이 작품은 부여 계약 N ° 201024과 N ° 202,213 (유럽 스트로크 네트워크)에서 유럽 공동체의 7 번째 프레임 워크 프로그램 (FP7/2007-2013)에 의해 재정 지원되었다. 추가 자금은 Bundesministerium FÜR Bildung 놀이 Forschung (행정 연구 베를린 센터)와 독일 Forschungsgemeinschaft (Exzellenzcluster NEUROCURE)에 의해 주어졌다. 저자는 애니메이션 제작을위한 운영 및 엘크 루드비히 (샤히떼 비디오 서비스) 동안 지원 Mareike 티엘크을 (실험적 신경과에 대한 Dep., 샤히떼 베를린) 감사드립니다.

높은 품질과 재현성을 보장하기 위해, 우리는 표준 운영 절차 (SOP)의 사용을 권장합니다. 이 비디오에서는, 같은 연구실에서 개발하고 사용하는 출판 솝스이 적용됩니다 1. 1. 중간 대뇌 동맥 폐색 <ol><li> 마우스는 수의 직원들과 상담에 적절한 마취 정권과 anaesthetized 있습니다. (1.5 예를 들어 유도 - 1.0 2% Isoflurane 및 유지 보수 - 3분의 2 N2O와 기화기를 사용하여 3분의 1 O2에서 Isoflurane 1.5 %). <ol><li> 생쥐의 체온은 36.5 ° C에서 ± 0.5 ° C 열판과 함께 수술하는 동안 유지됩니다. 마우스의 직장 온도에 따라 체온이 피드백 제어 가열 패드는, 매우 좋습니다. </li><li> 적절한 에이전트 (예 : 70% 에틸 알코올)과 피부와 주변 모피를 소독하고 나중에 그것을 건조. </li></ol></li><li> 정중선 목 절개가 만들어이며, 부드러운 조직은 따로 뽑아 있습니다. </li><li> 왼쪽 일반 경동맥 (LCCA)는 신중하게 주변 신경 (vagal 신경을 해치지 않고) 및 묶인이 6.0/7.0 문자열을 사용하여 만든에서 무료로 해부하는 것입니다. 5.0 문자열도 사용할 수 있습니다. </li><li> 왼쪽 외부 경동맥의 동맥 (LECA)는 다음 구분하고 두 번째 매듭이 이루어집니다. </li><li> 다음, 왼쪽 내부 경동맥의 동맥 (LICA)는 고립되고 매듭은 6.0 필라멘트와 준비가되어 있습니다. </li><li> 왼쪽 내부 경동맥의 동맥 (LICA)과 왼쪽 pterygopalatine 동맥 (LPA)의 좋은 경치를 취득 후, 두 동맥은 microvascular 클립을 사용하여 잘린 있습니다. </li><li> 그것은 LECA과 LICA에 bifurcates 전에 작은 구멍이 LCCA에서 절단합니다. 그것은 클립에서 멈출 때까지 실리콘 경화제 혼합물 (아래 참조)과 코팅 8.0 나일론으로 만든 monofilament는 다음 LICA에 소개됩니다. 관심은 후두 동맥을 입력하지 지불되어야합니다. (그림 1) </li><li> 필라멘트는 윌리스의 원안에 LMCA의 기원을 막다하는 LICA에 삽입되는 동안 잘린 동맥이 열립니다. </li><li> LICA에 세 번째 매듭은 위치에 필라멘트를 해결하기 위해 닫힙니다. </li><li> 마우스는 볼륨 보충으로 subcutaneously 호수 0.5 ML를 받게됩니다. 통증의 경우, Lidocaine 젤은 몸에 붙이기도 상처에 적용됩니다. </li><li> reperfusion가 의도하는 경우, 마우스가 30 숙박 - 온수 케이지 90 분 폐색, 상처는 작은 봉합 클립과 함께 폐쇄 수 있습니다. 이후, 두 번째 마취가 수행, ICA에서 세 번째 매듭이 순간 열리고 필라멘트가 철회됩니다. </li><li> 나머지 봉합이 짧아되고 피부는 수술 치료와 적응합니다. </li><li> 모든 동물은 위에서 설명한대로 두 번째 볼륨 보충을 받게됩니다. </li><li> 동물은 체온의 제어 두 시간 동안 열띤 새장에 넣어 있습니다. <ol><li> 동물 불편의 흔적에 대한 수술 후 매일 점검해야합니다. 마우스 일부 게시물 외과 체중 감량을 보여줄 수 있어요. 그들은 먹는 장려하는 바닥에 놓인 배양 - 접시에 으깬 음식을받을 수 있습니다. 