeneity of stroke by age, and sex. , 3-12 (2004).</li><li>Thom, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heart+disease+and+stroke+statistics--2006+update:+a+report+from+the+American+Heart+Association+Statistics+Committee+and+Stroke+Statistics+Subcommittee.">Heart disease and stroke statistics--2006 update: a report from the American Heart Association Statistics Committee and Stroke Statistics Subcommittee.</a> Circulation. 113, e85-e151 (2006).</li><li>Mohr, J. 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> Middle cerebral artery disease. , 123-166 (2004).</li><li>Longa, E. Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reversible+middle+cerebral+artery+occlusion+without+craniectomy+in+rats.">Reversible middle cerebral artery occlusion without craniectomy in rats.</a> Stroke. 20, 84-91 (1989).</li><li>Chou, W. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neutrophil+protein+kinase+Cdelta+as+a+mediator+of+stroke-reperfusion+injury.">Neutrophil protein kinase Cdelta as a mediator of stroke-reperfusion injury.</a> J. Clin. Invest. 114, 49-56 (2004).</li><li>Hudgins, W. R., Garcia, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+electrocautery,+atmospheric+exposure,+and+surgical+retraction+on+the+permeability+of+the+blood-brain-barrier.">The effect of electrocautery, atmospheric exposure, and surgical retraction on the permeability of the blood-brain-barrier.</a> Stroke. 1, 375-380 (1970).</li><li>Carmichael, S. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rodent+models+of+focal+stroke:+size,+mechanism,+and+purpose.">Rodent models of focal stroke: size, mechanism, and purpose.</a> NeuroRx. 2, 396-409 (2005).</li><li>Durukan, A., Tatlisumak, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acute+ischemic+stroke:+overview+of+major+experimental+rodent+models,+pathophysiology,+and+therapy+of+focal+cerebral+ischemia.">Acute ischemic stroke: overview of major experimental rodent models, pathophysiology, and therapy of focal cerebral ischemia.</a> Pharmacol. Biochem. Behav. 87, 179-197 (2007).</li><li>Taniguchi, H., Andreasson, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+hypoxic-ischemic+encephalopathy+model+of+perinatal+ischemia.">The hypoxic-ischemic encephalopathy model of perinatal ischemia.</a> J Vis Exp. , (2008).</li></ol>

ليس لدينا أية مصالح متضاربة المحتملة في الكشف عنها.

ويشيع استخدام MCAO في الفئران لنقص تروية الدماغ نموذج التنسيق بين البشر. أصبح استخدام الفئران لدراسات السكتة الدماغية أكثر تواترا بسبب توافر سلالات معدلة وراثيا وخروج المغلوب. هناك بعض التفاصيل الهامة أن نلاحظ في البروتوكول : <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> فمن الضروري للحفاظ على درجة حرارة الجسم الماوس أثناء الجراحة وقبل أن يتعافى تماما من التخدير. درجة حرارة الجسم له تأثيرات على مدى احتشاء ؛ انخفاض درجة حرارة الجسم وارتفاع الحرارة يزيد من حجم احتشاء. </li><li style=";text-align:right;direction:rtl"> حين فضح وعزل التقييم القطري المشترك واللجنة الاقتصادية لأفريقيا ، وتجنب إلحاق أضرار العصب المبهم في مكان قريب والقصبة الهوائية ، التي يمكن أن تزيد من حجم واستمرارية انخفاض احتشاء. </li><li style=";text-align:right;direction:rtl"> أبدا إدراج خياطة حيدة أكثر من 10 ملم تمرير التشعب. ويمكن للخياطة إدراج بعيدا خرم الشريان الدماغي الأمامي ويؤدي إلى نزف في الدماغ. يمكن رأى المقاومة عندما متقدمة حول خياطة 90-10 ملم وراء النقطة التشعب. إذا حدث هذا ، يجب على الباحث وقف التقدم في خياطة الجروح وتأكيد المسافة. </li></ol>

<!DOCTYPE html PUBLIC "-//W3C//DTD XHTML 1.0 Transitional//EN" "http://www.w3.org/TR/xhtml1/DTD/xhtml1-transitional.dtd">
<html xmlns="http://www.w3.org/1999/xhtml" xml:lang="en" lang="en">	
		
