Bu çalışma Amerikan Kalp Derneği Büyükşehir Güneydoğu Ortaklık (09GRNT2060421), Amerikan Tıp Derneği, ve Florida Klinik ve Translational Bilim Enstitüsü Üniversitesi bağışları ile desteklenmiştir. Adam Mekke bir NIH / NINDS, NRSA predoctoral fellow (F30 NS-060335) &#39;dir. Robert Regenhardt Hipertansiyon Florida Multidisipliner Eğitim Programı Üniversitesi (T32 HL-083810) den predoctoral burs desteğini aldı.

Bu protokol Florida Üniversitesi Kurumsal Hayvan Bakım ve Kullanım Kurulu (IACUC) tarafından onaylanmış ve uygun olduğunu &quot;Bakım Rehberi ve Laboratuvar Hayvanlarının Kullanımı&quot; (sekizinci baskı, Ulusal Bilimler Akademisi, 2011) oldu. Malzemeler <ol><li> Hayvanlar: Ameliyat sırasında 250-300 g Sekiz haftalık, erkek, Sprague Dawley sıçan (Charles River Çiftlikler, Wilmington, ABD MA). </li><li> Anestezi <ol class="lalpha"><li> İnhalasyon anestezi sistemi (VetEquip A.Ş., Pleasanton, CA, USA) </li><li> Izofluran anestezi (Baxter İlaç sanayii, Deerfield, IL, ABD) </li></ol></li><li> Stereotaksik sistemi (David Kopf Instruments, Tujunga, CA, USA) <ol class="lalpha"><li> Küçük hayvan stereotaksik sistemi </li><li> Sıçanlarda Non-rüptürü kulak çubukları </li><li> Sıçanlar için gaz anestezi kafa tutucu </li></ol></li><li> Sıcaklık ayarı<ol class="lalpha"><li> BAT-12 mikroprob termometre (Dünya Precision Instruments, Inc, Sarasota, FL, ABD) </li><li> T / POMPA, TP600 Termal battaniye (Gaymar Industries, Inc, Orchard Park, NY, ABD) </li></ol></li><li> Cerrahi aletler <ol class="lalpha"><li> Bisturi sapı ve 11. bıçak, iris forseps, Graefe forseps, tornavida bulldog klemp Retraktörler, 26 ayar eğimli iğne (Dünya Precision Instruments, Inc, Sarasota, FL, USA) 10 ul şırınga </li><li> Mikromotor matkap ve stereotaksik tutucu, Quintessential Stereotaksik Enjektör (Stoelting, Wood Dale, IL, ABD) </li><li> 1,0 mm yuvarlak matkap bur, 1,0 mm ters koni matkap frez (Roboz Cerrahi Instrument Co, Inc, Gaithersburg, MD, USA) </li></ol></li><li> Cerrahi Malzemeleri <ol class="lalpha"><li> 2.4 mm şaft uzunluğu, 21-gauge kılavuz kanül [uzun kaide altında 4mm] ve kanül kukla (Plastik biri Roanoke vidalar 0-80 X 3/32 Montaj, VA, USA) </li><li> Jet protez akrilik ve sıvı (Lang Diş Manufacturing Co, Inc, Wheeling, IL, ABD) </li><li> 3,0 naylon sütür (Oasis, Mettawa, IL, ABD) </li><li> Pamuklu çubuklar, Puralube göz merhemi (Fisher Scientific, Pittsburg, PA, USA) </li><li> Elektrikli saç kesme makineleri (Oster, Providence, RI, ABD) </li></ol></li><li> Kimyasallar <ol class="lalpha"><li> Endotelin-1 (Amerikan Peptid, Sunnyvale, CA, USA) </li><li> Klorheksidin% 2 (Agrilabs, Joseph, MO, ABD) </li><li> Buprenorfin HCl (Hospira Inc, Lake Forest, IL, ABD) </li></ol></li><li> Görselleştirme Ekipmanları <ol class="lalpha"><li> Cerrahi mikroskop (Seiler Instrument ve Üretim; St Louis, MO, USA) </li><li> Fiber Optik aydınlatıcı (TechniQuip Corp, Livermore, CA, USA) </li></ol></li><li> Lazer Doppler flowmetre sistemi (ADInstruments, Inc, Colorado Springs, CO, ABD) <ol class="lalpha"><li> Standard Kalem Probu </li><li> Prob tutucu </li><li> Kan Debimetre </li><li> LabChart 7 ile PowerLab 4/30 </li></ol></li><li> Infarkt hacminin ölçümü <ol class="lalpha"><li> Sıçan beyin matriks (Zivic-Miller Lab., Inc, Allison Park, PA, USA) </li><li> 2,3,5-Triphenyftetrazoliumchlorid (Sigma-Aldrich Co, St Louis, MO, ABD), PBS içinde% 0.05 ile seyreltildi </li><li> Flatbed tarayıcı (Epson Perfection V30, Epson America, Inc, Long Beach, CA, USA) </li><li> Image J yazılımı (ImageJ 1.42q yazılım, ABD Ulusal Sağlık Enstitüleri, Bethesda, MA, ABD) </li></ol></li></ol> 1. Ön cerrahi Adımlar <ol><li> Ameliyat öncesinde, sıçanlar su ve kemirgen chow ücretsiz erişim ile 00:12 aydınlık / karanlık döngüsü altında yer alır. </li><li> Anestezi arka pençe çimdik çekme artık sıçan kadar bir indüksiyon odasında% 100 O 2 gaz karışımı içinde% 4 izofluran ile indüklenir. </li><li> ASeptik tekniği steril eldiven, steril cerrahi aletler, ve steril cerrahi örtü 11 kullanımı da dahil olmak üzere bu işlem sırasında devam etmelidir. </li><li> Baş tacı elektrikli saç makasları ile traş edilir. </li><li> Sıçan sıcaklık kontrollü bir çalışma yüzeyi (termal battaniye) üzerinde yatan bir emici ped üzerine yüzüstü pozisyonda yerleştirilir. </li><li> Kafa gaz anestetik yüz maskesi yerleştirilmesi ile başlayarak stereotaksik alet içine yerleştirilir. </li><li> Sonraki, kulak çubukları yerleştirilir ve sıkılır. </li><li> Prosedür anestezi sırasında% 100 O 2 gaz karışımı içinde izofluran% 2 ile korunur. </li><li> Yağlayıcı oftalmik merhem hem gözlere uygulanır ve göz kapaklarının cerrahi prosedür sırasında göz kuruma önlemek için kapatılır. </li><li> Rektal sıcaklık probu ± 0.5 ° C sıcaklıkta 37 sabit bir hayvansal çekirdek sıcaklığı korumak için eklenir </li><li> Stereotaksik sıkıca tutulan baş ilecihaz, cerrahi alanda% 2 klorheksidin ve tuzlu üç kez dönüşümlü olarak temizlenir. </li></ol> 2. Cerrahi Adımlar <ol><li> Bir neşter kullanılarak, orta hat kesi kulak (üstün ense hattı) arasına gözlerin en kaudal yönleri (nasion) den kalvaryumun örten cilt üzerinde yapılır. </li><li> Deri sonra 3 bulldog kelepçeler ile yanal geri çekilir. </li><li> Bağ dokusu çoklu yapılar açıkça görülebilir, böylece kuru pamuklu kullanarak kafatası kaldırılır. Bunlar bregma, koronal sütür ve sağ lateral kafa sırtı bulunur. Pamuklu bezler cerrahi alanında kan çıkarmak için kullanılır. </li><li> Ameliyat mikroskobu kullanarak, bregma bulunur, ve 1.0 mm boyunca matkap bur bregma de sıfır olana kadar stereotaksik manipulators ayarlanır. </li><li> Matkap frez ardından 1.6 mm ön ve bregma lateral 5,2 mm taşınır. </li><li> Kafatası nüfuz A frez deliği Cann için delinirula yerleştirme (Şekil 1). Fazla enkaz ve kan sürekli pamuklu kullanılarak temizlenir. </li></ol> Bu noktada, bir kılavuz kanül sokulabilir (adım 7) ya da bur delik aracılığıyla bir direkt ET-1, enjeksiyon (16 doğrudan adım devam) gerçekleştirilebilir. <ol start="7"><li> 3 montaj vidaları için Sonraki, bur delikleri 1,0 mm ters koni matkap frez (Şekil 1) kullanarak kafatası parsiyel kalınlıkta ile delinir. Bir delik koronal sütür ve sutur lateral 1-2 mm anterior 1-2 mm her frontal kemikte