음식 7 일 동안 매일 바뀝니다. </li></ol></li></ol> 2. 섐 운영 <ol><li> 가짜 작업에 필라멘트가 LMCA을 막다에 삽입 및 인스턴트 reperfusion (1.8)을 허용하는 즉시 철회합니다. 두 번째 마취를 포함하여 - 후속 작업 대뇌 ischaemia (1.14 1.9)를받은 동물에서 수행되는 동일합니다. </li></ol> 3. 필라멘트의 준비 <ol><li> 필라멘트의 불임을 고려하여야한다. 멸균 장비뿐만 아니라 필라멘트의 적절한 처리의 사용은 나중에 불임 수술을위한 필수 조건이다. 일반적인 살균 방법의 많은 필라멘트의 품질을 악화시킬 수 있으므로 필라멘트의 소독은 어렵습니다. 그러나 이러한 방사선으로 방법, 자외선 또는 γ 선, 또는 화학 살균과 함께 예를 들어, 산화 에틸렌과 같은 반응성이 매우 높은 가스와 예를 들어, 적용됩니다. </li><li> 8.0 나일론 필라멘트는 현미경으로 11mm의 길이로 절단됩니다 </li><li> 필라멘트 팁은 Xantopren M 점막 및 활성 NF Optosil의 경화제의 혼합 8 mm의 길이를 통해 완전히 고르게 코팅되어야합니다 </li></ol> 4. 대표 결과 다른 모터와 행동 적자의 결과, 혈액 흐름 제한의 기간에 따라 다릅니다. 모두 30 뇌성 ischaemia 60 분, 이후 대부분의 경우 동물 측면 밀어 감소 저항을 표시하고 운동의 방해로 인해 돌고. 전면 리에서 flexor 위치로 Milder 병변 명단MBS. 이 쉽게 관찰할 표지판이 작전의 성공을위한 기본 점수로 사용할 수 있습니다. 2 Morphologically 병변은 조직학 또는 자기 공명 영상 (MRI)을 사용하여 평가하실 수 있습니다. ischaemia 30 분 striatum에 한정 주로의 연결을 세포 사망의 원인 반면, 중간 대뇌 동맥의 폐색 60 분이 striatum과 네크로 텍스 모두 포함하는 영역에서 조직 pannecrosis을 생산하고 있습니다. 3 (그림 2)를 경색 볼륨 측면에서, 우리는 기대 운영 일련의 30 %보다 낮은 표준 편차. 60 분 후 20 % - 사망률은 ischaemia 10 30 분 후에 약 5 % 폐색 시간에 따라 달라집니다. 또 최소한의 침입 가능성은 성공의 직접적인 제어를 허용하는 작업 동안 레이저 도플러 flowmetry (LDF)의 사용이다. 개별 동물에서 10 감소 - preocclusion 값의 20 %가 명확하게 초점 대뇌 국소 빈혈의 성공적인 유도를 나타내는 4가 그러나, LDF는 interindividual 비교하는 방법으로 사용할 수 없습니다 LDF는 혈액의 양적 변화 (백분율)를 측정할 수 있기 때문입니다. 작고 제한된 조직 샘플 볼륨 내에서 흐름. 그것은 혈류 감소의 공간적 범위에 대한 정보를 제공하지 않습니다 5. 보행 분석 6,7, Rotarod 8, 폴 테스트 9,10, 접착제 제거 시험 11,12, 계단 테스트 13,14, 래더 렁 테스트 15,16와 모리스 물 미로를 포함하여 뇌졸중 후 행동 측면을 평가하기 위해 몇 가지 테스트가 있습니다 17. 모든 테스트에서, MCAo를 받게 쥐가 덜 성공적으로 제어 동물보다 수행합니다. <img src="/files/ftp_upload/2423/2423fig1.jpg" alt="그림 1" /> 그림 1. 마우스의 뇌 (배경에 묘사된) 제공하는 선박의 건축 계획. 다른 변종은 유사 나타날 수 있습니다, 예를 들어 후두 동맥 때로는 내부 경동맥에서 떠납니다. <img src="/files/ftp_upload/2423/2423fig2.jpg" alt="그림 2" /> 그림 2. 근위 MCAo 모델에 reperfusion의 다른 시간 지점 이후에 전형적인 병변 크기의 도식 그림. 중간에 기능 활동 MCAo 후 대뇌 혈류의 전형적인 코스는 그려져 있습니다. (MCAo : 중간 대뇌 동맥 폐색, LDF : 레이저 도플러 유량 측정)

