Die aktuelle Forschung wird von der Solorz-Zak-Forschungsstiftung gefördert.

Alle chirurgischen Verfahren wurden die ethischen Überprüfung unterzogen und von der Regierung von Oberbayern (Referenznummer: 55.2-1-54-2532.3-13-13 und -2532-136-11) zugelassen. Tiere sind männliche C57BL / 6 Mäuse mit einem Körpergewicht von etwa 25 g auf. 1. Vorbereitung der Tiere <ol><li> Induzieren Anästhesie, indem Sie die Maus in eine Kammer. Spülen Sie die Kammer mit 5% Isofluran, bis das Tier das Bewusstsein verliert. </li><li> Injizieren vorgemischten Narkose intraperitoneal: Fentanyl (0,05 mg / kg), Midazolam (5 mg / kg) und Medetomidin (0,5 mg / kg). Überprüfen Reflexe vor und während des Verfahrens regelmäßig. Reinjizierst ein Drittel der ursprünglichen Menge Stunden Anästhesie erhalten. </li><li> Intubieren orotracheal mit einem Rohr aus einem Katheter 20 G 11 vorgenommen. Für Intubation beheben Sie das Tier auf einer schrägen Plattform (30 °), die Zunge zurückziehen mit gebogenen Pinzette, visualisieren die Stimmbänder unter einem Operationsmikroskop und stecken Sie den Schlauch in die Luftröhrewährend der Inspiration. </li><li> Platzieren Sie die Maus in Bauchlage und die korrekte Platzierung der Röhrchen mit einem Stück Baumwolle oder einem microcapnograph. </li><li> Schließen Sie das Intubationstubus an die Atemschutzmaske. Lüften Sie die Maus mit Raumluft mit 25% Sauerstoff, ergänzt mit einer Frequenz von 180 bis 220 Atemzüge / min und einem Schlagvolumen von 200-250 ul. </li><li> Schließen Sie das Intubationstubus auf die microcapnograph. Pflegen Sie die endexspiratorischen pCO 2 bei 30 mmHg durch Anpassung der Beatmungsfrequenz. </li><li> Legen Sie eine rektale Temperatursonde und legen Sie das Tier auf einem Heizkissen, um die 37 ° C Körperkerntemperatur aufrecht zu erhalten. </li><li> Tragen Sie die Ring Pulsoximeter Sensor an der rechten Hinterpfote. </li><li> Öffnen Sie die Haut über dem Schädel mit einer Schere. Der Schnitt sollte etwa 0,5 cm lang und zwischen Ohr und Auge. </li><li> Präparieren Sie die linke Schläfenmuskel mit einem Skalpell aus dem Schläfenbein. </li><li> Kleben Sie die Laser-Doppler-Durchflussmesser (LDF) Sonde aufder linke Schläfenbein. Halten der Sonde in einer festen Position, bis der Kleber aushärtet. </li><li> Bohren Sie ein Loch von ca. 1,5 mm Durchmesser mit einem Zahnbohrer in den linken Schläfenbein. Kühlen Sie die Knochen mit Kochsalzlösung, um Hitzeschäden zu vermeiden. </li><li> Legen Sie die ICP-Sonde in die Schädelhöhle. Schieben nach vorn dorsal wie möglich zu Hirngewebeschäden und Blutungen zu verhindern. </li><li> Wenn die Sonde an der richtigen Position, zu beheben, und verschließen Sie diese mit Zement. Lassen Sie den Zement trocken für 5 min. </li><li> Drehen Sie die Maus vorsichtig zu einer Rückenlage. </li><li> Für kontinuierliche Blutdrucküberwachung, Katheterisierung der linken Oberschenkelarterie. </li><li> Verbinden Sie den Katheter mit dem Oberschenkelblutdrucküberwachungsgerät. </li></ol> 2. SAH Induktions <ol><li> Öffnen Sie die Haut mit einer Schere vom Brustbein