Cerrahi teknik, başlangıçta, Medical College of Georgia Dr William D. Hill laboratuvarda elde edildi. Yazarlar ayrıca, Dr. David A. Rempe ve Landa Prifti diseksiyon kamera kullanımı için teşekkür etmek istiyorum. Bu araştırma Ulusal Sağlık Enstitüsü, NS041744, NS051279, F31 NS064700 ve AHA 30815697D tarafından desteklenmiştir.

Bu protokol, Rochester komitesinin araştırma hayvanların etik (UÇAR) ayrılmış Üniversitesi tarafından onaylanmıştır. Aseptik teknikler protokol sırasında takip edilmelidir. Steril eldiven ve maske kullanılması gereklidir. Protokol sırasında kullanılan ekipman, malzeme, kimyasallar, araçlar, Tablo 1&#39;de açıklanmıştır. 1. Pre-cerrahi Hazırlık <ol><li> Hemen ameliyat sonrası ve ardından ilk 24 saat post-operatif dönem boyunca her 3-5 saat, 2 saat önce cerrahi buprenorfin (0.05mg/kg) subkutan farelere enjekte. </li><li> Tüm cerrahi aletler, gazlı bez sünger, pamuk uçlu aplikatörler otoklav ile sterilize edin. Cerrahi aletler, sağ kullanmadan önce kuru steril gazlı bez cerrahisi ve hava içinde% 70 etanol içinde tutun. % 70 etanol ile çalışma alanı püskürtün. </li><li> % 3 isofluorane-% 20 oksijen gaz karışımı ile bir anestezik vaporizatör kullanarak fare anestezisi. Debimetre ile oksijen miktarını ayarlayın. Parmak veya kuyruk pinching (tepkisiz) tarafından anestezi seviyesini test edin. % 2 (v / v) isofluorane ile anestezi seviyesini korumak. </li><li> Farenin gözleri suni gözyaşları uygulayın ve cerrahi işlem sırasında gözün zarar görmesini önlemek için dikkatli kullanın. Lateral pozisyonda sağ tarafta fare yerleştirin. </li><li> Tıraş, sol göz ve sol kulak hayvan kesme makineleri kullanarak taban arasındaki sol tarafındaki alanda. Betadin çözüm ve pamuk uçlu aplikatörler alan ve hafifçe ovuşturarak kullanarak% 70 etanol arasında değişen alan temizleyin. Gerektiği kadar tekrarlayın. </li><li> 37 vücut ısısını korumak için bir rektal prob bağlı bir ısıtma yastığı ° C üzerinde fare yerleştirin. Mineral yağ kullanarak rektal prob yerleştirin. </li><li> Mikroskop altında sağ tarafta fare konumunu ve bant ile sabitleyin. Bir gazlı bez sünger bir pencere kesin ve cerrahi alanı kapsayacak. </li></ol> 2. Cerrahi Prosedür ve MCA ligasyon <ol><li> Sol göz ve ince düz makas kullanılarak sol kulak tabanı arasındaki dikey bir kesi olun. Kavisli hemostat cerrahi alanında açık tutmak için kullanın. </li><li> Yaylı makas kullanılarak temporal kas yatay bir kesi yapmak ve biraz forseps ile yavaşça çekerek kafatası temporal kas ayrı. </li><li> Kavisli bir forseps ile çene kemiği tutarak bir 18G iğne kullanarak zigomatik ark ve squamosal kemiğin birleştiği küçük bir subtemporal kraniyotomi olun. </li><li> MCA ile kafatası kemik rongeurs küçük parçalara kaldırarak Açığa. Zigomatik ark ve orbital içeriği bu işlem sırasında hasar olmamalıdır. Bu işlem sırasında doku aşırı kuruma oluşursa, pamuk uçlu aplikatörler ile steril PBS geçerlidir. </li><li> Küçük damar cauterizer kullanarak MCA distal kısmının Arter. </li><li> Temporal kas orijinal konumuna geri koyun ve 5-0 naylon sütür cerrahi kesi sitenizi kapatın. </li><li> Anestezi durdurun ve rektal prob kaldırmak. 2. doz subkutan buprenorfin (0.05mg/kg) bir fare enjekte edilir. Bir ısıtma paneli ile 37 ° C tutuldu kafesine fare dön. </li><li> Sonraki 24 saat süreyle herhangi bir rahatsızlık (iştah azalması / su tüketimi, hunched duruş, artan solunum, saç Pilo-dikilen) için fare yakından izlemek. 