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R. <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+suture+size+and+carotid+clip+application+upon+blood+flow+and+infarct+volume+after+permanent+and+temporary+middle+cerebral+artery+occlusion+in+mice.">Effect of suture size and carotid clip application upon blood flow and infarct volume after permanent and temporary middle cerebral artery occlusion in mice.</a> Brain Res. 970, 1-2 (2003).</li><li>Beckmann, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High+resolution+magnetic+resonance+angiography+non-invasively+reveals+mouse+strain+differences+in+the+cerebrovascular+anatomy+in+vivo.">High resolution magnetic resonance angiography non-invasively reveals mouse strain differences in the cerebrovascular anatomy in vivo.</a> Magn Reson Med. 44, 252-258 (2000).</li><li>Barone, F. C., Knudsen, D. J., Nelson, A. H., Feuerstein, G. Z., Willette, R. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+strain+differences+in+susceptibility+to+cerebral+ischemia+are+related+to+cerebral+vascular+anatomy.">Mouse strain differences in susceptibility to cerebral ischemia are related to cerebral vascular anatomy.</a> J Cereb Blood Flow Metab. 13, 683-692 (1993).</li><li>Florian, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-term+hypothermia+reduces+infarct+volume+in+aged+rats+after+focal+ischemia.">Long-term hypothermia reduces infarct volume in aged rats after focal ischemia.</a> Neurosci Lett. 438, 180-185 (2008).</li><li>Noor, R., Wang, C. X., Shuaib, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+hyperthermia+on+infarct+volume+in+focal+embolic+model+of+cerebral+ischemia+in+rats.">Effects of hyperthermia on infarct volume in focal embolic model of cerebral ischemia in rats.</a> Neurosci Lett. 349, 130-132 (2003).</li><li>Barber, P. A., Hoyte, L., Colbourne, F., Buchan, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Temperature-regulated+model+of+focal+ischemia+in+the+mouse:+a+study+with+histopathological+and+behavioral+outcomes.">Temperature-regulated model of focal ischemia in the mouse: a study with histopathological and behavioral outcomes.</a> Stroke. 35, 1720-1725 (2004).</li><li>Shin, H. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mild+induced+hypertension+improves+blood+flow+and+oxygen+metabolism+in+transient+focal+cerebral+ischemia.">Mild induced hypertension improves blood flow and oxygen metabolism in transient focal cerebral ischemia.</a> Stroke. 39, 1548-1555 (2008).</li><li>Bottiger, B. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Global+cerebral+ischemia+due+to+cardiocirculatory+arrest+in+mice+causes+neuronal+degeneration+and+early+induction+of+transcription+factor+genes+in+the+hippocampus.">Global cerebral ischemia due to cardiocirculatory arrest in mice causes neuronal degeneration and early induction of transcription factor genes in the hippocampus.</a> Brain Res Mol Brain Res. 65, 135-142 (1999).