		<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>Isoflurane</td>
<td>Chemical	</td>
<td>Aerrane, Baxter</td>
<td>95045-588</td>
<td>&nbsp;</td>
<tr>
<td>V-10 Anesthesia system</td>
<td>Equipment</td>
<td>VetEquip</td>
<td>901807</td>
<td>&nbsp;</td>
<tr>
<td>TR-200 Temp Controller</td>
<td>Equipment</td>
<td>Fine Science Tools</td>
<td>21060</td>
<td>&nbsp;</td>
<tr>
<td>Electric clipper</td>
<td>Equipment</td>
<td>Braintree</td>
<td>CLP-9931</td>
<td>&nbsp;</td>
<tr>
<td>Dissecting microscope</td>
<td>Equipment</td>
<td>Nikon</td>
<td>SMZ745T</td>
<td>&nbsp;</td>
<tr>
<td>Retractor system</td>
<td>Equipment</td>
<td>Braintree</td>
<td>ACD-014</td>
<td>&nbsp;</td>
<tr>
<td>Bipolar coagulator</td>
<td>Equipment</td>
<td>Howard Instrument</td>
<td>64000</td>
<td>&nbsp;</td>
<tr>
<td>Silk suture</td>
<td>Material</td>
<td>Harvard Apparatus</td>
<td>510479</td>
<td>&nbsp;</td>
<tr>
<td>Monofilament suture</td>
<td>Material</td>
<td>Harvard Apparatus</td>
<td>723351</td>
<td>&nbsp;</td>
<tr>
<td>Vascular clamp</td>
<td>Equipment</td>
<td>Fine Science Tools</td>
<td>00396-01</td>
<td>&nbsp;</td>
<tr>
<td>Vannas scissor</td>
<td>Equipment</td>
<td>Fine Science Tools</td>
<td>15000-08</td>
<td>&nbsp;</td>
<tr>
<td>Brain matrix</td>
<td>Equipment</td>
<td>Braintree</td>
<td>BS-2000C</td>
<td>&nbsp;</td>
<tr>
<td>2,3,5-triphenyltetrazolium chloride</td>
<td>Chemical</td>
<td>Sigma-Aldrich</td>
<td>T8877</td>
<td>&nbsp;</td>
<tr>
<td>10% neutral buffer formalin</td>
<td>Chemical</td>
<td>Sigma-Aldrich</td>
<td>HT5011</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

mouse model of middle cerebral artery occlusion

السكتة الدماغية هي الأكثر شيوعا مرض عصبي قاتل في الولايات المتحدة (1). الغالبية من السكتات الدماغية (88 ٪) نتيجة انسداد الأوعية الدموية في الدماغ (السكتة الدماغية) 2. منذ السكتات الدماغية أكثر (~ 80 ٪) تحدث في أراضي الشريان الدماغي الأوسط (MCA) 3 ، وركزت العديد من النماذج الحيوانية السكتة الدماغية التي تم وضعها على هذا الشريان. نموذج حيدة داخل اللمعة من انسداد الشريان الدماغي الأوسط (MCAO) ينطوي على ادراج خيوط جراحية في الشريان السباتي الخارجي والترابط إلى الأمام في الشريان السباتي الداخلي (ICA) حتى غيض occludes أصل MCA ، مما أدى إلى وقف تدفق الدم واحتشاء الدماغ لاحقة في 4 أراضي MCA. ويمكن استخدام هذه التقنية لنموذج إطباق دائمة أو عابرة 5. إذا تمت إزالة خياطة بعد فترة معينة (30 دقيقة ، ح 1 ، ح ​​أو 2) ، ويتحقق ضخه (MCAO عابرة) ، وإذا ترك خيوط في مكانه (24 ساعة) الإجراء هو مناسبة كنموذج للMCAO دائمة . هذه التقنية لا تتطلب قطع القحف ، وهو إجراء جراحة الأعصاب لإزالة جزء من الجمجمة ، والتي قد تؤثر على الضغط داخل الجمجمة ، ودرجة الحرارة 6. أصبح الأسلوب الأكثر استخداما لتقليد دائم ونقص تروية دماغية عابر التنسيق في الجرذان والفئران 7،8. لتقييم مدى احتشاء دماغي ، ونحن وصمة عار شرائح الدماغ مع 2،3،5 - triphenyltetrazolium كلوريد (TTC) لتحديد نقص تروية أنسجة المخ 9. في هذا الفيديو ، ونحن لشرح طريقة MCAO وتحديد حجم احتشاء بواسطة تلطيخ TTC.