bis zum Kinn (2 cm). Präparieren Sie die Bindegewebe unverblümt und drücken Sie die Speicheldrüsen beiseite. </li><li> Setzen Sie die linke A. carotis communis (CCA) und mobilisieren sie. Preserve dieVagusnerv, die in der gleichen Bindegewebe als CCA läuft. Bewegen Schädel und entlarven und zu mobilisieren, die ICA und den Rechnungshof mit der gleichen Technik. </li><li> Ligieren des ECA so weit kranial wie möglich. </li><li> Vereinbaren zwei weitere Ligationen für das Filament auf der ECA. </li><li> Verschließen Sie die CCA und der ICA vorübergehend mit microclips. Positionieren Sie die microclips mit einem Mikroclip Applikator. Stellen Sie sicher, die Clips richtig durch leichtes Ziehen sie wieder angewendet. </li><li> Schneiden Sie ein Loch für das Filament Einsetzen in den ECA mit einem Schiff Schere. </li><li> Legen Sie eine Prolene 5-0 Faden mit 12 mm Länge in den ECA. </li><li> Schließen Sie die Einstichstelle mit einem verabredeten Ligation. </li><li> Entfernen Sie die microclips mit einem Mikroclip-Applikator von der CCA und der ICA. </li><li> Schieben Sie den Faden mit einer Pinzette in die ICA, bis der ICP steigt. Ein plötzlicher Anstieg des ICP zeigt Blutungen Induktion. </li><li> Ziehen Sie den Faden sofort abzubinden und die ECA durch Schließen sowohl prearranged Liga nacheinander. Dies verhindert ein Ausbluten der Insertionsstelle. </li><li> Naht der Hautwunde. </li><li> Überwachen Sie die physiologischen Parameter des Tieres für weitere 20 min. </li><li> ICP entfernen und LDF-Sonden und Naht der Hautwunde. </li></ol> 3. Ende des Experiments <ol><li> Perfusion des Tieres transkardial mit 20 ml Kochsalzlösung (Raumtemperatur), gefolgt von 20 ml von 4% PFA in PBS (4 ° C). </li><li> Präparieren Sie das Gehirn aus dem Schädel. Schneiden Sie die Schädel in der Mittellinie und zwischen den Augenhöhlen. Dann ziehen die Knochen aus dem Gehirn. </li><li> Bewerten Sie die Blutverteilung im Subarachnoidalraum. </li></ol> 4. Überlegungen in der Sache des Überlebens Chirurgie (nicht im Video gezeigt) <ol><li> Inject Carprofen (4 mg / kg subkutan) für die postoperative Analgesie direkt nach Narkoseeinleitung. Während der postoperativen Beobachtungszeitraum Carprofen (4 mg / kg subkutan) wird alle 24 Std. </li&gt; <li> Statt invasive Blutdruckmessung nutzen eine nicht-invasive Blutdrucküberwachungssystem. </li><li> Zur Beendigung der Narkose inject Antagonisierung Mittel subkutan Naloxon (1,2 mg / kg), Flumazenil (0,5 mg / kg) und Atipamezol (2,5 mg / kg). </li><li> Extubiert das Tier. </li><li> Nach Wiedererlangung der Reflexe legte das Tier in einen vorgeheizten Kammer. Halten des Tieres bei etwa 32 ° C für 24 Stunden. </li><li> Tiere werden regelmäßig auf Spontanatmung und deren Allgemeinzustand in den ersten Stunden nach der Operation überprüft. Wenn der Hirnstamm betroffenen Tiere haben Probleme mit der Atmung und sollte eingeschläfert werden. </li><li> Die Tiere werden täglich für ihre Allgemeinzustand und das Körpergewicht als auch für neurologische und sensorische Defizite 15 überprüft. </li></ol>