</li><li>Kapinya, K. J., Prass, K., Dirnagl, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isoflurane+induced+prolonged+protection+against+cerebral+ischemia+in+mice:+a+redox+sensitive+mechanism.">Isoflurane induced prolonged protection against cerebral ischemia in mice: a redox sensitive mechanism.</a> Neuroreport. 13, 1431-1435 (2002).</li><li>Endres, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanisms+of+stroke+protection+by+physical+activity.">Mechanisms of stroke protection by physical activity.</a> Ann Neurol. 54, 582-590 (2003).</li><li>Gertz, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Physical+activity+improves+long-term+stroke+outcome+via+endothelial+nitric+oxide+synthase-dependent+augmentation+of+neovascularization+and+cerebral+blood+flow.">Physical activity improves long-term stroke outcome via endothelial nitric oxide synthase-dependent augmentation of neovascularization and cerebral blood flow.</a> Circ Res. 99, 1132-1140 (2006).</li><li>Richter, S. H., Garner, J. P., Wurbel, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Environmental+standardization:+cure+or+cause+of+poor+reproducibility+in+animal+experiments.">Environmental standardization: cure or cause of poor reproducibility in animal experiments.</a> Nat Methods. 6, 257-261 (2009).</li><li>Liesz, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+spectrum+of+systemic+immune+alterations+after+murine+focal+ischemia:+immunodepression+versus+immunomodulation.">The spectrum of systemic immune alterations after murine focal ischemia: immunodepression versus immunomodulation.</a> Stroke. 40, 2849-2858 (2009).</li><li>Meisel, C., Schwab, J. M., Prass, K., Meisel, A., Dirnagl, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Central+nervous+system+injury-induced+immune+deficiency+syndrome.">Central nervous system injury-induced immune deficiency syndrome.</a> Nat Rev Neurosci. 6, 775-786 (2005).</li><li>Engel, O., Meisel, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Models+of+Infection+Before+and+After+Stroke:+Investigating+New+Targets.">Models of Infection Before and After Stroke: Investigating New Targets.</a> Infect Disord Drug Targets. 9, (2010).</li><li>Dirnagl, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bench+to+bedside:+the+quest+for+quality+in+experimental+stroke+research.">Bench to bedside: the quest for quality in experimental stroke research.</a> J Cereb Blood Flow Metab. 26, 1465-1478 (2006).</li><li>Macleod, M. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Good+laboratory+practice:+preventing+introduction+of+bias+at+the+bench.">Good laboratory practice: preventing introduction of bias at the bench.</a> Stroke. 40, 50-52 (2009).</li><li>Group, S. T. A. I. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recommendations+for+standards+regarding+preclinical+neuroprotective+and+restorative+drug+development.">Recommendations for standards regarding preclinical neuroprotective and restorative drug development.</a> Stroke. 30, 2752-2758 (1999).</li></ol>