Watch this Scientific Journal Video about Mouse Model of Middle Cerebral Artery Occlusion at JoVE.com

Mouse Model of Middle Cerebral Artery Occlusion

علينا أن نظهر في شريط الفيديو وسيلة لإنتاج وسط انسداد الشريان الدماغي في فئران بالغة باستخدام حيدة داخل اللمعة. علينا أن نبين كيفية تقييم مدى احتشاء الدماغ بواسطة 2،3،5 - triphenyltetrazolium تلطيخ (TTC) كلوريد.

نموذج الفأر من انسداد الشريان الدماغي الأوسط

Stroke is the most common fatal neurological disease in the United States 1. The majority of strokes (88%) result from blockage of blood vessels in the ...

mouse-model-of-middle-cerebral-artery-occlusion

Google

Research

JoVE Journal

Medicine

77.2K Views.  Ernest Gallo Clinic and Research Center, University of California, San Francisco. علينا أن نظهر في شريط الفيديو وسيلة لإنتاج وسط انسداد الشريان الدماغي في فئران بالغة باستخدام حيدة داخل اللمعة. علينا أن نبين كيفية تقييم مدى احتشاء الدماغ بواسطة 2،3،5 - triphenyltetrazolium تلطيخ (TTC) كلوريد.

Video: Mouse Model of Middle Cerebral Artery Occlusion

Modeling Stroke in Mice - Middle Cerebral Artery Occlusion with the Filament Model

Stroke is among the most frequent causes of death and adult disability, especially in highly developed countries. However, treatment options to date are very limited. To meet the need for novel therapeutic approaches, experimental stroke research frequently employs rodent models of focal cerebral ischaemia. Most researchers use permanent or transient occlusion of the middle cerebral artery (MCA) in mice or rats.
Proximal occlusion of the middle cerebral artery (MCA) via the intraluminal suture technique (so called filament or suture model) is probably the most frequently used model in experimental stroke research. The intraluminal MCAO model offers the advantage of inducing reproducible transient or permanent ischaemia of the MCA territory in a relatively non-invasive manner. Intraluminal approaches interrupt the blood flow of the entire territory of this artery. Filament occlusion thus arrests flow proximal to the lenticulo-striate arteries, which supply the basal ganglia. Filament occlusion of the MCA results in reproducible lesions in the cortex and striatum and can be either permanent or transient. In contrast, models inducing distal (to the branching of the lenticulo-striate arteries) MCA occlusion typically spare the striatum and primarily involve the neocortex. In addition these models do require craniectomy. In the model demonstrated in this article, a silicon coated filament is introduced into the common carotid artery and advanced along the internal carotid artery into the Circle of Willis, where it blocks the origin of the middle cerebral artery. In patients, occlusions of the middle cerebral artery are among the most common causes of ischaemic stroke. Since varying ischemic intervals can be chosen freely in this model depending on the time point of reperfusion, ischaemic lesions with varying degrees of severity can be produced. Reperfusion by removal of the occluding filament at least partially models the restoration of blood flow after spontaneous or therapeutic (tPA) lysis of a thromboembolic clot in humans.
In this video we will present the basic technique as well as the major pitfalls and confounders which may limit the predictive value of this model.

Filamentous occlusion of the Middle cerebral artery is a common model for studying ischemic stroke in mice.

السكتة الدماغية النمذجة في الفئران -- الشرق انسداد الشريان الدماغي مع نموذج الشعيرة

Stroke is among the most frequent causes of death and adult disability, especially in highly developed countries. However, treatment options to date are ...

Intraluminal Middle Cerebral Artery Occlusion (MCAO) Model for Ischemic Stroke with Laser Doppler Flowmetry Guidance in Mice

Stroke is the third leading cause of death and the leading cause of disability in the world, with an estimated cost of near $70 billion 
in the United States in 20091,2. The intraluminal middle cerebral artery occlusion (MCAO) model was developed by Koizumi4 in 1986 to simulate this 
impactful human pathology in the rat. A modification of the MCAO method was later presented by Longa3. Both techniques have been widely used to identify 
molecular mechanisms of brain injury resulting from ischemic stroke and potential therapeutic modalities5. This relatively noninvasive method in rats has been 
extended to use in mice to take advantage of transgenic and knockout strains6,7. To model focal cerebral ischemia, an intraluminal suture is advanced via 
the internal carotid artery to occlude the base of the MCA. Retracting the suture after a specified period of time mimics spontaneous reperfusion, but the 
suture can also be permanently retained. This video will be demonstrating the two major approaches for performing intraluminal MCAO procedure in mice in a 
stepwise fashion, as well as providing insights for potential drawbacks and pitfalls. The ischemic brain tissue will subsequently be stained by 
2,3,5-triphenyltetrazolium chloride (TTC) to evaluate the extent of cerebral infarction8.