24 saat için buprenorfin subkutan her 3-5 saatlik bir ameliyat sonrası fare enjekte edilir. Bu dönemde kurtarma jel ile fare sağlayın. </li></ol> 3. TTC Boyama ve İnme Hacminin Belirlenmesi <ol><li> Ameliyat sonrası yirmi dört saat derin fare uyutmak transcardially fosfat% 4 TTC (w / v) kullanarak 15 dakika sonra 10 dakika için% 4 paraformaldehid çözüm için tamponlu salin (PBS) ile fare serpmek orta akış hızında mini peristaltik pompa. Beyin çıkarın ve bir gecede% 4 paraformaldehid çözüm içine yerleştirin. </li><li> Beyin kesit bloğu içine yerleştirin. Beyin tıraş bıçakları kullanarak 1mm kalınlığında dilimler halinde dilimleyin. </li><li> Milimetrenin ölçekli cetvel yanında dilimleri yerleştirin. Diseksiyon mikroskobu bağlı bir dijital kamera kullanarak dilimleri Fotoğraf. </li><li> Görüntü J Yazılım (http://rsbweb.nih.gov/ij/) kullanarak karşı sitenin toplam alanı ipsilateral sitesi olmayan enfarkte alan çıkarılarak vuruş alanını hesaplayınız. Atım hacmi dilimleri 22 kontur istifleme alanı hesaplayın. </li></ol> 4. Yol Görüntü J yazılım kullanımı <ol><li> Dosya menüsünde tıklayarak Görüntü J yazılım analiz edilecek görüntü dosyasını açın. </li><li> Programında &#39;düz çizgi&#39; sekmesini tıklayın. Cetvel üzerinde iki kenar boşlukları arasındaki düz bir çizgi çizin. &#39;Analiz&#39; menüsünü tıklayın ve &#39;set ölçekli&#39; seçeneğini seçin. 1 olarak bilinen mesafe, mm gibi uzunluk birimi ayarlayın. </li><li> &#39;Freehand daire&#39; sekmesini tıklayın. Kontralateral yarımkürede anahat bir daire çizin. &#39;Analiz&#39; menüsünü tıklayın ve &#39;tedbir&#39; seçeneğini seçin. Çizilen dairenin alanı gösteren hesaplama penceresi açılır. </li><li> Ipsilateral hemisferde hariç inme (beyaz) alanı çevresinde başka bir daire çizin. Adım 4.3 &#39;te belirtilen bölge ölçün. Yeni değeri hesaplama penceresine eklenecektir. 1. ve 2. değerler arasındaki fark, aşağıdaki formül olarak gösterilir infarkt alanında temsil eder. </li><li> Adımları 4.2-4.4 tekrarlayarak her dilim vuruş alanını hesaplayınız. (ΣA n) her dilim hesaplanan inme alanının toplamı alın. Bu her dilim kalınlığı 1mm olduğu gerçeğini bilerek atım hacmi temsil eder. ΣA n x 1mm (her dilim kalınlığı) = Strok hacmi </li></ol> 5. Temsilcisi Sonuçlar: Fare kalıcı MCA ligasyonu ile elde infarktlar genellikle kortikal. Ancak, MCA lentikülostriat dal proksimalinde bağlandı ise subkortikal lezyonlar elde etmek mümkündür. MCA ligasyonu sonrası felç hacimleri, 10mm ³, 35mm ³ 19, 23 değişebilir. Inme hacimleri ise 23 20mm, 35mm ³ ³ arasında Filiano ark MRI görüntülerinden tespit Moyanova ark TTC boyama ile hesaplanan atım hacimlerine 19, 10mm, 22mm ³ ³ arasında. Belki ligasyon tam yerini ve inme hacimlerini ölçmek için kullanılan farklı yöntemler bu farklılıkların olası nedeni. Şekil 1, vahşi tip C57/BL6 fare TTC boyanmış beyin 24 saat yazılan kalıcı MCA ligasyonu gösterilmiştir. İnme sitesi görünümü (Şekil 1A, B) beyaz. Şekil 1A ve B farklı açılardan infarkt sitesi göstermektedir. Şekil 2 (Şekil 2A-G) posterior anterior gelen TTC lekeli inme beynin 1mm kalınlığında dilimler gösterir. 24 saat ameliyat sonrası infarkt hacmi hesaplandı. Infarkt hacmi ~ 23mm ³. <img src="/files/ftp_upload/3039/3039fig1.jpg" alt="Şekil 1"/> Şekil 1. İnme sitesi. AB Temsilciliği, TTC MCA ligasyonu sonrası tüm beyin görüntüleri, 24 saat boyandı. Beyaz alan infarkt temsil eder. <img src="/files/ftp_upload/3039/3039fig2.jpg" alt="Şekil 2"/> Şekil 2. Temsilcisi sonuçları (A&gt; G), TTC lekeli 1mm beyin bölümleri, 24 saat sonra MCAL. Dilimler posterior anterior hizalanır. Beyaz alan infarkt temsil eder.

