Questo lavoro è stato sostenuto dalle concessioni dal American Heart Association Greater Affiliato Sud-Est (09GRNT2060421), l&#39;American Medical Association, e presso l&#39;Università della Florida clinica e traslazionale Science Institute. Adam Mecca è un NIH / NINDS, compagno NRSA predoctoral (F30 NS-060335). Robert Regenhardt ricevuto il sostegno predoctoral borsa di studio presso l&#39;Università di programma di formazione multidisciplinare in Florida Ipertensione (T32 HL-083810).

Questo protocollo è stato approvato dalla cura degli animali e del Comitato Istituzionale uso (IACUC) presso l&#39;Università della Florida, ed è in conformità con la &quot;Guida per la cura e l&#39;uso di animali da laboratorio&quot; (ottava edizione, Accademia Nazionale delle Scienze, 2011). Materiale <ol><li> Animali: otto settimane di età, di sesso maschile, Sprague Dawley (Charles River, Farms Wilmington, MA, USA) del peso di 250-300 g al momento dell&#39;intervento chirurgico. </li><li> Anestesia <ol class="lalpha"><li> Inalazione sistema di anestesia (VetEquip Inc., Pleasanton, CA, USA) </li><li> Isoflurano anestetico (Farmaceutici Baxter, Deerfield, IL, USA) </li></ol></li><li> Stereotassico sistema (David Kopf Instruments, Tujunga, CA, USA) <ol class="lalpha"><li> Piccolo animale sistema stereotassico </li><li> Non rottura orecchio barre per i ratti </li><li> Gas testa porta l&#39;anestesia per i ratti </li></ol></li><li> Regolazione della temperatura<ol class="lalpha"><li> BAT-12 Termometro microsonda (World Precision Instruments, Inc., Sarasota, FL, Stati Uniti d&#39;America) </li><li> T / POMPA, TP600 coperta termica (Gaymar Industries, Inc., Orchard Park, NY, Stati Uniti d&#39;America) </li></ol></li><li> Strumenti chirurgici <ol class="lalpha"><li> Manico bisturi e # 11 della lama, iris forcipe, Graefe pinze, divaricatori bulldog pinza, cacciavite, siringa da 10 ml con 26 gauge smussato (Strumenti di precisione del Mondo, Inc., Sarasota, FL, Stati Uniti d&#39;America) </li><li> Micromotore trapano e supporto stereotassica, Quintessential stereotassico Injector (Stoelting, Wood Dale, IL, USA) </li><li> 1,0 millimetri fresa trapano rotondo, 1,0 mm di cono rovesciato trapano fresa (Roboz chirurgico Instrument Co., Inc., Gaithersburg, MD, USA) </li></ol></li><li> Forniture chirurgiche <ol class="lalpha"><li> Viti di montaggio 0-80 x 3/32 con 2.4 mm di lunghezza dell&#39;albero, da 21 gauge cannula guida [4 millimetri a lungo sotto il piedistallo] e manichino cannula (Materie plastiche uno, Roanoke, VA, USA) </li><li> Jet acrilico protesi e liquidi (Lang Dental Manufacturing Co., Inc., Wheeling, IL, USA) </li><li> 3,0 nylon sutura (Oasis, Mettawa, IL, USA) </li><li> Tamponi di cotone, unguento oculare Puralube (Fisher Scientific, Pittsburgh, PA, USA) </li><li> Tosatrici elettriche (Oster, Providence, RI, USA) </li></ol></li><li> Prodotti chimici <ol class="lalpha"><li> L&#39;endotelina-1 (American Peptide, Sunnyvale, CA, USA) </li><li> Clorexidina al 2% (Agrilabs, St. Joseph, MO, USA) </li><li> Buprenorfina HCl (Hospira Inc., Lake Forest, IL, USA) </li></ol></li><li> Visualizzazione Attrezzature <ol class="lalpha"><li> Microscopio chirurgico (Strumento Seiler e Manufacturing, St. Louis, MO, USA) </li><li> Fibra ottica illuminatore (TechniQuip Corp., Livermore, CA, USA) </li></ol></li><li> Laser Doppler flussimetria sistema (ADInstruments, Inc., Colorado Springs, CO, USA) <ol class="lalpha"><li> StMatita sonda Andard </li><li> Porta sonda </li><li> Sangue flussometro </li><li> Powerlab 4/30 con LabChart 7 </li></ol></li><li> Misurazione del