Intraluminal Middle Cerebral Artery Occlusion MCAO Model for Ischemic Stroke with Laser Doppler Flowmetry Guidance in Mice

The intraluminal middle cerebral artery occlusion (MCAO) model is the most frequent used model among experimental ischemic stroke models. Here we will demonstrate the entire model in detail with the guide of Laser Doppler flowmetry, and its representative results.

الشريان الدماغي الأوسط انسداد داخل اللمعة (MCAO) النموذجي للاصابة بالسكتة الدماغية مع توجيه الليزر دوبلر Flowmetry في الفئران

Stroke is the third leading cause of death and the leading cause of disability in the world, with an estimated cost of near $70 billion 
in the United ...

The Application Of Permanent Middle Cerebral Artery Ligation in the Mouse

Focal cerebral ischemia is among the most common type of stroke seen in patients. Due to the clinical significance there has been a prolonged effort to develop suitable animal models to study the events that unfold during ischemic insult. These techniques include transient or permanent, focal or global ischemia models using many different animal models, with the most common being rodents.
The permanent MCA ligation method which is also referred as pMCAo in the literature is used extensively as a focal ischemia model in rodents 1-6. This method was originally described for rats by Tamura et al. in 1981 7. In this protocol a craniotomy was used to access the MCA and the proximal regions were occluded by electrocoagulation. The infarcts involve mostly cortical and sometimes striatal regions depending on the location of the occlusion. This technique is now well established and used in many laboratories 8-13. Early use of this technique led to the definition and description of &#x201C;infarct core&#x201D; and &#x201C;penumbra&#x201D; 14-16, and it is often used to evaluate potential neuroprotective compounds 10, 12, 13, 17. Although the initial studies were performed in rats, permanent MCA ligation has been used successfully in mice with slight modifications 18-20 .
This model yields reproducible infarcts and increased post-survival rates. Approximately 80% of the ischemic strokes in humans happen in the MCA area 21 and thus this is a very relevant model for stroke studies. Currently, there is a paucity of effective treatments available to stroke patients, and thus there is a need for good models to test potential pharmacological compounds and evaluate physiological outcomes. This method can also be used for studying intracellular hypoxia response mechanisms in vivo. 
Here, we present the MCA ligation surgery in a C57/BL6 mouse. We describe the pre-surgical preparation, MCA ligation surgery and 2,3,5 Triphenyltetrazolium chloride (TTC) staining for quantification of infarct volumes.

Middle cerebral artery (MCA) ligation is a technique to study focal cerebral ischemia in animal models. In this method, the middle cerebral artery is exposed by craniotomy and ligated by cauterization. This method gives highly reproducible infarct volumes and increased post-operative survival rates compared to other methods available.

تطبيق الربط الدائم الشريان المخي الأوسط في الماوس

Focal cerebral ischemia is among the most common type of stroke seen in patients. Due to the clinical significance there has been a prolonged effort to ...

Endothelin-1 Induced Middle Cerebral Artery Occlusion Model for Ischemic Stroke with Laser Doppler Flowmetry Guidance in Rat