<ol>
	<li>Cahill, J., Zhang, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Subarachnoid+hemorrhage:+is+it+time+for+a+new+direction.">Subarachnoid hemorrhage: is it time for a new direction.</a> Stroke. 40, 86-87 (2009).</li><li>van Gijn, J., Kerr, R. S., Rinkel, G. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Subarachnoid+haemorrhage.">Subarachnoid haemorrhage.</a> Lancet. 369, 306-318 (2007).</li><li>Lin, C. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+murine+model+of+subarachnoid+hemorrhage-induced+cerebral+vasospasm.">A murine model of subarachnoid hemorrhage-induced cerebral vasospasm.</a> J. Neurosci. Methods. 123, 89-97 (2003).</li><li>Altay, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+method+for+subarachnoid+hemorrhage+to+induce+vasospasm+in+mice.">A novel method for subarachnoid hemorrhage to induce vasospasm in mice.</a> J. Neurosci. Methods. 183, 136-140 (2009).</li><li>Feiler, S., Friedrich, B., Scholler, K., Thal, S. C., Plesnila, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Standardized+induction+of+subarachnoid+hemorrhage+in+mice+by+intracranial+pressure+monitoring.">Standardized induction of subarachnoid hemorrhage in mice by intracranial pressure monitoring.</a> J. Neurosci. Methods. 190, 164-170 (2010).</li><li>Bederson, J. B., Germano, I. M., Guarino, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cortical+blood+flow+and+cerebral+perfusion+pressure+in+a+new+noncraniotomy+model+of+subarachnoid+hemorrhage+in+the+rat.">Cortical blood flow and cerebral perfusion pressure in a new noncraniotomy model of subarachnoid hemorrhage in the rat.</a> Stroke. 26, 1086-1091 (1995).</li><li>Veelken, J. A., Laing, R. J., Jakubowski, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Sheffield+model+of+subarachnoid+hemorrhage+in+rats.">The Sheffield model of subarachnoid hemorrhage in rats.</a> Stroke. 26, 1279-1283 (1995).</li><li>Kamii, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Amelioration+of+vasospasm+after+subarachnoid+hemorrhage+in+transgenic+mice+overexpressing+CuZn-superoxide+dismutase.">Amelioration of vasospasm after subarachnoid hemorrhage in transgenic mice overexpressing CuZn-superoxide dismutase.</a> Stroke. 30, 867-871 (1999).</li><li>Parra, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+model+of+subarachnoid+hemorrhage+associated+cerebral+vasospasm:+methodological+analysis.">Mouse model of subarachnoid hemorrhage associated cerebral vasospasm: methodological analysis.</a> Neurol. Res. 24, 510-516 (2002).</li><li>Broderick, J. P., Brott, T. G., Duldner, J. E., Tomsick, T., Leach, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Initial+and+recurrent+bleeding+are+the+major+causes+of+death+following+subarachnoid+hemorrhage.">Initial and recurrent bleeding are the major causes of death following subarachnoid hemorrhage.</a> Stroke. 25, 1342-1347 (1994).</li><li>Thal, S. C., Plesnila, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Non-invasive+intraoperative+monitoring+of+blood+pressure+and+arterial+pCO2+during+surgical+anesthesia+in+mice.">Non-invasive intraoperative monitoring of blood pressure and arterial pCO2 during surgical anesthesia in mice.</a> J. Neurosci. Methods. 159, 261-267 (2007).</li><li>Hockel, K., Trabold, R., Scholler, K., Torok, E., Plesnila, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impact+of+anesthesia+on+pathophysiology+and+mortality+following+subarachnoid+hemorrhage+in+rats.">Impact of anesthesia on pathophysiology and mortality following subarachnoid hemorrhage in rats.</a> Exp. Transl. Stroke Med. 4, 5 (2012).</li><li>Wang, Z., Schuler, B., Vogel, O., Arras, M., Vogel, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=What+is+the+optimal+anesthetic+protocol+for+measurements+of+cerebral+autoregulation+in+spontaneously+breathing+mice?.">What is the optimal anesthetic protocol for measurements of cerebral autoregulation in spontaneously breathing mice?.</a> Exp. Brain Res. 207, 249-258 (2010).</li><li>Schwartz, A. Y., Masago, A., Sehba, F. A., 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=Experimental+models+of+subarachnoid+hemorrhage+in+the+rat:+a+refinement+of+the+endovascular+filament+model.">Experimental models of subarachnoid hemorrhage in the rat: a refinement of the endovascular filament model.</a> J. Neurosci. Methods. 96, 161-167 (2000).</li><li>Feiler, S., Plesnila, N., Thal, S. C., Zausinger, S., Scholler, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contribution+of+matrix+metalloproteinase-9+to+cerebral+edema+and+functional+outcome+following+experimental+subarachnoid+hemorrhage.">Contribution of matrix metalloproteinase-9 to cerebral edema and functional outcome following experimental subarachnoid hemorrhage.</a> Cerebrovasc. Dis. 32, 289-295 (2011).</li></ol>