volume dell&#39;infarto <ol class="lalpha"><li> Rat cervello matrice (Zivic Miller-Lab., Inc., Allison Park, PA, USA) </li><li> 2,3,5-trifeniltetrazolio cloruro (Sigma-Aldrich, St Louis, MO, USA) diluito a 0,05% in PBS </li><li> Scanner piano (Epson Perfection V30, Epson America, Inc., Long Beach, CA, USA) </li><li> Image J software (ImageJ 1.42q software, US National Institutes of Health, Bethesda, MA, USA) </li></ol></li></ol> 1. Pre-chirurgiche Steps <ol><li> Prima dell&#39;intervento, i topi sono alloggiati sotto una 12:12 ciclo luce / buio con libero accesso all&#39;acqua e roditori chow. </li><li> Anestesia viene indotta con 4% isoflurano in 100% O 2 miscela di gas in una camera di induzione fino ratto non ritiri a pinch zampa posteriore. </li><li> Latecnica settico dovrebbe essere mantenuta durante questa procedura, compreso l&#39;uso di guanti sterili, sterili strumenti chirurgici, e un telo chirurgico sterile 11. </li><li> La parte superiore della testa è rasata con tosatrici elettriche. </li><li> Il ratto viene posto in posizione prona su un tampone assorbente sdraiata su una temperatura controllata superficie operativa (coperta termica). </li><li> La testa viene posta nell&#39;apparecchio stereotassico partire posizionamento della maschera frontale del gas anestetico. </li><li> Successivamente, le barre auricolari sono inseriti e serrati. </li><li> Durante la procedura di anestesia viene mantenuta con isofluorano 2% in 100% O 2 miscela di gas. </li><li> Unguento oftalmico lubrificante viene applicato a entrambi gli occhi, e le palpebre sono chiuse per prevenire l&#39;essiccamento occhio durante la procedura chirurgica. </li><li> Una sonda di temperatura rettale è inserito per mantenere una temperatura interna costante di 37 animali ± 0,5 ° C. </li><li> Con la testa saldamente nella stereotassicodispositivo, l&#39;area chirurgica è purificato con alternanza di clorexidina al 2% e soluzione fisiologica per tre volte. </li></ol> 2. Fasi chirurgiche <ol><li> Utilizzando un bisturi, una incisione mediana è fatta sulla pelle sovrastante la calvarium dagli aspetti più caudali degli occhi (nasion) fra le orecchie (linea nucale superiore). </li><li> La pelle viene quindi retratto lateralmente con 3 pinze bulldog. </li><li> Tessuto connettivo è rimosso dal cranio con tamponi di cotone asciutti modo che molteplici strutture può essere visto chiaramente. Questi includono bregma, la sutura coronale, e la cresta a destra laterale del cranio. Tamponi di cotone sono usati per rimuovere il sangue dal campo operatorio. </li><li> Utilizzando il microscopio operatorio, bregma si trova, ed i manipolatori stereotassiche vengono regolati fino alla rotonda 1,0 millimetri fresa trapano viene azzerato a bregma. </li><li> La fresa trapano è poi spostato a 1,6 mm e 5,2 mm anteriore laterale bregma. </li><li> Un foro fresa che penetra il cranio si è esercitato per cannula posizionamento (Figura 1). Detriti in eccesso e il sangue sono continuamente spengono con tamponi di cotone. </li></ol> A questo punto, una cannula guida può essere inserito (passo 7) o diretta ET-1 attraverso il foro di iniezione può essere eseguita fresa (procedere direttamente al punto 16). <ol start="7"><li> Successivamente, fori fresa per 3 viti di montaggio si sono esercitati nello spessore parziale del cranio con un 1,0 mm di cono rovesciato trapano fresa (Figura 1). Un buco si è esercitato in ogni osso frontale di circa 1-2 mm anteriormente alla sutura coronale e 1-2 mm lateralmente alla sutura sagittale. Un buco si è esercitato nell&#39;osso parietale circa 2-3 mm posteriormente