<ol>
	<li>Britton, M., Rafols, J., Alousi, S., Dunbar, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effects+of+middle+cerebral+artery+occlusion+on+central+nervous+system+apoptotic+events+in+normal+and+diabetic+rats.">The effects of middle cerebral artery occlusion on central nervous system apoptotic events in normal and diabetic rats.</a> Int J Exp Diabesity Res. 4, 13-20 (2003).</li><li>Ciceri, P., Rabuffetti, M., Monopoli, A., Nicosia, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Production+of+leukotrienes+in+a+model+of+focal+cerebral+ischaemia+in+the+rat.">Production of leukotrienes in a model of focal cerebral ischaemia in the rat.</a> Br J Pharmacol. 133, 1323-1329 (2001).</li><li>Jin, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Delayed+transplantation+of+human+neural+precursor+cells+improves+outcome+from+focal+cerebral+ischemia+in+aged+rats.">Delayed transplantation of human neural precursor cells improves outcome from focal cerebral ischemia in aged rats.</a> Aging Cell. 9, 1076-1083 (2010).</li><li>McCaig, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evolution+of+GADD34+expression+after+focal+cerebral+ischaemia.">Evolution of GADD34 expression after focal cerebral ischaemia.</a> Brain Res. 1034, 51-61 (2005).</li><li>Shirotani, T., Shima, K., Chigasaki, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+studies+of+extracellular+metabolites+in+the+striatum+after+distal+middle+cerebral+artery+occlusion+in+stroke-prone+spontaneously+hypertensive+rats.">In vivo studies of extracellular metabolites in the striatum after distal middle cerebral artery occlusion in stroke-prone spontaneously hypertensive rats.</a> Stroke. 26, 878-884 (1995).</li><li>Wu, Y. P., Tan, C. K., Ling, E. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Expression+of+Fos-like+immunoreactivity+in+the+brain+and+spinal+cord+of+rats+following+middle+cerebral+artery+occlusion.">Expression of Fos-like immunoreactivity in the brain and spinal cord of rats following middle cerebral artery occlusion.</a> Exp Brain Res. 115, 129-136 (1997).</li><li>Tamura, A., Graham, D. I., McCulloch, J., Teasdale, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Focal+cerebral+ischaemia+in+the+rat:+1.+Description+of+technique+and+early+neuropathological+consequences+following+middle+cerebral+artery+occlusion.">Focal cerebral ischaemia in the rat: 1. Description of technique and early neuropathological consequences following middle cerebral artery occlusion.</a> J Cereb Blood Flow Metab. 1, 53-60 (1981).</li><li>Bederson, J. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rat+middle+cerebral+artery+occlusion:+evaluation+of+the+model+and+development+of+a+neurologic+examination.">Rat middle cerebral artery occlusion: evaluation of the model and development of a neurologic examination.</a> Stroke. 17, 472-476 (1986).</li><li>Carswell, H. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+and+gender+influences+on+sensitivity+to+focal+cerebral+ischemia+in+the+stroke-prone+spontaneously+hypertensive+rat.">Genetic and gender influences on sensitivity to focal cerebral ischemia in the stroke-prone spontaneously hypertensive rat.</a> Hypertension. 33, 681-685 (1999).</li><li>Mary, V., Wahl, F., Uzan, A., Stutzmann, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enoxaparin+in+experimental+stroke:+neuroprotection+and+therapeutic+window+of+opportunity.">Enoxaparin in experimental stroke: neuroprotection and therapeutic window of opportunity.</a> Stroke. 32, 993-999 (2001).</li><li>Menzies, S. A., Hoff, J. T., Betz, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Middle+cerebral+artery+occlusion+in+rats:+a+neurological+and+pathological+evaluation+of+a+reproducible+model.">Middle cerebral artery occlusion in rats: a neurological and pathological evaluation of a reproducible model.</a> Neurosurgery. 31, 100-107 (1992).