delinir. Bir delik koronal sütür ve kılavuz kanül frez deliği ipsilateral sutur lateral 2-3 mm posterior 2-3 mm kadar paryetal kemikte delinir. Üç 0-80 x 3/32 montaj vidaları bu bur deliklere yerleştirilir ve kanül tutan çimento için destek sağlayacaktır. Vidalar sadece dura matere zarar vermemeye olarak 2 veya 3 tur kadar gelişmiş olmalıdır. </li&gt; <li> Kılavuz kanül stereotaksik kanül tutucu yerleştirilir ve bregma yer almaktadır. </li><li> Kılavuz kanül bregma de sıfır olana kadar stereotaksik manipülatörleri ayarlanır. Kanül 1.6 mm ön ve bregma lateral 5,2 mm uzaklıkta frez deliği taşınır. </li><li> Son olarak, kılavuz kanül bregma ventral 4.5 mm nihai uç pozisyon ile bur delik içine indirilir. </li><li> Lazer Doppler probu yerleştirilmesi (isteğe bağlı) <ol class="lalpha"><li> LDF kullanarak serebral kan akımını izlemek için, bir prob tutucu diş çimento ile kılavuz kanül takılma öncesinde pozisyon içine yerleştirilebilir. </li><li> Prob tutucu tabanı küçük kama şeklinde sekmesi dışında kaide ile aynı hizada kesilir. </li><li> Prob tutucu sonra kılavuz kanül posterior ve medial odaklı sekmesini (Şekil 1) ile lateral kafa sırt sadece medial yerleştirilir. </li><li> Prob tutucu ve guide kanül diş çimento kullanılarak birbirine yapıştırılmıştır. </li></ol></li><li> Diş çimento sonra yerinde kanül sabitlemek için kullanılır. Çimento her üç vida ile temas halinde olan ve kanül tüm taban çevreler. </li><li> Çimento önce kanül tutucu kaldırılması için tamamen kuru olmalıdır. Bu yaklaşık 5 dakika sürer. </li></ol> Bu adımlardan sonra, cerrahi kesi kapalı olabilir ve kanül kukla kılavuz kanül içine ve vidalı olabilir. Alternatif olarak, ET-1 indüklenmiş MCAO kanül implant ameliyat sonrası iyileşme süresinden sonra sıçan üzerinde gerçekleştirilebilir. Bu yöntem için, 19 ileri yapılmalı ve adımlar 14-18 daha sonraki bir zamanda gerçekleştirilebilir. Aynı ameliyat sırasında kılavuz kanül implant ve ET-1 enjeksiyon işlemini yapmak için bir sonraki adım 14 yapılmalıdır. <ol start="14"><li> Infüzyon şırınga ET-1 ile yüklenir (PBS içinde 80 uM ile seyreltilmiş), ve daha sonra, stereotaksik enjektörü monte edilir. </li><li>İğne ucu kılavuz kanül tarafındaki ucunda sıfır olana kadar stereotaksik manipülatörleri ayarlanır. </li><li> İğne ucu kılavuz kenar kanül ventral 17.2 mm arasında bir konuma kılavuz kanül vasıtasıyla azaltılır. Bir kılavuz kanül kullanılmış ise, iğne ucu bregma de sıfırlanır ve bregma ventral bir pozisyon 8,7 mm bur delik boyunca aşağı iner. </li><li> 80 uM ET1 3 ul dakika başına 1 ul bir oranda demlenir. </li><li> Infüzyon yavaş kaldırılır sonra eksiksiz ve sonra şırınga 3 dakika boyunca yerinde kalır. </li><li> Kesi 3.0 naylon dikiş ve kanül kukla kanül içine vidalanır ile kapatılır. </li><li> Uygun analjezik dozu (0.05-0.1 mg / kg yani buprenorfin) iyileşme döneminde ağrı ve rahatsızlık en aza indirmek için ameliyat sonrası kullanılmalıdır. </li><li> Sıçan cerrahi seti kaldırılır ve yumuşak gıda ve su ücretsiz, kolay erişim, hipotermi önlemek için kuru ve sıcak bir kurtarma alanına yerleştirilir. </li&gt; </ol>