Stroke is the number one cause of disability and third leading cause of death in the world, costing an estimated $70 billion in the United States in 20091, 2. Several models of cerebral ischemia have been developed to mimic the human condition of stroke. It has been suggested that up to 80% of all strokes result from ischemic damage in the middle cerebral artery (MCA) area3. In the early 1990s, endothelin-1 (ET-1) 4 was used to induce ischemia by applying it directly adjacent to the surface of the MCA after craniotomy. Later, this model was modified 5 by using a stereotaxic injection of ET-1 adjacent to the MCA to produce focal cerebral ischemia. The main advantages of this model include the ability to perform the procedure quickly, the ability to control artery constriction by altering the dose of ET-1 delivered, no need to manipulate the extracranial vessels supplying blood to the brain as well as gradual reperfusion rates that more closely mimics the reperfusion in humans5-7. On the other hand, the ET-1 model has disadvantages that include the need for a craniotomy, as well as higher variability in stroke volume8. This variability can be reduced with the use of laser Doppler flowmetry (LDF) to verify cerebral ischemia during ET-1 infusion. Factors that affect stroke variability include precision of infusion and the batch of the ET-1 used6. Another important consideration is that although reperfusion is a common occurrence in human stroke, the duration of occlusion for ET-1 induced MCAO may not closely mimic that of human stroke where many patients have partial reperfusion over a period of hours to days following occlusion9, 10. This protocol will describe in detail the ET-1 induced MCAO model for ischemic stroke in rats. It will also draw attention to special considerations and potential drawbacks throughout the procedure.

Several animal models of cerebral ischemia have been developed to simulate the human condition of stroke. This protocol describes the endothelin-1 (ET-1) induced middle cerebral artery occlusion (MCAO) model for ischemic stroke in rats. In addition, important considerations, advantages, and shortcomings of this model are discussed.

Endothelin-1 المستحثة انسداد الشريان الدماغي الأوسط نموذج للالسكتة الدماغية مع التوجيه بالليزر Flowmetry دوبلر في الجرذ

Stroke is the number one cause of disability and third leading cause of death in the world, costing an estimated $70 billion in the United States in ...

Utilizing a Cranial Window to Visualize the Middle Cerebral Artery During Endothelin-1 Induced Middle Cerebral Artery Occlusion

Creation of a cranial window is a method that allows direct visualization of structures on the cortical surface of the brain1-3. This technique can be performed in many locations overlying the rat cerebrum, but is most easily carried out by creating a craniectomy over the readily accessible frontal or parietal bones. Most frequently, we have used this technique in combination with the endothelin-1 middle cerebral artery occlusion model of ischemic stroke to quantify the changes in middle cerebral artery vessel diameter that occur with injection of endothelin-1 into the brain parenchyma adjacent to the proximal MCA4, 5. In order to visualize the proximal portion of the MCA during endothelin -1 induced MCAO, we use a technique to create a cranial window through the temporal bone on the lateral aspect of the rat skull (Figure 1). Cerebral arteries can be visualized either with the dura intact or with the dura incised and retracted. Most commonly, we leave the dura intact during visualization since endothelin-1 induced MCAO involves delivery of the vasoconstricting peptide into the brain parenchyma. This bypasses the need to incise the dura directly over the visualized vessels for drug delivery. This protocol will describe how to create a cranial window to visualize cerebral arteries in a step-wise fashion, as well as how to avoid many of the potential pitfalls pertaining to this method.

This article describes a method for visualizing rat cerebral arteries through a cranial window using temporal craniectomy in order to view proximal portions of the middle cerebral artery (Figure 1). This versatile method can be combined with various techniques of drug delivery to measure cerebral artery reactivity in vivo.

الاستفادة من وجود نافذة القحف لتصور الشريان الدماغي الأوسط خلال Endothelin-1 انسداد الشريان الدماغي الأوسط المستحثة

Creation of a cranial window is a method that allows direct visualization of structures on the cortical surface of the brain1-3. This technique can be ...

Optimized System for Cerebral Perfusion Monitoring in the Rat Stroke Model of Intraluminal Middle Cerebral Artery Occlusion

The translational potential of pre-clinical stroke research depends on the accuracy of experimental modeling. Cerebral perfusion monitoring in animal models of acute ischemic stroke allows to confirm successful arterial occlusion and exclude subarachnoid hemorrhage. Cerebral perfusion monitoring can also be used to study intracranial collateral circulation, which is emerging as a powerful determinant of stroke outcome and a possible therapeutic target. Despite a recognized role of Laser Doppler perfusion monitoring as part of the current guidelines for experimental cerebral ischemia, a number of technical difficulties exist that limit its widespread use. One of the major issues is obtaining a secure and prolonged attachment of a deep-penetration Laser Doppler probe to the animal skull. In this video, we show our optimized system for cerebral perfusion monitoring during transient middle cerebral artery occlusion by intraluminal filament in the rat. We developed in-house a simple method to obtain a custom made holder for twin-fibre (deep-penetration) Laser Doppler probes, which allow multi-site monitoring if needed. A continuous and prolonged monitoring of cerebral perfusion could easily be obtained over the intact skull.