</li><li>Iaci, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glial+growth+factor+2+promotes+functional+recovery+with+treatment+initiated+up+to+7+days+after+permanent+focal+ischemic+stroke.">Glial growth factor 2 promotes functional recovery with treatment initiated up to 7 days after permanent focal ischemic stroke.</a> Neuropharmacology. 59, 640-649 (2010).</li><li>Richard, M. J. P., Khan, B. V. C. B. J., Saleh, T. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cellular+mechanisms+by+which+lipoic+acid+confers+protection+during+the+early+stages+of+cerebral+ischemia:+A+possible+role+for+calcium.">Cellular mechanisms by which lipoic acid confers protection during the early stages of cerebral ischemia: A possible role for calcium.</a> Neuroscience Research. , (2011).</li><li>Heiss, W. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Progressive+derangement+of+periinfarct+viable+tissue+in+ischemic+stroke.">Progressive derangement of periinfarct viable tissue in ischemic stroke.</a> J Cereb Blood Flow Metab. 12, 193-203 (1992).</li><li>Nedergaard, M., Gjedde, A., Diemer, N. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Focal+ischemia+of+the+rat+brain:+autoradiographic+determination+of+cerebral+glucose+utilization,+glucose+content,+and+blood+flow.">Focal ischemia of the rat brain: autoradiographic determination of cerebral glucose utilization, glucose content, and blood flow.</a> J Cereb Blood Flow Metab. 6, 414-424 (1986).</li><li>Nowicki, J. P., Assumel-Lurdin, C., Duverger, D., MacKenzie, E. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Temporal+evolution+of+regional+energy+metabolism+following+focal+cerebral+ischemia+in+the+rat.">Temporal evolution of regional energy metabolism following focal cerebral ischemia in the rat.</a> J Cereb Blood Flow Metab. 8, 462-473 (1988).</li><li>Butcher, S. P., Bullock, R., Graham, D. I., McCulloch, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Correlation+between+amino+acid+release+and+neuropathologic+outcome+in+rat+brain+following+middle+cerebral+artery+occlusion.">Correlation between amino acid release and neuropathologic outcome in rat brain following middle cerebral artery occlusion.</a> Stroke. 21, 1727-1733 (1990).</li><li>Arlicot, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Detection+and+quantification+of+remote+microglial+activation+in+rodent+models+of+focal+ischaemia+using+the+TSPO+radioligand+CLINDE.">Detection and quantification of remote microglial activation in rodent models of focal ischaemia using the TSPO radioligand CLINDE.</a> Eur J Nucl Med Mol Imaging. 37, 2371-2380 (2010).</li><li>Moyanova, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Protective+role+for+type+4+metabotropic+glutamate+receptors+against+ischemic+brain+damage.">Protective role for type 4 metabotropic glutamate receptors against ischemic brain damage.</a> J Cereb Blood Flow Metab. , (2010).</li><li>Ortolano, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Advances+in+imaging+of+new+targets+for+pharmacological+intervention+in+stroke:+real-time+tracking+of+T-cells+in+the+ischaemic+brain.">Advances in imaging of new targets for pharmacological intervention in stroke: real-time tracking of T-cells in the ischaemic brain.</a> Br J Pharmacol. 159, 808-811 (2010).</li><li>O'Neill, M. J., A, C. J. <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+cerebral+ischemia.">Rodent models of focal cerebral ischemia.</a> Current Protocols in Neuroscience. , 9.6.1-9.6.32 (2000).</li><li>Lin, T. N., He, Y. Y., Wu, G., Khan, M., Hsu, C. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+brain+edema+on+infarct+volume+in+a+focal+cerebral+ischemia+model+in+rats.">Effect of brain edema on infarct volume in a focal cerebral ischemia model in rats.</a> Stroke. 24, 117-121 (1993).</li><li>Filiano, A. J., Tucholski, J., Dolan, P. J., Colak, G., Johnson, G. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transglutaminase+2+protects+against+ischemic+stroke.">Transglutaminase 2 protects against ischemic stroke.</a> Neurobiol Dis. 39, 334-343 (2010).</li></ol>