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Çıkar çatışması ilan etti.

ET-1 ile indüklenen MCAO düzenli birden sıçan suşları kullanılmıştır deneysel iskemik inme kurulmuş bir modeldir. Bu model 5, 14 kullanırken böyle sıçan suşu, hayvan yaşı, vücut ısısı, anestezi yöntemi ve operatör uzmanlığı gibi birçok değişken, infarkt hacimleri artan değişkenliği yol açabilir. Ancak, bazı araştırmacılar bu modelin avantajları nispeten non-invaziv yaklaşım, ET-1 ile serebral kan akımı doz yanıt, ve bilinçli sıçanlar 14, 15 ET-1 infü...

<table border="1"><tbody><tr><td></td><td></td><td></td><td><ol><li> Hayvanlar: Ameliyat sırasında 250-300 g Sekiz haftalık, erkek, Sprague Dawley sıçan (Charles River Çiftlikler, Wilmington, ABD MA). </li><li> Anestezi <ol class="lalpha"><li> İnhalasyon anestezi sistemi (VetEquip A.Ş., Pleasanton, CA, USA) </li><li> Izofluran anestezi (Baxter İlaç sanayii, Deerfield, IL, ABD) </li></ol></li><li> Stereotaksik sistemi (David Kopf Instruments, Tujunga, CA, USA) <ol class="lalpha"><li> Küçük hayvan stereotaksik sistemi </li><li> Sıçanlarda Non-rüptürü kulak çubukları </li><li> Sıçanlar için gaz anestezi kafa tutucu </li></ol></li><li> Sıcaklık ayarı <ol class="lalpha"><li> BAT-12 mikroprob termometre (Dünya Precision Instruments, Inc, Sarasota, FL, ABD) </li><li> T / POMPA, TP600 Termal battaniye (Gaymar Industries, Inc, Orchard Park, NY, ABD) </li></ol></li><li> Cerrahi aletler <ol class="lalpha"><li> Bisturi sapı ve 11. bıçak, iris forseps, Graefe forseps, tornavida bulldog klemp Retraktörler, 26 ayar eğimli iğne (Dünya Precision Instruments, Inc, Sarasota, FL, USA) 10 ul şırınga </li><li> Mikromotor matkap ve stereotaksik tutucu, Quintessential Stereotaksik Enjektör (Stoelting, Wood Dale, IL, ABD) </li><li> 1,0 mm yuvarlak matkap bur, 1,0 mm ters koni matkap frez (Roboz Cerrahi Instrument Co, Inc, Gaithersburg, MD, USA) </li></ol></li><li> Cerrahi Malzemeleri <ol class="lalpha"><li> 2.4 mm şaft uzunluğu, 21-gauge kılavuz kanül [uzun kaide altında 4mm] ve kanül kukla (Plastik biri, Roanoke, VA, USA) ile vidaları 0-80 X 3/32 Montaj </li><li> Jet protez akrilik ve sıvı (Lang Diş Manufacturing Co, Inc, Wheeling, IL, ABD) </li><li> 3,0 naylon sütür (Oasis, Mettawa, IL, ABD) </li><li> Pamuklu çubuklar, Puralube göz merhemi (Fisher Scientific, Pittsburg, PA, ABD) </li><li> Elektrikli saç kesme makineleri (Oster, Providence, RI, ABD) </li></ol></li><li> Kimyasallar <ol class="lalpha"><li> Endotelin-1 (Amerikan Peptid, Sunnyvale, CA, USA) </li><li> Klorheksidin% 2 (Agrilabs, Joseph, MO, ABD) </li><li> Buprenorfin HCl (Hospira Inc, Lake Forest, IL, ABD) </li></ol></li><li> Görselleştirme Ekipmanları <ol class="lalpha"><li> Cerrahi mikroskop (Seiler Instrument ve Üretim; St Louis, MO, USA) </li><li> Fiber Optik aydınlatıcı (TechniQuip Corp, Livermore, CA, USA) </li></ol></li><li> Lazer Doppler flowmetre sistemi (ADInstruments, Inc, Colorado Springs, CO, ABD) <ol class="lalpha"><li> Standart Kalem Probu </li><li> Prob tutucu </li><li> Kan Debimetre </li><li> LabChart 7 ile PowerLab 4/30 </li></ol></li><li> Infarkt hacminin ölçümü <ol class="lalpha"><li> Sıçan beyinmatriks (Zivic-Miller Lab., Inc, Allison Park, PA, USA) </li><li> 2,3,5-Triphenyftetrazoliumchlorid (Sigma-Aldrich Co, St Louis, MO, ABD), PBS içinde% 0.05 ile seyreltildi </li><li> Flatbed tarayıcı (Epson Perfection V30, Epson America, Inc, Long Beach, CA, USA) </li><li> Image J yazılımı (ImageJ 1.42q yazılım, ABD Ulusal Sağlık Enstitüleri, Bethesda, MA, ABD) </li></ol></li></ol></td></tr></tbody></table>