alla sutura coronale e 2-3 mm lateralmente alla sutura sagittale ipsilaterale al foro di guida cannula fresa. Tre 0-80 x 3/32 viti di montaggio sono posti in questi buchi fresa e fornirà il supporto per il cemento che tiene nella cannula. Le viti devono essere solo advanced 2 o 3 giri in modo da non danneggiare la dura madre. </li&gt; <li> La cannula guida è inserito nel supporto stereotassica cannula e bregma si trova. </li><li> I manipolatori stereotassiche vengono regolati fino a quando la cannula guida è azzerato a bregma. La cannula viene spostato nel foro situato fresa anteriore 1,6 millimetri e 5,2 mm laterale bregma. </li><li> Infine, la cannula guida è abbassato nel foro della fresa con la posizione della punta finale di 4,5 mm al bregma ventrale. </li><li> Laser Doppler sonda collocamento di flusso (opzionale) <ol class="lalpha"><li> Al fine di monitorare il flusso ematico cerebrale utilizzando LDF, un porta sonda può essere collocato in posizione prima apposizione della cannula guida con cemento dentale. </li><li> La base porta sonda viene tagliata a filo con il piedistallo, tranne per la scheda a forma di piccolo cuneo. </li><li> Il supporto sonda viene poi collocato posteriormente alla cannula guida e appena mediale alla cresta laterale del cranio con la linguetta orientata medialmente (Figura 1). </li><li> Il titolare sonda e gUIDA cannula sono apposti insieme con cemento dentale. </li></ol></li><li> Cemento dentale viene quindi utilizzato per fissare la cannula in posizione. Il cemento è in contatto con tutte e tre le viti e circonda l&#39;intera base della cannula. </li><li> Il cemento dovrebbe essere completamente asciutto prima della rimozione del supporto cannula. Questa operazione richiede circa 5 minuti. </li></ol> Dopo queste operazioni, l&#39;incisione chirurgica può essere chiuso e il manichino cannula può essere avvitato nella cannula guida. In alternativa, ET-1 indotta MCAO può essere eseguita sul ratto dopo un periodo di recupero dalla chirurgia impianto cannula. Per questo metodo, passo 19 deve essere eseguita dopo e passi 14-18 possono essere eseguite in un secondo momento. Effettuare guida impianto cannula e ET-1 iniezione durante l&#39;intervento stesso, passo 14 deve essere eseguita dopo. <ol start="14"><li> La siringa di infusione è caricato con ET-1 (diluito a 80 pM in PBS) e poi montate nel iniettore stereotassico. </li><li> Ilmanipolatori stereotassiche vengono regolati finché la punta dell&#39;ago viene azzerato al bordo della cannula guida. </li><li> La punta dell&#39;ago è abbassato attraverso la cannula guida in una posizione di 17,2 millimetri ventrale al bordo della cannula guida. Se una cannula guida non viene utilizzato, la punta dell&#39;ago viene azzerato al bregma e abbassato attraverso il foro di una fresa 8,7 millimetri posizione ventrale al bregma. </li><li> 3 ml di 80 mM ET1 viene infusa a una velocità di 1 ml al minuto. </li><li> La siringa viene lasciato in posizione per 3 minuti dopo l&#39;infusione è completa e quindi lentamente rimosso. </li><li> L&#39;incisione viene chiusa con sutura di nylon 3,0 e il manichino cannula viene avvitata nella cannula. </li><li> Una dose di analgesico appropriato (ad esempio buprenorfina a 0,05-0,1 mg / kg) deve essere utilizzato dopo l&#39;intervento chirurgico per ridurre al minimo il dolore e il disagio durante il periodo di recupero. </li><li> Il ratto viene rimosso dalla sala operatoria e collocato in un ambiente caldo, zona di recupero asciutto per prevenire l&#39;ipotermia, con connessione, facile accesso a cibo e acqua dolce. </li&gt; </ol>