Autoren haben nichts zu offenbaren.

Behandlungsmöglichkeiten nach SAB sind knapp und meist unwirksam. Deshalb ist die Pathophysiologie der post hämorrhagischen Hirnschäden muss, um neue therapeutische Ziele zu identifizieren und zu entwickeln neue therapeutische Ansätze weiter zu verstehen. Standardisierte und reproduzierbare Tiermodellen auch in gentechnisch veränderten Tieren, also Mäuse, sind entscheidend für solche Untersuchungen. Der CWP-Modell hat sich ein weit verbreitetes Modell für SAH, wie es ähnelt der Pathophysiologie des Men...

SAH ist die Schlaganfall-Subtyp mit der geringsten positiven Auswirkungen für den Patienten: 40% der Patienten sterben innerhalb eines Monats nach der Blutung ein und Überlebende haben nur selten eine klinisch positiven Ergebnis. Die große Mehrheit der spontanen sahs (80%) werden durch Ruptur der intrakraniellen Aneurysmen, die meist entlang der vorderen und hinteren Kommunikation Arterie, die A. basilaris und A. cerebri media (MCA) 2 befinden verursacht. Solche Aneurysmen sind schwierig in Tieren zu modellieren und damit Tiermodellen von SAH werden entweder durch Injektion von Blut in den Subarachnoidalraum / Hirnkammern oder durch endovaskuläre Perforation einer subarachnoidalen Gefäß durchgeführt. Eigenblutinjektion in die Cisterna magna ist einfach durchzuführen und reproduzierbarer als das Blutvolumen kann direkt gesteuert werden 3. Leider sind einige Aspekte der Pathophysiologie SAH, z. B. dieGefäßverletzung, können mit diesem Verfahren nicht modelliert werden. Ein anderer technischer Ansatz für die Induktion von SAH ist die Eröffnung einer Vene intracisternal 4. , Die intraluminale CWP an der MCA Zweig scheint jedoch das Verfahren, dass die Modelle die Pathophysiologie des Menschen am engsten 5 sein. Die Methode wurde von Bederson und Kollegen und gleichzeitig durch Veelken 6,7 und Kollegen entwickelt und zuerst in Ratten beschrieben. Später wurde die intraluminale Perforation Modell mit Mäusen 8,9 angepasst. Ein Faden wird in der A. carotis externa (ECA) eingeführt und zur Schädelbasis über die Arteria carotis interna (ICA). An der Verzweigungsstelle des MCA perforiert das Filament das Schiff und induziert eine Blutung in den Subarachnoidalraum an der Schädelbasis. Das Blut verteilt dann in den verbleibenden Subarachnoidalraum entlang Spalten und Blutgefäße. Blutungen durch Gerinnselbildung an der Stelle der Perforation aufgehört, aber rebleedings, whICH sind häufig bei Patienten 10, auftreten kann schädlich. Entsprechend wurde die endovaskuläre Filament Modell ein weit verbreitetes Modell SAH in den letzten Jahren. Der am häufigsten genannte Nachteil des Filaments Perforation Modell ist, dass Blutungen Volumen kann nicht direkt gesteuert werden, und können daher variabel sein. Diese Variabilität kann durch strenge Kontrolle der Tierphysiologie und Post-hämorrhagischen ICP deutlich reduziert werden. Mäuse haben den großen Vorteil, dass eine große Anzahl von genetisch veränderten Stämme sind verfügbar. Aufgrund ihrer geringen Größe chirurgischen Verfahren neigen jedoch dazu, komplexer zu sein als in größeren Spezies, zB Ratten oder Kaninchen. Deshalb ist die Verkleinerung von Techniken für die Ratten, Mäuse entwickelten oft nicht zu den gewünschten Ergebnissen, zB als Mäuse haben eine sehr begrenzte Körpergewicht und Blutvolumen nicht-invasive Techniken zur Blutdruck-und Blutgasanalyse sowie für die Hämoglobinsättigung und der Herzfrequenz führenmüssen angewendet werden, wann immer möglich. Dementsprechend ist das Ziel der aktuellen Publikation, um den Faden Perforation Modell für SAH in Mäusen beschreiben und zu zeigen, wie dieses Modell in einer standardisierten und hoch reproduzierbare Art und Weise durchgeführt werden.

<table border="1">
	<tbody>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td>Equipment</td>
		</tr>
		<tr>
			<td>operation microscope</td>
			<td>Leica</td>
			<td>KL2500</td>
			<td></td>
		</tr>
		<tr>
			<td>isoflurane vaporizer</td>
			<td>Harvard Instruments</td>
			<td>Continuous Flow Vaporizer</td>
			<td></td>
		</tr>
		<tr>
			<td>respirator</td>
			<td>Hugo Sachs</td>
			<td>Minivent 845</td>
			<td></td>
		</tr>
		<tr>
			<td>microcapnograph</td>
			<td>Hugo Sachs</td>
			<td>Type 340</td>
			<td></td>
		</tr>
		<tr>
			<td>temperature controller</td>
			<td>FHC</td>
			<td>DC Temperature Controller</td>
			<td></td>
		</tr>
		<tr>
			<td>dental drill</td>
			<td>Paggen</td>
			<td>Labset- N</td>
			<td></td>
		</tr>
		<tr>
			<td>ICP monitor</td>
			<td>Codman</td>
			<td>ICP monitor</td>
			<td></td>
		</tr>
		<tr>
			<td>blood pressure monitor</td>
			<td>AD Instruments</td>
			<td>Bridge Amp FE221</td>
			<td></td>
		</tr>
		<tr>
			<td>syringe pump</td>
			<td>World Precision Instruments</td>
			<td>SP101IZ</td>
			<td></td>
		</tr>
		<tr>
			<td>pulsoximeter</td>
			<td>Kent Scientific</td>
			<td>MouseSTAT</td>
			<td></td>
		</tr>
		<tr>
			<td>LDF</td>
			<td>Perimed</td>
			<td>Periflux 5000</td>
			<td></td>
		</tr>
		<tr>
			<td>analog data monitor</td>
			<td>AD Instruments</td>
			<td>Power Lab 16/35</td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td>Material</td>
		</tr>
		<tr>
			<td>cement for ICP probe fixation</td>
			<td>Speiko</td>
			<td>Carboxylate cement</td>
			<td></td>
		</tr>
		<tr>
			<td>glue for LDF probe fixation</td>
			<td>Bob Smith Industries</td>
			<td>Cyanoacrylate glue (Maxi Cure and Insta Set)</td>
			<td></td>
		</tr>
		<tr>
			<td>venous catheter</td>
			<td>Johnson &#38; Johnson</td>
			<td>Jelco winged i.v. catheter; REF 4076</td>
			<td>modified intubation tube</td>
		</tr>
		<tr>
			<td>tubing for femoral catheter</td>
			<td>Smiths Medical</td>
			<td>Fine Bore Polythene Tubing; ID 0.28 mm OD 0.61 mm; REF 800/100/100</td>
			<td>cut to 30 cm length</td>
		</tr>
		<tr>
			<td>filament for vessel perforation</td>
			<td>Ethicon</td>
			<td>Prolene 5-0</td>
			<td>cut to 12 mm length</td>
		</tr>
		<tr>
			<td>surgical equipment</td>
			<td>Fine Scientific Instruments</td>
			<td>forceps medical #5, vessel scissors 8 cm, microclip 4 mm jaw</td>
			<td></td>
		</tr>
	</tbody>
</table>