동물 실험 Landesamt fuer 아퍼 놀이 Soziales, 베를린, 독일에 의해 명시된 지침과 규정에 따라 수행되었다.

과도, 근위 MCA 폐색 18,19의 모델은 여기서 환자에서 허혈성 뇌졸중의 가장 일반적인 유형 중 하나를 모방한 것이었 제시. 변수 reperfusion 시간에 따라 20, 모델이 일시적 허혈성 발작 (TIA)에서 대형에 이르기까지 손상의 다른 등급을 제공합니다 허혈성 반구의 주요 부분을 포함 infarcts. 이것은 뇌졸중 후 연구자가 다른 pathophysiological 메커니즘을 공부하실 수 있습니다. 20,21 수술은 짧은 기간에 수행하고 높은 재현성 병변을 생성하실 수 있습니다. 그럼에도 불구하고,이 confounders의 철저한 관리가 필요합니다. 작업 기술 22 작은 차이가 경색에 다른 효과에 대한 계정 수 있습니다. 23,24을뿐만 아니라, 뇌성 혈관 해부학 차이로 인해 서로 다른 마우스 변종은 다른 결과를 보여줍니다. 25,26 바디케어 온도가 저체온증 더 심각한 적자에 작은 병변 27 고열로 이어지는으로 신경 손상에 영향을 미칩니다. 따라서 28 일, 온도 제어 및 유지 보수가이 모델에 관련성이있다., 혈압 및 혈액 가스는 또한 29 결과의 중요한 confounders하고 필요 모니터 수 있습니다. 신속하고 최소한의 침입 방법 중 30,31 사용 (비침습 적절한 혈압 측정, 쉬운 접근 혈액 수집 사이트)을 권장합니다. 일부 neuroprotective 효과를 가지고, 및 / 또는 vasodilators 수 있으므로 마취의 선택은 Isoflurane 예를 들어 같은, 또한 매우 중요합니다. 따라서 32, 마취에 대한 노출이 가능하고 표준처럼 짧게해야합니다. 우리는 더 이상보다 15 분 수술을받은 동물을 제외할 수 있습니다. 외과 사이트를 면도하는 것은 microabrasions과 염증을 생산하고 머리 조각을 출시. 이것은 더 이상 염증을 증가하고 뇌졸중 pathophysiology에 영향을 미치는 수 있습니다 현지 감염을 육성 수 있습니다. 주택 조건, 농축 특히 사용, 뇌졸중 결과에 영향을 미칠 수 있으며, 표준화 및 연구 보고서에서 설명한다. 재현성에 대한 환경 농축과 그 효과를 6,33,34 사용 논의의 문제입니다. 35 또 다른 중요한 confounder가 특히 ischaemia 36 이상 시간 이후에 감염 뇌졸중 유발 위험이있는 추가 병적 증가 죽음에 이르게한다. 37,38가 감염은 5 일 3시 경에에서 증상 해짐에 따라,이 같은 모델 후속 장기에 중요한 결과를했습니다. 뇌졸중, 표준화 및 품질 관리에 대한 새로운 치료 전략의 개발에 대한 검색 결과가 관련 생산 잠복기 translational 뇌졸중 연구 매우 중요하다 39 &quot;좋은 실험실 연습&quot;40,41 요구하는가. : <ol><li> 사용 동물의 적절한 자세한 설명; </li><li> 샘플 크기 계산과 예상 효과의 크기에보고; </li><li> 연구하기 전에 설정 포함하고 배제 기준; </li><li> 그룹으로 할당 임의화; </li><li> 조사에 관한 할당 은폐; </li><li> 분석에서 제외 동물을보고; </li><li> 결과 멀게 평가; </li><li> 관심과 연구 기금의 잠재적인 충돌을보고. </li></ol>