Kalıcı MCA ligasyonu yöntemi yüksek tekrarlanabilir infarkt hacmi ve mevcut diğer yöntemlere göre artış post-operatif sağkalım oranları verir. Prosedürün kolay ve kısa bir süre (~ 30 dakika) daha pratik hale getirir. Bu yöntem yaygın olarak, hem de fare ve sıçanlar kullanılır. Bu teknik bir stereomikroskop altında bir invaziv cerrahi gerektirir. Bu nedenle, bir mikroskop altında faaliyet gösteren ve başarılı bir kraniotomi mükemmelleştirmek tecrübesi çok önem...

<table border="1"><tbody><tr><th> ADI </th><th> FİRMA VE KATALOG NUMARASI </th></tr><tr><td> A. Çözümleri ve Kimyasallar </td><td></td></tr><tr><td> Povidon-İyot Çözüm, Cerrahi Scrub </td><td> Aplicare, 82-209 </td></tr><tr><td> % 70 Etanol </td><td> Koptec, V1001 </td></tr><tr><td> 2,3,5 triphenyltetrazolium klorür (TTC) </td><td> Sigma, T8877 </td></tr><tr><td> 4% Paraformaldehit </td><td> Elektron Mikroskopi Bilimler, 159SP </td></tr><tr><td> 7.4: Fosfat Tuzlu, pH Tamponlu </td><td> Üretilen ve laboratuvar sterilize. </td></tr><tr><td> Madeni Yağ </td><td> PML Microbiologicals, R6570 </td></tr><tr><td> Isofluorane </td><td> Butler Şirket, 029.405 </td></tr><tr><td> Buprenorfin (0.3mg/ml) </td><td> Hospira, NDC 0409-2012-32 </td></tr><tr><td> Suni gözyaşları </td><td> Butler Şirket, 007.312 </td></tr><tr><td> Canlandırıcı Jel </td><td> Clear H 2 O, 72-01-5022 </td></tr><tr><td> B. Cerrahi Malzemeleri ve Ekipmanları </td><td></td></tr><tr><td> C57/BL6 fare, 10-12 haftalık </td><td> Koloni üniversite vivaryum yetiştirilen </td></tr><tr><td> % 100 oksijen deposu </td><td> Airgas </td></tr><tr><td> Anestezik Buharlaştırıcı ve Akış metre </td><td> Surgivet, Model 100 </td></tr><tr><td> Rektal Probe Isıtma Pedi </td><td> Güzel Bilim Araçları, TR200 </td></tr><tr><td> Diseksiyon mikroskobu </td><td> Zeiss, STEMI 2000 </td></tr><tr><td> Isıtma paneli </td><td> Petco Hizmetleri Ltd 0307-013 </td></tr><tr><td> Küçük Gemi Cauterizer Seti </td><td> Güzel Bilim Araçları, 18.000-00 </td></tr><tr><td> Mini Peristaltik Pompa </td><td> Harvard Apparatus, MPII </td></tr><tr><td> 1 ml şırınga </td><td> BD, 329.650 </td></tr><tr><td> 18G iğne </td><td> Becton Dickinson, 305.196 </td></tr><tr><td> Cerrahi 5.0 Naylon Sütür </td><td> Ethilon, 698H </td></tr><tr><td> Blok Kesit </td><td> Kent Bilimsel, 1mm </td></tr><tr><td> Tek Kenar Razor Blades </td><td> Elektron Mikroskopi Bilimler, 71.962 </td></tr><tr><td> Etiket Şeridini </td><td> Scienceware, Bel-sanat Ürünler F13463-2075 </td></tr><tr><td> Tül Süngerler </td><td> Kendall curity 2252, Johnson ve Johnson 6415 </td></tr><tr><td> Pamuk İpucu Aplikatörler </td><td> Püriten 806-WCL </td></tr><tr><td> C. Cerrahi Aletler </td><td></td></tr><tr><td> Cerrahi Bahar Makas </td><td> Güzel Bilim Araçları 15.003-08 </td></tr><tr><td> Microdissecting forseps açılı </td><td> Güzel Bilim Araçları 5 / 45 </td></tr><tr><td> Microdissecting forseps kavisli </td><td> Güzel Bilim Araçlar 5 Ti </td></tr><tr><td> İnce düz makas </td><td> Güzel Bilim Araçları 14.568-12 </td></tr><tr><td> Eğimli kemik rongeur </td><td> Güzel Bilim Araçları 16.021-14 </td></tr><tr><td> Hartman Hemostat Eğri </td><td> Güzel Bilim Araçları 13.003-10 </td></tr></tbody></table> Tablo 1: MCA ligasyonu işlemi için kullanılan malzemeler, araç ve kimyasalların listesi. A. Çözümleri ve Kimyasallar, B. Cerrahi Malzemeleri ve Ekipmanları, C. Cerrahi Aletler.