a murine model of subarachnoid hemorrhage

In diesem Video-Veröffentlichung eine standardisierte Maus-Modell der Subarachnoidalblutung (SAB) wird vorgestellt. Blutung wird durch endovaskuläre Kreis von Willis Perforation (CWP) induziert und durch Hirndruck (ICP) Überwachung bewährt. Dadurch wird eine homogene Verteilung im Blut Liquorräume um den arteriellen Kreislauf-und Kleinhirnrisse erreicht. Körpertemperatur, den systemischen Blutdruck, Herzfrequenz, und Hämoglobinsättigung: Tierphysiologie wird durch Intubation, Beatmung und kontinuierliche Online-Überwachung von verschiedenen physiologischen und Herz-Kreislauf-Parameter gepflegt. Dadurch wird die Hirndurchblutung Druck kann genau überwacht werden, was zu einem weniger variablen Volumen von ausgetretenen Blutes. Dies ermöglicht eine bessere Standardisierung der endovaskulären Filament Perforation in Mäusen und macht das ganze Modell sehr gut reproduzierbar. So ist es für pharmakologische und pathophysiologische Studien in Wildtyp-und genetisch leicht verfügbarly veränderten Mäusen.

Sterblichkeit Sobald die Operationstechnik beherrscht das Verfahren nicht zu entlocken jede intraoperative Mortalität. Auch Blutungen in nahezu allen Tieren erreicht werden. Die postoperative Mortalität ist mit 30-40% die meisten Tiere sterben am Tag 1 nach der Operation (Abbildung 5). ICP-Werte nach SAB Die ICP-Blutungen vor etwa 4 mmHg. Blutende Ergebnisse in einem starken Anstieg des ICP bis zu 120 mmHg. ICP-Werte dann...

Watch this Scientific Journal Video about A Murine Model of Subarachnoid Hemorrhage at JoVE.com

A Murine Model of Subarachnoid Hemorrhage

Eine standardisierte Maus-Modell der Subarachnoidalblutung durch intraluminale Kreis von Willis Perforation beschrieben. Schiff Perforation und Subarachnoidalblutung durch intrakranielle Drucküberwachung überwacht. Neben verschiedenen Vitalparameter erfasst und gesteuert werden, um physiologische Bedingungen aufrecht zu erhalten.

Einem Mausmodell der Subarachnoidalblutung

In this video publication a standardized mouse model of subarachnoid  hemorrhage (SAH) is presented. Bleeding is induced by endovascular Circle of  Willis ...

a-murine-model-of-subarachnoid-hemorrhage

Research

JoVE Journal

Medicine

19.5K Aufrufe.  University of Munich Medical Center.  Eine standardisierte Maus-Modell der Subarachnoidalblutung durch intraluminale Kreis von Willis Perforation beschrieben. Schiff Perforation und Subarachnoidalblutung durch intrakranielle Drucküberwachung überwacht. Neben verschiedenen Vitalparameter erfasst und gesteuert werden, um physiologische Bedingungen aufrecht zu erhalten. 