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		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Surgical scissors (skin cut)</td>
<td>&nbsp;</td>
<td>FST</td>
<td>14028-10</td>
<td>&nbsp;</td>
<tr>
<td>2 Dumont forceps (Medical #5 straight tip)</td>
<td>&nbsp;</td>
<td>FST</td>
<td>11253-20</td>
<td>&nbsp;</td>
<tr>
<td>Spring scissors (according to Vannas)</td>
<td>&nbsp;</td>
<td>FST</td>
<td>15000-00</td>
<td>&nbsp;</td>
<tr>
<td>Applying forceps for micro clamps</td>
<td>&nbsp;</td>
<td>FST</td>
<td>00072-14</td>
<td>&nbsp;</td>
<tr>
<td>Micro vascular clamp (e.g. S&amp;G B1-V)</td>
<td>&nbsp;</td>
<td>FST</td>
<td>00396-01</td>
<td>Also Serrefine possible</td>
</tr>
<tr>
<td>2 Hemostats according to Hartmann</td>
<td>&nbsp;</td>
<td>FST</td>
<td>13002-10</td>
<td>&nbsp;</td>
<tr>
<td>Needle holder (according Olsen-Hegar or other)</td>
<td>&nbsp;</td>
<td>FST</td>
<td>12002-14</td>
<td>&nbsp;</td>
<tr>
<td>Standard anatomical forceps (for wound closure)</td>
<td>&nbsp;</td>
<td>FST</td>
<td>11000-12</td>
<td>&nbsp;</td>
<tr>
<td>5/0 sutures </td>
<td>&nbsp;</td>
<td>Suprama</td>
<td>&nbsp;</td>
<td>for vascular ligatures</td>
</tr>
<tr>
<td>6/0 sutures </td>
<td>&nbsp;</td>
<td>Suprama</td>
<td>&nbsp;</td>
<td>for skin suture, 5/0 also possible</td>
</tr>
<tr>
<td>Lidocaine (e.g. Xylocain Gel 2%)</td>
<td>&nbsp;</td>
<td>Astra Zeneca</td>
<td>&nbsp;</td>
<td>or other local pain relief</td>
</tr>
<tr>
<td>Dissecting microscope (stereo microscope), magnification 6x to max. 40x</td>
<td>&nbsp;</td>
<td>Leica Zeiss</td>
<td>MZ6 Stemi2000C</td>
<td>&nbsp;</td>
<tr>
<td>Cold light source</td>
<td>&nbsp;</td>
<td>Leica</td>
<td>KL1500</td>
<td>&nbsp;</td>
<tr>
<td>Temperature feedback controlled heating pad system</td>
<td>&nbsp;</td>
<td>FST</td>
<td>21052-00</td>
<td>With mouse pad and small probe</td>
</tr>
<tr>
<td>Anaethesia system for isoflurane</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Isoflurane</td>
<td>&nbsp;</td>
<td>Abott</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Veterinary Recovery Chamber</td>
<td>&nbsp;</td>
<td>Peco Services</td>
<td>V1200</td>
<td>Heated cage</td>
</tr>
<tr>
<td>8.0 nylon filament</td>
<td>&nbsp;</td>
<td>Suprama</td>
<td>TEL181005</td>
<td>for coating</td>
</tr>
<tr>
<td>Scalpel</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>For cutting filament in pieces</td>
</tr>
<tr>
<td>Ruler</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>To cut correct length of filament</td>
</tr>
<tr>
<td>Xantopren M Mucosa</td>
<td>&nbsp;</td>
<td>Heraeus Kulzer</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Activator Universal Plus</td>
<td>&nbsp;</td>
<td>Heraeus Kulzer</td>
<td>&nbsp;</td>
<td>Optosil - Xantopren</td>
</tr>
<tr>
<td>2 Dumont forceps (Medical #5 straight tip)</td>
<td>&nbsp;</td>
<td>FST</td>
<td>11253-20</td>
<td>&nbsp;</td>
<tr>
<td>A soft underlay for storing and grasping the uncoated filaments</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>e.g. swabs, foam, &#x2026;</td>
</tr>
<tr>
<td>Receptacle for new build filaments</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>e.g. petri dish, flexible foam,&#x2026;</td>
</tr>		</tbody>
		</table>
		</div>

modeling stroke in mice - middle cerebral artery occlusion with the filament model