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L&#39;ET-1 indotta MCAO è un modello sperimentale consolidata di ictus ischemico che viene regolarmente utilizzato in molteplici ceppi di ratto. Molte variabili, come ceppo ratto, animale età, temperatura corporea, metodo di anestesia, e competenza operatore può portare ad un aumento dei volumi di variabilità infarto quando si utilizza questo modello 5, 14. Tuttavia, alcuni ricercatori hanno dimostrato che i vantaggi di questo modello sono l&#39;approccio relativamente non-invasiva, dose-risposta del flu...

<table border="1"><tbody><tr><td></td><td></td><td></td><td><ol><li> Animali: otto settimane di età, di sesso maschile, Sprague Dawley (Charles River, Farms Wilmington, MA, USA) del peso di 250-300 g al momento dell&#39;intervento chirurgico. </li><li> Anestesia <ol class="lalpha"><li> Inalazione sistema di anestesia (VetEquip Inc., Pleasanton, CA, USA) </li><li> Isoflurano anestetico (Farmaceutici Baxter, Deerfield, IL, USA) </li></ol></li><li> Stereotassico sistema (David Kopf Instruments, Tujunga, CA, USA) <ol class="lalpha"><li> Piccolo animale sistema stereotassico </li><li> Non rottura orecchio barre per i ratti </li><li> Gas testa porta l&#39;anestesia per i ratti </li></ol></li><li> Regolazione della temperatura <ol class="lalpha"><li> BAT-12 Termometro microsonda (World Precision Instruments, Inc., Sarasota, FL, Stati Uniti d&#39;America) </li><li> T / POMPA, TP600 coperta termica (Gaymar Industries, Inc., Orchard Park, NY, Stati Uniti d&#39;America) </li></ol></li><li> Strumenti chirurgici <ol class="lalpha"><li> Manico bisturi e # 11 della lama, iris forcipe, Graefe pinze, divaricatori bulldog pinza, cacciavite, siringa da 10 ml con 26 gauge smussato (Strumenti di precisione del Mondo, Inc., Sarasota, FL, Stati Uniti d&#39;America) </li><li> Micromotore trapano e supporto stereotassica, Quintessential stereotassico Injector (Stoelting, Wood Dale, IL, USA) </li><li> 1,0 millimetri fresa trapano rotondo, 1,0 mm di cono rovesciato trapano fresa (Roboz chirurgico Instrument Co., Inc., Gaithersburg, MD, USA) </li></ol></li><li> Forniture chirurgiche <ol class="lalpha"><li> Viti di montaggio 0-80 x 3/32 con 2.4 mm di lunghezza dell&#39;albero, da 21 gauge cannula guida [4 millimetri a lungo sotto il piedistallo] e manichino cannula (Materie plastiche uno, Roanoke, VA, USA) </li><li> Jet acrilico protesi e liquidi (Lang Dental Manufacturing Co., Inc., Wheeling, IL, USA) </li><li> 3,0 nylon sutura (Oasis, Mettawa, IL, USA) </li><li> Tamponi di cotone, l&#39;unguento oculare Puralube (Fisher scientific, Pittsburg, PA, USA) </li><li> Tosatrici elettriche (Oster, Providence, RI, USA) </li></ol></li><li> Prodotti chimici <ol class="lalpha"><li> L&#39;endotelina-1 (American Peptide, Sunnyvale, CA, USA) </li><li> Clorexidina al 2% (Agrilabs, St. Joseph, MO, USA) </li><li> Buprenorfina HCl (Hospira Inc., Lake Forest, IL, USA) </li></ol></li><li> Visualizzazione Attrezzature <ol class="lalpha"><li> Microscopio chirurgico (Strumento Seiler e Manufacturing, St. Louis, MO, USA) </li><li> Fibra ottica illuminatore (TechniQuip Corp., Livermore, CA, USA) </li></ol></li><li> Laser Doppler flussimetria sistema (ADInstruments, Inc., Colorado Springs, CO, USA) <ol class="lalpha"><li> Matita sonda standard </li><li> Porta sonda </li><li> Sangue flussometro </li><li> Powerlab 4/30 con LabChart 7 </li></ol></li><li> Misurazione del volume dell&#39;infarto <ol class="lalpha"><li> Rat cervellomatrice (Zivic Miller-Lab., Inc., Allison Park, PA, USA) </li><li> 2,3,5-trifeniltetrazolio cloruro (Sigma-Aldrich, St Louis, MO, USA) diluito a 0,05% in PBS </li><li> Scanner piano (Epson Perfection V30, Epson America, Inc., Long Beach, CA, USA) </li><li> Image J software (ImageJ 1.42q software, US National Institutes of Health, Bethesda, MA, USA) </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