the application of permanent middle cerebral artery ligation in the mouse

Fokal serebral iskemi, inme hastalarında görülen en yaygın türü arasında yer alıyor. Klinik önemi nedeniyle İskemiden sırasında ortaya çıkan olaylardan çalışmak için uygun hayvan modelleri geliştirmek için uzun bir çaba olmuştur. Bu tekniklerin en yaygın olan kemirgenler, birçok farklı hayvan modelleri kullanarak, geçici veya kalıcı fokal veya global iskemi modelleri içermektedir. Kemirgenler 1-6 fokal iskemi modeli olarak da literatürde pMCAo denir kalıcı MCA ligasyonu yöntemi yaygın olarak kullanılır. Bu yöntem başlangıçta Tamura ve arkadaşları tarafından sıçanlarda için tarif edilmiştir. 1981 7. Bu protokol bir kraniotomi MCA ve proksimal bölgelerde elektrokoagülasyon tıkalı erişmek için kullanılır oldu. Infarktlar oklüzyonu konuma bağlı olarak çoğunlukla kortikal ve bazen striatal bölgeleri içerir. Bu teknik, şimdi iyi kurulmuş ve birçok laboratuvarda 8-13 kullanılmaktadır. Erken bu tekniğin kullanımı &quot;infarkt çekirdek&quot; ve &quot;penumbra&quot; 14-16 tanımı ve açıklaması yol açtı, ve genellikle, potansiyeli nöroprotektif bileşikler 10, 12, 13, 17 değerlendirmek için kullanılır . Sıçanlarda yapılan ilk çalışmalar olmasına rağmen, kalıcı MCA ligasyonu küçük modifikasyonlar 18-20 farelerde başarıyla kullanılmaktadır. Bu model, tekrarlanabilir infarktlar ve artan sonrası sağ kalım oranları elde edilir. Iskemik inme insanlarda yaklaşık% 80 MCA alan 21 olur ve böylece bu vuruş çalışmaları için bir çok ilgili bir model . Şu anda, orada inmeli hastalarda etkili tedaviler oldukça kıt ve böylece potansiyel farmakolojik bileşikler test ve fizyolojik sonuçlarını değerlendirmek için iyi bir model için bir ihtiyaç vardır. Bu yöntem ayrıca, in vivo hücre içi hipoksi yanıt mekanizmaları eğitim için kullanılabilir . Burada, bir C57/BL6 fare MCA ligasyonu cerrahi sunuyoruz. Biz ameliyat öncesi hazırlık, MCA ligasyonu cerrahi ve infarkt hacimleri ölçümü için 2,3,5 triphenyltetrazolium klorür (TTC) ile boyanarak açıklar.

Watch this Scientific Journal Video about The Application Of Permanent Middle Cerebral Artery Ligation in the Mouse at JoVE.com

The Application Of Permanent Middle Cerebral Artery Ligation in the Mouse

Orta serebral arter (MCA) ligasyonu, hayvan modellerinde fokal serebral iskemi çalışma bir tekniktir. Bu yöntemde, orta serebral arter kraniotomi maruz kalmaktadır ve koterizasyon tarafından bağlandı. Bu yöntem son derece tekrarlanabilir infarkt hacimleri ve mevcut diğer yöntemlere göre artış post-operatif sağkalım oranları verir.

Fare Daimi Orta Serebral Arter ligasyon Of Uygulama

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 ...

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19.5K Görüntüleme.  University of Rochester. Orta serebral arter (MCA) ligasyonu, hayvan modellerinde fokal serebral iskemi çalışma bir tekniktir. Bu yöntemde, orta serebral arter kraniotomi maruz kalmaktadır ve koterizasyon tarafından bağlandı. Bu yöntem son derece tekrarlanabilir infarkt hacimleri ve mevcut diğer yöntemlere göre artış post-operatif sağkalım oranları verir.