Cerebral perfusion monitoring has been demonstrated to improve accuracy in ischemic stroke models. Technical difficulties often limit the use of this essential tool for cerebrovascular research. In this video, an optimized system is shown to obtain a single or multi-site hemodynamic monitoring during intraluminal middle cerebral artery occlusion in rats.

الأمثل نظام لرصد الإرواء الدماغي في نموذج الفئران من السكتة الدماغية داخل اللمعة انسداد الشريان الدماغي الأوسط

The translational potential of pre-clinical stroke research depends on the accuracy of experimental modeling. Cerebral perfusion monitoring in animal ...

Embolic Middle Cerebral Artery Occlusion (MCAO) for Ischemic Stroke with Homologous Blood Clots in Rats

Clinically, thrombolytic therapy with use of recombinant tissue plasminogen activator (tPA) remains the most effective treatment for acute ischemic stroke. However, the use of tPA is limited by its narrow therapeutic window and by increased risk of hemorrhagic transformation. There is an urgent need to develop suitable stroke models to study new thrombolytic agents and strategies for treatment of ischemic stroke. At present, two major types of ischemic stroke models have been developed in rats and mice: intraluminal suture MCAO and embolic MCAO. Although MCAO models via the intraluminal suture technique have been widely used in mechanism-driven stroke research, these suture models do not mimic the clinical situation and are not suitable for thrombolytic studies. Among these models, the embolic MCAO model closely mimics human ischemic stroke and is suitable for preclinical investigation of thrombolytic therapy. This embolic model was first developed in rats by Overgaard et al.1 in 1992 and further characterized by Zhang et al. in 19972. Although embolic MCAO has gained increasing attention, there are technical problems faced by many laboratories. To meet increasing needs for thrombolytic research, we present a highly reproducible model of embolic MCAO in the rat, which can develop a predictable infarct volume within the MCA territory. In brief, a modified PE-50 tube is gently advanced from the external carotid artery (ECA) into the lumen of the internal carotid artery (ICA) until the tip of the catheter reaches the origin of the MCA. Through the catheter, a single homologous blood clot is placed at the origin of the MCA. To identify the success of MCA occlusion, regional cerebral blood flow was monitored, neurological deficits and infarct volumes were measured. The techniques presented in this paper should help investigators to overcome technical problems for establishing this model for stroke research.

Embolic Middle Cerebral Artery Occlusion MCAO for Ischemic Stroke with Homologous Blood Clots in Rats

In this paper, we demonstrate a standard method for producing an embolic middle cerebral artery occlusion with homologous blood clots (fibrin-rich) in adult rat. This model closely mimics human ischemic stroke and is suitable for preclinical study of thrombolytic therapy for ischemic stroke.

صمي الأوسط انسداد الشريان الدماغي (MCAO) لالدماغية السكتة الدماغية مع مثلي تجلط الدم في الجرذان

Clinically, thrombolytic therapy with use of recombinant tissue plasminogen activator (tPA) remains the most effective treatment for acute ischemic ...

Quantification of Neurovascular Protection Following Repetitive Hypoxic Preconditioning and Transient Middle Cerebral Artery Occlusion in Mice

Experimental animal models of stroke are invaluable tools for understanding stroke pathology and developing more effective treatment strategies. A 2 week protocol for repetitive hypoxic preconditioning (RHP) induces long-term protection against central nervous system (CNS) injury in a mouse model of focal ischemic stroke. RHP consists of 9 stochastic exposures to hypoxia that vary in both duration (2 or 4 hr) and intensity (8% and 11% O2). RHP reduces infarct volumes, blood-brain barrier (BBB) disruption, and the post-stroke inflammatory response for weeks following the last exposure to hypoxia, suggesting a long-term induction of an endogenous CNS-protective phenotype. The methodology for the dual quantification of infarct volume and BBB disruption is effective in assessing neurovascular protection in mice with RHP or other putative neuroprotectants. Adult male Swiss Webster mice were preconditioned by RHP or duration-equivalent exposures to 21% O2 (i.e. room air). A 60 min transient middle cerebral artery occlusion (tMCAo) was induced 2 weeks following the last hypoxic exposure. Both the occlusion and reperfusion were confirmed by transcranial laser Doppler flowmetry. Twenty-two hr after reperfusion, Evans Blue (EB) was intravenously administered through a tail vein injection. 2 hr later, animals were sacrificed by isoflurane overdose and brain sections were stained with 2,3,5- triphenyltetrazolium chloride (TTC). Infarcts volumes were then quantified. Next, EB was extracted from the tissue over 48 hr to determine BBB disruption after tMCAo. In summary, RHP is a simple protocol that can be replicated, with minimal cost, to induce long-term endogenous neurovascular protection from stroke injury in mice, with the translational potential for other CNS-based and systemic pro-inflammatory disease states.