endothelin-1 induced middle cerebral artery occlusion model for ischemic stroke with laser doppler flowmetry guidance in rat

İnme 2009 1, 2 ABD&#39;de tahmini 70000000000 $ maliyetlendirme, sakatlık ve dünyada ölüm üçüncü önde gelen nedenidir bir numaralı nedenidir. Serebral iskemi çeşitli modeller inme insanlık durumu taklit etmek için geliştirilmiştir. Bu orta serebral arter (OSA) alanında 3 iskemik hasardan tüm vuruşları sonucu% 80 o kadar ileri sürülmüştür. 1990&#39;ların başlarında, endotelin-1 (ET-1) 4 kranyotomi sonra MCA yüzeyine doğrudan doğruya bitişik uygulayarak ve iskemi indüklemek için kullanılmıştır. Daha sonra, bu model fokal serebral iskemi üretmek için MCA ET-1, bitişik bir stereotaksik enjeksiyon ile 5 modifiye edilmiştir. Bu modelin ana avantajları hızlı yordamı gerçekleştirmek için yeteneği, ET-1 dozunda değiştirerek arter daralmasına kontrol etme yeteneği teslim, beyne kan tedarik yanı sıra kademeli reperfusi ekstrakraniyal damarların işlemek gerek vardıroranları o daha yakından 5-7 insanlarda reperfüzyon taklit eder. Diğer taraftan, ET-1 model bir kranyotomi için ihtiyaç yanı sıra, inme hacim 8 daha yüksek değişkenlik aşağıdaki dezavantajları vardır. Bu değişkenlik ET-1 infüzyonu sırasında serebral iskemi doğrulamak için lazer Doppler flowmetre (LDF) kullanımı ile azaltılabilir. Strok değişkenliği etkileyen faktörler infüzyon ve ET-1 toplu hassas 6. kullanılan içerir. Başka bir önemli husus da reperfüzyon insan felç sık karşılaşılan bir durumdur ki, ET-1 indüklenmiş MCAO için tıkanma süresinin yakından, insan felç birçok hasta tıkanma 9 gün sonraki saatlik bir süre boyunca kısmi reperfüzyon kalmamıştır taklit olmayabilir 10. Bu protokol ayrıntılı sıçanlarda iskemik inme için ET-1 ile indüklenen MCAO modeli anlatacağız. Aynı zamanda işlem boyunca özel hususlar ve potansiyel sakıncaları dikkat çekecek.