행정 죽음과 특히 고도로 발달한 국가에서 성인 장애의 가장 자주 원인 중 하나입니다. 그러나, 현재까지 치료 옵션이 매우 제한됩니다. 소설 치료 접근 방법의 필요성을 충족시키기 위해, 실험 연구는 뇌졸중 자주 초점 대뇌 ischaemia의 쥐 모델을 사용합니다. 대부분의 연구자들은 생쥐이나 쥐가 중간 대뇌 동맥 (MCA)의 영구 또는 과도 폐색을 사용합니다. intraluminal 봉합 기술 (때문에 필라멘트 또는 봉합 모델이라고도 함)를 통해 중간 대뇌 동맥 (MCA)의 근위 폐색은 아마 실험 뇌졸중 연구에 가장 자주 사용되는 모델입니다. intraluminal MCAO 모델은 비교적 비침습 방식으로 MCA 영역의 재현성 과도 또는 영구적인 ischaemia을 유도하는 장점을 제공합니다. Intraluminal 접근이 동맥의 전체 영토의 혈액 흐름을 방해. 필라멘트 폐색 따라서 체포은 기초 신경을 공급 lenticulo - 흔적 동맥에 근위 흐름. 필라멘트 피질과 striatum에서 재현할 병변의 MCA 결과의 폐색 및 영구 또는 일시적 중 하나가 될 수 있습니다. 반면, 말초 유도 모델 (lenticulo - 흔적 동맥 분기까지) MCA 폐색 일반적으로 striatum을 살려 주로 네크로 텍스 포함됩니다. 또한 이러한 모델은 craniectomy이 필요합니다. 이 문서에서 보여주는 모델에서 실리콘 코팅 필라멘트는 일반적으로 경동맥에 도입하고 중간 대뇌 동맥의 기원을 차단 윌리스의 서클에 내부 경동맥 따라 고급. 환자에서 중간 대뇌 동맥의 허혈성 뇌졸중 occlusions는 가장 일반적인 원인 중입니다. 다양한 허혈성 간격은 허혈성 병변이 심각도 다양한 것은 생산할 수와 reperfusion의 시점에 따라이 모델에서 자유롭게 선택할 수 있기 때문에. 부분적으로 인간의 thromboembolic 응고의 자연 또는 치료 (tPA) 용해 후 혈류의 복원 모델을 최소한 occluding 필라멘트의 제거에 의해 Reperfusion. 이 비디오에서 우리는 기본적인 기술뿐만 아니라이 모델의 예측 가치를 제한할 수있는 주요 함정과 confounders를 제공합니다.

Watch this Scientific Journal Video about Modeling Stroke in Mice - Middle Cerebral Artery Occlusion with the Filament Model at JoVE.com

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

중간 대뇌 동맥의 폐색 Filamentous은 생쥐에서 허혈성 뇌졸중 공부를위한 일반적인 모델입니다.

마우스의 모델링 스트로크 - 필라멘트 모델 중간 대뇌 동맥 폐색

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

modeling-stroke-mice-middle-cerebral-artery-occlusion-with-filament

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JoVE Journal

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91.0K 조회수.  Center for Stroke Research Berlin, Charité Universitätsmedizin. 중간 대뇌 동맥의 폐색 Filamentous은 생쥐에서 허혈성 뇌졸중 공부를위한 일반적인 모델입니다.

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

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.

중간 대뇌 동맥 폐색의 마우스 모델

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 ...

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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.

마우스의 레이저 도플러 Flowmetry 안내와 허혈성 뇌졸중에 대한 중간 대뇌 동맥 Intraluminal 폐색 (MCAO) 모델

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 ...

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.

쥐의 레이저 도플러 Flowmetry 안내와 허혈성 뇌졸중에 대한 엔도 테린-1 유도 중간 뇌 동맥 폐색 모델

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.

엔도 테린-1 유도 중간 뇌 동맥 폐색 동안 중동 뇌동맥을 시각화하기 위해 두개골 창 활용

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.

Intraluminal 중간 대뇌 동맥 폐색의 쥐 뇌졸중 모델에서 뇌 관류 모니터링에 최적화 된 시스템

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.

마우스의 모델링 스트로크 : 말초 중간 대뇌 동맥의 영구 응고

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.

흰쥐의 동종 혈액 혈전과 허혈성 뇌졸중에 대한 전성 중간 대뇌 동맥 폐색 (MCAO)

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.

마우스의 반복 저산소 전처리 및 과도 중간 대뇌 동맥 폐쇄 후 신경 혈관 보호의 정량화

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

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.

세 쥐의 경동맥 폐색으로 영구 말초 중간 대뇌을 수행하면 두피 허혈 지속적인 장애를 연구하는

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

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.