Video: The Application Of Permanent Middle Cerebral Artery Ligation in the Mouse

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 ...

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 ...

Permanent Cerebral Vessel Occlusion via Double Ligature and Transection

Stroke is a leading cause of death, disability, and socioeconomic loss worldwide. The majority of all strokes result from an interruption in blood flow (ischemia) 1. Middle cerebral artery (MCA) delivers a great majority of blood to the lateral surface of the cortex 2, is the most common site of human stroke 3, and ischemia within its territory can result in extensive dysfunction or death 1,4,5. Survivors of ischemic stroke often suffer loss or disruption of motor capabilities, sensory deficits, and infarct. In an effort to capture these key characteristics of stroke, and thereby develop effective treatment, a great deal of emphasis is placed upon animal models of ischemia in MCA.
Here we present a method of permanently occluding a cortical surface blood vessel. We will present this method using an example of a relevant vessel occlusion that models the most common type, location, and outcome of human stroke, permanent middle cerebral artery occlusion (pMCAO). In this model, we surgically expose MCA in the adult rat and subsequently occlude via double ligature and transection of the vessel. This pMCAO blocks the proximal cortical branch of MCA, causing ischemia in all of MCA cortical territory, a large portion of the cortex. This method of occlusion can also be used to occlude more distal portions of cortical vessels in order to achieve more focal ischemia targeting a smaller region of cortex. The primary disadvantages of pMCAO are that the surgical procedure is somewhat invasive as a small craniotomy is required to access MCA, though this results in minimal tissue damage. The primary advantages of this model, however, are: the site of occlusion is well defined, the degree of blood flow reduction is consistent, functional and neurological impairment occurs rapidly, infarct size is consistent, and the high rate of survival allows for long-term chronic assessment.

Permanent Cerebral Vessel Occlusion via Double Ligature and Transection

We describe a highly reproducible method for the permanent occlusion of a rodent major cerebral blood vessel. This technique can be accomplished with very little peripheral damage, minimal blood loss, a high rate of long-term survival, and consistent infarct volume commensurate with the human clinical population.

Kalıcı Serebral Gemi Tıkanıklığı<em&gt; Ile</em&gt; Çift Ligature ve transeksiyonu

Stroke is a leading cause of death, disability, and socioeconomic loss worldwide. The majority of all strokes result from an interruption in blood flow ...

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 ...

Permanent Ligation of the Left Anterior Descending Coronary Artery in Mice: A Model of Post-myocardial Infarction Remodelling and Heart Failure

Heart failure is a syndrome in which the heart fails to pump blood at a rate commensurate with cellular oxygen requirements at rest or during stress. It is characterized by fluid retention, shortness of breath, and fatigue, in particular on exertion. Heart failure is a growing public health problem, the leading cause of hospitalization, and a major cause of mortality. Ischemic heart disease is the main cause of heart failure.
Ventricular remodelling refers to changes in structure, size, and shape of the left ventricle. This architectural remodelling of the left ventricle is induced by injury (e.g., myocardial infarction), by pressure overload (e.g., systemic arterial hypertension or aortic stenosis), or by volume overload. Since ventricular remodelling affects wall stress, it has a profound impact on cardiac function and on the development of heart failure. A model of permanent ligation of the left anterior descending coronary artery in mice is used to investigate ventricular remodelling and cardiac function post-myocardial infarction. This model is fundamentally different in terms of objectives and pathophysiological relevance compared to the model of transient ligation of the left anterior descending coronary artery. In this latter model of ischemia/reperfusion injury, the initial extent of the infarct may be modulated by factors that affect myocardial salvage following reperfusion. In contrast, the infarct area at 24 hr after permanent ligation of the left anterior descending coronary artery is fixed. Cardiac function in this model will be affected by 1) the process of infarct expansion, infarct healing, and scar formation; and 2) the concomitant development of left ventricular dilatation, cardiac hypertrophy, and ventricular remodelling.
Besides the model of permanent ligation of the left anterior descending coronary artery, the technique of invasive hemodynamic measurements in mice is presented in detail.