1. Post-Op nörolojik değerlendirme Hayvanlarda bilinç kavuşur sonra testler çeşitli denge, kavrama kuvveti, pençe yerleştirerek, postural asimetri ve tırmanma merdiven içeren nörolojik defisitler değerlendirmek için kullanılabilir. Ayçiçeği tohumu görevi motor ve infarkt hacmi 7, 12 arasında anlamlı bir ilişki vardır duyusal fonksiyon brüt değerlendirmesidir. Açılması ve 5 ayçiçeği tohumu tüketen bu görevi sırasında, fareler zamanlanmış. Beş tohum...

Watch this Scientific Journal Video about Endothelin-1 Induced Middle Cerebral Artery Occlusion Model for Ischemic Stroke with Laser Doppler Flowmetry Guidance in Rat at JoVE.com

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

Serebral iskemi Çeşitli hayvan modellerinde inme insanlık durumu simüle etmek için geliştirilmiştir. Bu protokol sıçanlarda iskemik inme için endotelin-1 (ET-1) bağlı orta serebral arter tıkanıklığı (MCAO) modeli anlatılmaktadır. Buna ek olarak, önemli hususlar, avantajlar, ve bu modelin eksiklikler ele alınmaktadır.

Rat Lazer Doppler Flowmetri Rehberlik ile İskemik İnme için Endotelin-1 Bağlı Orta Serebral Arter Tıkanıklığı Modeli

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 ...

endothelin-1-induced-middle-cerebral-artery-occlusion-model-for

Research

JoVE Journal

Medicine

19.8K Görüntüleme.  University of Florida. Serebral iskemi Çeşitli hayvan modellerinde inme insanlık durumu simüle etmek için geliştirilmiştir. Bu protokol sıçanlarda iskemik inme için endotelin-1 (ET-1) bağlı orta serebral arter tıkanıklığı (MCAO) modeli anlatılmaktadır. Buna ek olarak, önemli hususlar, avantajlar, ve bu modelin eksiklikler ele alınmaktadır.

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

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.

Fare Modelleme İnme - Filament Model Orta Serebral Arter Tıkanıklığı

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

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Mouse Model of Middle Cerebral Artery Occlusion

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 brain (ischemic stroke) 2. Since most ischemic strokes (~80%) occur in the territory of middle cerebral artery (MCA) 3, many animal stroke models that have been developed have focused on this artery. The intraluminal monofilament model of middle cerebral artery occlusion (MCAO) involves the insertion of a surgical filament into the external carotid artery and threading it forward into the internal carotid artery (ICA) until the tip occludes the origin of the MCA, resulting in a cessation of blood flow and subsequent brain infarction in the MCA territory 4. The technique can be used to model permanent or transient occlusion 5. If the suture is removed after a certain interval (30 min, 1 h, or 2 h), reperfusion is achieved (transient MCAO); if the filament is left in place (24 h) the procedure is suitable as a model of permanent MCAO. This technique does not require craniectomy, a neurosurgical procedure to remove a portion of skull, which may affect intracranial pressure and temperature 6. It has become the most frequently used method to mimic permanent and transient focal cerebral ischemia in rats and mice 7,8. To evaluate the extent of cerebral infarction, we stain brain slices with 2,3,5-triphenyltetrazolium chloride (TTC) to identify ischemic brain tissue 9. In this video, we demonstrate the MCAO method and the determination of infarct size by TTC staining.

We demonstrate in the video a method for producing a middle cerebral artery occlusion in adult mice using an intraluminal monofilament. We also show how to evaluate the extent of cerebral infarction by 2,3,5-triphenyltetrazolium chloride (TTC) staining.

Orta Serebral Arter Tıkanıklığı Fare Modeli

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 ...

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.

Fare lazer Doppler flowmetre Rehberlik İskemik İnme intraluminal Orta Serebral Arter Tıkanıklığı (MCAO) Model

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 ...