Serie: A Murine Model of Subarachnoid Hemorrhage

A Low Mortality Rat Model to Assess Delayed Cerebral Vasospasm After Experimental Subarachnoid Hemorrhage

Objective: To characterize and establish a reproducible model that demonstrates delayed cerebral vasospasm after aneurysmal subarachnoid hemorrhage (SAH) in rats, in order to identify the initiating events, pathophysiological changes and potential targets for treatment.
Methods: Twenty-eight male Sprague-Dawley rats (250 - 300 g) were arbitrarily assigned to one of two groups - SAH or saline control. Rat subarachnoid hemorrhage in the SAH group (n=15) was induced by double injection of autologous blood, 48 hr apart, into the cisterna magna. Similarly, normal saline (n=13) was injected into the cisterna magna of the saline control group. Rats were sacrificed on day five after the second blood injection and the brains were preserved for histological analysis. The degree of vasospasm was measured using sections of the basilar artery, by measuring the internal luminal cross sectional area using NIH Image-J software. The significance was tested using Tukey/Kramer's statistical analysis.
Results: After analysis of histological sections, basilar artery luminal cross sectional area were smaller in the SAH than in the saline group, consistent with cerebral vasospasm in the former group. In the SAH group, basilar artery internal area (.056 &mu;m &plusmn; 3) were significantly smaller from vasospasm five days after the second blood injection (seven days after the initial blood injection), compared to the saline control group with internal area (.069 &plusmn; 3; p=0.004). There were no mortalities from cerebral vasospasm.
Conclusion: The rat double SAH model induces a mild, survivable, basilar artery vasospasm that can be used to study the pathophysiological mechanisms of cerebral vasospasm in a small animal model. A low and acceptable mortality rate is a significant criterion to be satisfied for an ideal SAH animal model so that the mechanisms of vasospasm can be elucidated 7, 8. Further modifications of the model can be made to adjust for increased severity of vasospasm and neurological exams.

Aneurysmal subarachnoid hemorrhage (SAH) is bleeding that occurs into the subarachnoid space when an aneurysm ruptures. While the morbidity and mortality from this event has been on a decline due to improved treatment approaches, the risk of vasospasm after subarachnoid hemorrhage continues to be the same as it was several years ago. The importance of establishing a comprehensive and reproducible animal model to identify initiating events of cerebral vasospasm has been the focus of research since the first use of rats in an experimental vasospasm model in 1979 by Barry et al. Early work in rats demonstrated that a single injection of autologous blood into the cisterna magna led to acute (within minutes) but not delayed cerebral vasospasm 3, 6, 14. Here we characterize a low mortality SAH rat model that results in reproducible delayed vasospasm.

Eine niedrige Sterblichkeit Rattenmodell Verzögerte zerebralen Vasospasmus nach experimenteller Subarachnoidalblutung beurteilen

Objective: To characterize and establish a reproducible model that demonstrates delayed cerebral vasospasm after aneurysmal subarachnoid hemorrhage (SAH) ...

Forschung

Medizin

Murine Model of Wound Healing

Wound healing and repair are the most complex biological processes that occur in human life. After injury, multiple biological pathways become activated. Impaired wound healing, which occurs in diabetic patients for example, can lead to severe unfavorable outcomes such as amputation. There is, therefore, an increasing impetus to develop novel agents that promote wound repair. The testing of these has been limited to large animal models such as swine, which are often impractical. Mice represent the ideal preclinical model, as they are economical and amenable to genetic manipulation, which allows for mechanistic investigation. However, wound healing in a mouse is fundamentally different to that of humans as it primarily occurs via contraction. Our murine model overcomes this by incorporating a splint around the wound. By splinting the wound, the repair process is then dependent on epithelialization, cellular proliferation and angiogenesis, which closely mirror the biological processes of human wound healing. Whilst requiring consistency and care, this murine model does not involve complicated surgical techniques and allows for the robust testing of promising agents that may, for example, promote angiogenesis or inhibit inflammation. Furthermore, each mouse acts as its own control as two wounds are prepared, enabling the application of both the test compound and the vehicle control on the same animal. In conclusion, we demonstrate a practical, easy-to-learn, and robust model of wound healing, which is comparable to that of humans.

A murine model of cutaneous wound healing that can be used to assess therapeutic compounds in physiological and pathophysiological settings.

Mausmodell der Wundheilung

Wound healing and repair are the most complex biological processes that occur in human life. After injury, multiple biological pathways become activated. ...