This protocol describes repetitive hypoxic preconditioning, or brief exposures to systemic hypoxia that reduce infarct volumes and blood-brain barrier disruption following transient middle cerebral artery occlusion in mice. It also details dual quantification of infarct volume and blood-brain barrier disruption after stroke to assess the efficacy of neurovascular protection.

الكمي لحماية عائية عصبية بعد المتكرر نقص الأوكسجين شروط مسبقة وعابر الأوسط الشريان الدماغي انسداد في الفئران

Experimental animal models of stroke are invaluable tools for understanding stroke pathology and developing more effective treatment strategies. A 2 week ...

Surgical Approach for Middle Cerebral Artery Occlusion and Reperfusion Induced Stroke in Mice

Stroke is a leading cause of death worldwide and continues to be one of the major causes of long-term adult disabilities. About 87% of strokes are ischemic in origin and occur in the territory of the middle cerebral artery (MCA). Currently the only Food and Drug Administration (FDA) approved drug for the treatment of this devastating disease is tissue plasminogen activator (tPA). However, tPA has a small therapeutic window for administration (3 - 6 hr), and is only effective in 4% of the patients who actually receive it. Current research focuses on understanding the pathophysiology of stroke in order to find potential therapeutic targets. Thus, reliable models are crucial, and the MCA occlusion (MCAo) model (also termed the intraluminal filament or suture model) is deemed to be the most clinically relevant surgical model of ischemic stroke, and is fairly non-invasive and easily reproducible. Typically the MCAo model is used with rodents, especially with mice due to all the genetic variations available for this species. Here we describe (and present in the video) how to successfully perform the MCAo model (with reperfusion) in mice to generate reliable and reproducible data.

In order to understand the pathophysiology of stroke, it is important to use reliable models. This paper will describe one of the most frequently used stroke models in mice, termed the middle cerebral artery occlusion (MCAo) model (also termed the intraluminal filament or suture model) with reperfusion.

نهج الجراحي لانسداد الشريان الدماغي الأوسط وضخه المستحثة السكتة الدماغية في الفئران

Stroke is a leading cause of death worldwide and continues to be one of the major causes of long-term adult disabilities. About 87% of strokes are ...

Transient Middle Cerebral Artery Occlusion Model of Neonatal Stroke in P10 Rats

A number of animal models have been used to study hypoxic-ischemic injury, traumatic injury, global hypoxia, or permanent ischemia in both the immature and mature brain. Stroke occurs commonly in the perinatal period in humans, and transient ischemia-reperfusion is the most common form of stroke in neonates. The reperfusion phase is a critical component of injury progression, which occurs over a period of days to weeks, and of the endogenous response to injury. This postnatal day 10 (p10) rat model of transient middle cerebral artery occlusion (tMCAO) creates a unilateral, non-hemorrhagic focal ischemia-reperfusion injury that can be utilized to study the mechanisms of focal injury and repair in the full-term-equivalent brain. The injury pattern that is produced by tMCAO is consistent and highly reproducible and can be confirmed with MRI or histological analyses. The severity of injury can be manipulated through changes in occlusion time and other methods that will be discussed.

Neonatal stroke is a significant cause of early brain injury requiring a translational model with consistent focal injury patterns and high reproducibility in order to enable study. This study describes the detailed surgical procedure for creating a non-hemorrhagic, unilateral focal ischemia-reperfusion injury in full-term-equivalent rodents.

عابر الدماغي الأوسط الشريان انسداد نموذج من السكتة الدماغية حديثي الولادة في الفئران P10

A number of animal models have been used to study hypoxic-ischemic injury, traumatic injury, global hypoxia, or permanent ischemia in both the immature ...