Heart failure is the leading cause of hospitalization and a major cause of mortality. A model of permanent ligation of the left anterior descending coronary artery in mice is applied to investigate ventricular remodelling and cardiac dysfunction post-myocardial infarction. The technique of invasive hemodynamic measurements in mice is presented.

Farelerde Sol Ön İnen Koroner Arter Daimi ligasyonu: Post-miyokard Enfarktüs Remodelling ve Kalp Yetmezliği Bir Model

Heart failure is a syndrome in which the heart fails to pump blood at a rate commensurate with cellular oxygen requirements at rest or during stress. It ...

Performing Permanent Distal Middle Cerebral with Common Carotid Artery Occlusion in Aged Rats to Study Cortical Ischemia with Sustained Disability

Stroke typically occurs in elderly people with a range of comorbidities including carotid (or other arterial) atherosclerosis, high blood pressure, obesity and diabetes. Accordingly, when evaluating therapies for stroke in animals, it is important to select a model with excellent face validity. Ischemic stroke accounts for 80% of all strokes, and the majority of these occur in the territory of the middle cerebral artery (MCA), often inducing infarcts that affect the sensorimotor cortex, causing persistent plegia or paresis on the contralateral side of the body. We demonstrate in this video a method for producing ischemic stroke in elderly rats, which causes sustained sensorimotor disability and substantial cortical infarcts. Specifically, we induce permanent distal middle cerebral artery occlusion (MCAO) in elderly female rats by using diathermy forceps to occlude a short segment of this artery. The carotid artery on the ipsilateral side to the lesion was then permanently occluded and the contralateral carotid artery was transiently occluded for 60 min. We measure the infarct size using structural T2-weighted magnetic resonance imaging (MRI) at 24 hr and 8 weeks after stroke. In this study, the mean infarct volume was 4.5% &#177; 2.0% (standard deviation) of the ipsilateral hemisphere at 24 hr (corrected for brain swelling using Gerriet&#8217;s equation, n = 5). This model is feasible and clinically relevant as it permits the induction of sustained sensorimotor deficits, which is important for the elucidation of pathophysiological mechanisms and novel treatments.

Here we present a protocol to produce permanent distal middle cerebral artery occlusion in elderly female rats with simultaneous occlusion of the carotid arteries to generate large cortical infarcts and sustained deficits. We show confirmation of the lesion size using structural MRI at 24 hr and 8 weeks after stroke.

Yaşlı Sıçanlarda Ortak Karotis Arter Tıkanıklığı ile Kalıcı Distal Orta Serebral Sahne Kortikal İskemi Sürekli Engelli Eğitim için

Stroke typically occurs in elderly people with a range of comorbidities including carotid (or other arterial) atherosclerosis, high blood pressure, ...

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 ...

Middle Cerebral Artery Occlusion Allowing Reperfusion via Common Carotid Artery Repair in Mice

The ischemic stroke is a major cause of adult long-term disability and death worldwide. The current treatments available are limited, with only tissue plasminogen activator (tPA) as an approved drug treatment to target ischemic strokes. Current research in the field of ischemic stroke focuses on better understanding the pathophysiology of stroke, to develop and investigate novel pharmaceutical targets. Reliable experimental stroke models are crucial for the progression of potential treatments. The middle cerebral artery occlusion (MCAO) model is clinically relevant and the most frequently used surgical model of ischemic stroke in rodents. However, the outcomes of this model, such as lesion volume, are associated with high levels of variability, particularly in mice. The alternative MCAO model described here allows the reperfusion of the common carotid artery (CCA) and the increased perfusion of the middle cerebral artery (MCA) territory, using a tissue pad with fibrinogen-based sealant to repair the vessel, and the improved welfare of the mice by avoiding external carotid artery (ECA) ligation. This reduces the reliance on the Circle of Willis, which is known to be highly anatomically variable in mice. Representative data show that using this alternative surgical approach decreases the variability in lesion volumes between the traditional MCAO approach and the alternative approach described here.

Intraluminal filament occlusion of the middle cerebral artery is the most frequently used in vivo model of experimental stroke in rodents. An alternative surgical approach to allow common carotid artery repair is performed here, which allows the reperfusion of the common carotid artery and a full reperfusion to the middle cerebral artery territory.

Orta serebral arter tıkanıklığı ortak karotid arter onarım ile reperfüzyon farelerde izin

The ischemic stroke is a major cause of adult long-term disability and death worldwide. The current treatments available are limited, with only tissue ...