A New Murine Model of Endovascular Aortic Aneurysm Repair

Endovascular aneurysm exclusion is a validated technique to prevent aneurysm rupture. Long-term results highlight technique limitations and new aspects of Abdominal aortic aneurysm (AAA) pathophysiology. There is no abdominal aortic aneurysm endograft exclusion model cheap and reproducible, which would allow deep investigations of AAA before and after treatment. We hereby describe how to induce, and then to exclude with a covered coronary stentgraft an abdominal aortic aneurysm in a rat. The well known elastase induced AAA model was first reported in 19901 in a rat, then described in mice2. Elastin degradation leads to dilation of the aorta with inflammatory infiltration of the abdominal wall and intra luminal thrombus, matching with human AAA. Endovascular exclusion with small covered stentgraft is then performed, excluding any interactions between circulating blood and the aneurysm thrombus. Appropriate exclusion and stentgraft patency is confirmed before euthanasia by an angiography thought the left carotid artery. Partial control of elastase diffusion makes aneurysm shape different for each animal. It is difficult to create an aneurysm, which will allow an appropriate length of aorta below the aneurysm for an easy stentgraft introduction, and with adequate proximal and distal neck to prevent endoleaks. Lots of failure can result to stentgraft introduction which sometimes lead to aorta tear with pain and troubles to stitch it, and endothelial damage with post op aorta thrombosis. Giving aspirin to rats before stentgraft implantation decreases failure rate without major hemorrhage. Clamping time activates neutrophils, endothelium and platelets, and may interfere with biological analysis.

Histological and biochemical modifications after aneurysm endograft exclusion are still unclear. We describe a new model of endograft implantation on a murine aneurysm. Through thrombus analysis with and without persistant circulating blood stream interface, and new endovascular biomaterials evaluation, this model will help to better understand endovascular aneurysm exclusion pathobiology.

Ein neues Mausmodell der endovaskuläre Aortenaneurysma Repair

Endovascular aneurysm exclusion is a validated technique to prevent aneurysm rupture. Long-term results highlight technique limitations and new aspects of ...

Intrastriatal Injection of Autologous Blood or Clostridial Collagenase as Murine Models of Intracerebral Hemorrhage

Intracerebral hemorrhage (ICH) is a common form of cerebrovascular disease and is associated with significant morbidity and mortality. Lack of effective treatment and failure of large clinical trials aimed at hemostasis and clot removal demonstrate the need for further mechanism-driven investigation of ICH. This research may be performed through the framework provided by preclinical models. Two murine models in popular use include intrastriatal (basal ganglia) injection of either autologous whole blood or clostridial collagenase. Since, each model represents distinctly different pathophysiological features related to ICH, use of a particular model may be selected based on what aspect of the disease is to be studied. For example, autologous blood injection most accurately represents the brain&#39;s response to the presence of intraparenchymal blood, and may most closely replicate lobar hemorrhage. Clostridial collagenase injection most accurately represents the small vessel rupture and hematoma evolution characteristic of deep hemorrhages. Thus, each model results in different hematoma formation, neuroinflammatory response, cerebral edema development, and neurobehavioral outcomes. Robustness of a purported therapeutic intervention can be best assessed using both models. In this protocol, induction of ICH using both models, immediate post-operative demonstration of injury, and early post-operative care techniques are demonstrated. Both models result in reproducible injuries, hematoma volumes, and neurobehavioral deficits. Because of the heterogeneity of human ICH, multiple preclinical models are needed to thoroughly explore pathophysiologic mechanisms and test potential therapeutic strategies.

Preclinical models of intracerebral hemorrhage are utilized to mimic certain aspects of clinical disease. Thus, mechanisms of injury and potential therapeutic strategies may be explored. In this protocol, two models of intracerebral hemorrhage are described, intrastriatal (basal ganglia) injections of autologous blood or collagenase.

Intrastriataler Injektion von Eigenblut oder Clostridien-Kollagenase als Murine Modelle der Hirnblutung

Intracerebral hemorrhage (ICH) is a common form of cerebrovascular disease and is associated with significant morbidity and mortality. Lack of effective ...

A Preclinical Murine Model of Hepatic Metastases

Numerous murine models have been developed to study human cancers and advance the understanding of cancer treatment and development. Here, a preclinical, murine pancreatic tumor model of hepatic metastases via a hemispleen injection of syngeneic murine pancreatic tumor cells is described. This model mimics many of the clinical conditions in patients with metastatic disease to the liver. Mice consistently develop metastases in the liver allowing for investigation of the metastatic process, experimental therapy testing, and tumor immunology research.

A preclinical, murine model of hepatic metastases performed via a hemispleen injection technique.

Einer präklinischen Mausmodell der Lebermetastasen

Numerous murine models have been developed to study human cancers and advance the understanding of cancer treatment and development. Here, a preclinical, ...

The Rabbit Blood-shunt Model for the Study of Acute and Late Sequelae of Subarachnoid Hemorrhage: Technical Aspects

Early brain injury and delayed cerebral vasospasm both contribute to unfavorable outcomes after subarachnoid hemorrhage (SAH). Reproducible and controllable animal models that simulate both conditions are presently uncommon. Therefore, new models are needed in order to mimic human pathophysiological conditions resulting from SAH.
This report describes the technical nuances of a rabbit blood-shunt SAH model that enables control of intracerebral pressure (ICP). An extracorporeal shunt is placed between the arterial system and the subarachnoid space, which enables examiner-independent SAH in a closed cranium. Step-by-step procedural instructions and necessary equipment are described, as well as technical considerations to produce the model with minimal mortality and morbidity. Important details required for successful surgical creation of this robust, simple and consistent ICP-controlled SAH rabbit model are described.