Bilateral Common Carotid Artery Occlusion as an Adequate Preconditioning Stimulus to Induce Early Ischemic Tolerance to Focal Cerebral Ischemia

There is accumulating evidence, that ischemic preconditioning - a non-damaging ischemic challenge to the brain - confers a transient protection to a subsequent damaging ischemic insult. We have established bilateral common carotid artery occlusion as a preconditioning stimulus to induce early ischemic tolerance to transient focal cerebral ischemia in C57Bl6/J mice. In this video, we will demonstrate the methodology used for this study.

There is accumulating evidence, that ischemic preconditioning (PC) &#x2013; a non-damaging ischemic challenge to the brain - confers a transient protection to a subsequent damaging ischemic insult. We established bilateral common carotid artery occlusion (BCCAO) as a preconditioning stimulus to induce early ischemic tolerance (IT) to transient focal cerebral ischemia (induced by middle cerebral artery occlusion, MCAO) in C57Bl6/J mice.

Odak Serebral İskemi Erken İskemik Tolerans tetikleyebilecek bir Yeterli Önkoşullama Uyarıcı olarak bilateral Ortak Karotis Arter Tıkanıklığı

There is accumulating evidence, that ischemic preconditioning - a non-damaging ischemic challenge to the brain - confers a transient protection to a ...

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.

Endotelin-1 Bağlı Orta Serebral Arter Tıkanıklığı sırasında Orta Serebral Arter görselleştirin bir Kranial Pencere kullanmak

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.

Intraluminar Orta Serebral Arter Tıkanıklığı ve Sıçan Strok Model Serebral Perfüzyon İzleme için Optimize Sistemi

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

Modeling Stroke in Mice: Permanent Coagulation of the Distal Middle Cerebral Artery

Stroke is the third most common cause of death and a main cause of acquired adult disability in developed countries. Only very limited therapeutical options are available for a small proportion of stroke patients in the acute phase. Current research is intensively searching for novel therapeutic strategies and is increasingly focusing on the sub-acute and chronic phase after stroke because more patients might be eligible for therapeutic interventions in a prolonged time window. These delayed mechanisms include important pathophysiological pathways such as post-stroke inflammation, angiogenesis, neuronal plasticity and regeneration. In order to analyze these mechanisms and to subsequently evaluate novel drug targets, experimental stroke models with clinical relevance, low mortality and high reproducibility are sought after. Moreover, mice are the smallest mammals in which a focal stroke lesion can be induced and for which a broad spectrum of transgenic models are available. Therefore, we describe here the mouse model of transcranial, permanent coagulation of the middle cerebral artery via electrocoagulation distal of the lenticulostriatal arteries, the so-called &#8220;coagulation model&#8221;. The resulting infarct in this model is located mainly in the cortex; the relative infarct volume in relation to brain size corresponds to the majority of human strokes. Moreover, the model fulfills the above-mentioned criteria of reproducibility and low mortality. In this video we demonstrate the surgical methods of stroke induction in the &#8220;coagulation model&#8221; and report histological and functional analysis tools.

Various murine models of middle cerebral artery occlusion (MCAO) are widely used in experimental brain research. Here, we demonstrate the model of transcranial permanent distal MCAO which produces consistent cortical infarction of a size corresponding to damage imposed by the majority of human ischemic strokes.

Farelerde Modelleme İnme: Distal Orta Serebral Arter Daimi Pıhtılaşma

Stroke is the third most common cause of death and a main cause of acquired adult disability in developed countries. Only very limited therapeutical ...

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.

Sıçanlar Homolog Kan pıhtısı ile İskemik İnme emboli Orta Serebral Arter Tıkanıklığı (MCAO)

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

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.

Farelerde Orta Serebral Arter Tıkanıklığı ve Reperfüzyon Kaynaklı İnme Cerrahi Yaklaşım

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.

P10 Sıçanlarda Yenidoğan İnme Geçici Orta Serebral Arter Tıkanıklığı Modeli

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