L&#39;ictus è la prima causa di disabilità e la terza causa principale di morte nel mondo, che costano una cifra stimata di 70 miliardi negli Stati Uniti nel 2009 1, 2. Diversi modelli di ischemia cerebrale sono stati sviluppati per simulare la condizione umana di ictus. È stato suggerito che fino al 80% di tutte le corse da risultato nel danno ischemico cerebrale media (MCA) area 3. Nei primi anni 1990, endotelina-1 (ET-1) 4 è stato utilizzato per indurre ischemia applicandola direttamente adiacente alla superficie del MCA dopo craniotomia. Successivamente, questo modello è stato modificato 5 utilizzando una iniezione stereotassica di ET-1 adiacente alla MCA produrre ischemia cerebrale focale. I vantaggi principali di questo modello includono la capacità di eseguire la procedura rapida, la capacità di controllare costrizione dell&#39;arteria alterando la dose di ET-1 espresso, nessuna necessità di manipolare i vasi extracranici forniscono sangue al cervello e graduale reperfusisui tassi che più strettamente imita la riperfusione nell&#39;uomo 5-7. D&#39;altra parte, l&#39;ET-1 modello ha svantaggi che includono la necessità di una craniotomia, così come una maggiore variabilità della gittata sistolica 8. Tale variabilità può essere ridotta con l&#39;uso di laser Doppler flussimetria (LDF) per verificare ischemia cerebrale durante ET-1 infusione. I fattori che influenzano la variabilità ictus comprendono la precisione di infusione e il lotto del ET-1 utilizzato 6. Un&#39;altra considerazione importante è che, anche se la riperfusione è un evento comune nel tratto umano, la durata di occlusione per ET-1 indotta MCAO non strettamente imitare quello di ictus umano in cui molti pazienti hanno riperfusione parziale per un periodo di ore o giorni dopo occlusione 9, 10. Questo protocollo descrive in dettaglio l&#39;ET-1 indotta modello MCAO per l&#39;ictus ischemico nei ratti. Sarà inoltre richiamare l&#39;attenzione su considerazioni particolari e gli svantaggi potenziali durante tutta la procedura.

1. Post-Op valutazione neurologica Dopo che l&#39;animale riprenda coscienza, una grande varietà di test possono essere utilizzati per valutare i deficit neurologici, tra cui l&#39;equilibrio, forza di presa, messa zampa, asimmetria posturale e scala arrampicata. Il compito di semi di girasole è una valutazione lorda di funzione motoria e sensoriale che ha una correlazione significativa con il volume dell&#39;infarto 7, 12. Durante questa attività, i ratti sono a tempo durante l&#...

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

Modelli animali di ischemia cerebrale sono stati sviluppati per simulare la condizione umana di ictus. Questo protocollo descrive l&#39;endotelina-1 (ET-1) indotta da occlusione dell&#39;arteria cerebrale media (MCAO) modello per l&#39;ictus ischemico nei ratti. Inoltre, importanti considerazioni, i vantaggi ei limiti di tale modello sono discussi.

L&#39;endotelina-1 indotta cerebrale media occlusione dell&#39;arteria modello per l&#39;ictus ischemico con guida laser flussimetria Doppler 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 ...

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19.8K Views.  University of Florida. Modelli animali di ischemia cerebrale sono stati sviluppati per simulare la condizione umana di ictus. Questo protocollo descrive l'endotelina-1 (ET-1) indotta da occlusione dell'arteria cerebrale media (MCAO) modello per l'ictus ischemico nei ratti. Inoltre, importanti considerazioni, i vantaggi ei limiti di tale modello sono discussi.

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.

Corsa di modellazione in Mouse - Medio occlusione dell&#39;arteria cerebrale con il Modello Filament

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.

Modello murino di Mezzo occlusione dell&#39;arteria cerebrale

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.

Intraluminali occlusione dell&#39;arteria cerebrale media (MCAO) Modello per l&#39;ictus ischemico con la guida flussimetria laser Doppler in topi

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.

Bilaterale occlusione comune Arteria carotide come uno stimolo precondizionamento in grado di indurre tolleranza ischemica precoce di focale Ischemia Cerebrale

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.

Utilizzando una finestra craniale per visualizzare l&#39;arteria cerebrale media Durante endotelina-1 indotta da occlusione dell&#39;arteria cerebrale media

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.

Sistema ottimizzato per il monitoraggio perfusione cerebrale nel modello Rat Stroke of intraluminale occlusione dell&#39;arteria cerebrale media

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.

Corsa di modellazione in Mouse: Coagulazione permanente dell&#39;arteria cerebrale media distale

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.

Embolic occlusione dell&#39;arteria cerebrale media (MCAO) per l&#39;ictus ischemico con omologhi coaguli di sangue nei ratti

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.

Approccio chirurgico per occlusione dell&#39;arteria cerebrale media e riperfusione indotta ictus nei topi

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.

Transient Medio occlusione dell&#39;arteria cerebrale Modello di Stroke neonatale nei ratti P10

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