The experimental intracranial pressure-controlled blood shunt subarachnoid hemorrhage (SAH) model in the rabbit combines the standard procedures &#8212; subclavian artery cannulation and transcutaneous cisterna magna puncture, which enables close mimicking of human pathophysiological conditions after SAH. We present step-by-step instructions and discuss key surgical points for successful experimental SAH creation.

Das Kaninchen Blut-Shunt-Modell für die Untersuchung von akuten und Spätfolgen Subarachnoidalblutung: Technische Aspekte

Early brain injury and delayed cerebral vasospasm both contribute to unfavorable outcomes after subarachnoid hemorrhage (SAH). Reproducible and ...

A Murine Model of Irreversible and Reversible Unilateral Ureteric Obstruction

Obstruction of the kidney may affect native or transplanted kidneys and results in kidney injury and scarring. Presented here is a model of obstructive nephropathy induced by unilateral ureteric obstruction (UUO), which can either be irreversible (UUO) or reversible (R-UUO). In the irreversible UUO model, the ureter may be obstructed for variable periods of time in order to induce increasingly severe renal inflammation and interstitial fibrotic scarring. In the reversible R-UUO model the ureter is obstructed to induce hydronephrosis, tubular dilation and inflammation. After a suitable period of time the ureteric obstruction is then surgically reversed by anastomosis of the severed previously obstructed ureter to the bladder in order to allow complete decompression of the kidney and restoration of urinary flow to the bladder. The irreversible UUO model has been used to investigate various aspects of renal inflammation and scarring including the pathogenesis of disease and the testing of potential anti-inflammatory or anti-fibrotic therapies. The more challenging model of R-UUO has been used by some investigators and does offer significant research potential as it allows the study of inflammatory and immune processes and tissue remodeling in an injured and scarred kidney following the removal of the injurious stimulus. As a result, the R-UUO model offers investigators the opportunity to explore the resolution of kidney inflammation together with key aspects of tissue repair. These experimental models are of relevance to human disease as patients often present with obstruction of the renal tract that requires decompression and are commonly left with significant residual kidney impairment that has no current treatment options and may lead to eventual end stage kidney failure.

The murine model of irreversible unilateral ureteric obstruction (UUO) is presented together with the model of reversible UUO in which the ureteric obstruction is relieved by anastomosis of the severed ureter into the bladder. These models enable the study of renal inflammation and scarring as well as tissue remodeling.

Ein Mausmodell der irreversiblen und reversiblen Einseitige Ureterobstruktion

Obstruction of the kidney may affect native or transplanted kidneys and results in kidney injury and scarring. Presented here is a model of obstructive ...

Pre-clinical Orthotopic Murine Model of Human Prostate Cancer

To study the multifaceted biology of prostate cancer, pre-clinical in vivo models offer a range of options to uncover critical biological information about this disease. The human orthotopic prostate cancer xenograft mouse model provides a useful alternative approach for understanding the specific interactions between genetically and molecularly altered tumor cells, their organ microenvironment, and for evaluation of efficacy of therapeutic regimens. This is a well characterized model designed to study the molecular events of primary tumor development and it recapitulates the early events in the metastatic cascade prior to embolism and entry of tumor cells into the circulation. Thus it allows elucidation of molecular mechanisms underlying the initial phase of metastatic disease. In addition, this model can annotate drug targets of clinical relevance and is a valuable tool to study prostate cancer progression. In this manuscript we describe a detailed procedure to establish a human orthotopic prostate cancer xenograft mouse model.

Prostate cancer is the second most common cause of cancer-related deaths in the United States. An orthotopic cancer model provides a useful approach to understand the biology of prostate cancer and to evaluate the efficacy of therapeutic regimens. This protocol describes detailed steps necessary to establish an orthotopic prostate cancer mouse model.

Präklinische Orthotopische Mausmodell der menschlichen Prostata-Krebs

To study the multifaceted biology of prostate cancer, pre-clinical in vivo models offer a range of options to uncover critical biological information ...

Integrated Compensatory Responses in a Human Model of Hemorrhage

Hemorrhage is the leading cause of trauma-related deaths, partly because early diagnosis of the severity of blood loss is difficult. Assessment of hemorrhaging patients is difficult because current clinical tools provide measures of vital signs that remain stable during the early stages of bleeding due to compensatory mechanisms. Consequently, there is a need to understand and measure the total integration of mechanisms that compensate for reduced circulating blood volume and how they change during ongoing progressive hemorrhage. The body&#39;s reserve to compensate for reduced circulating blood volume is called the &#39;compensatory reserve&#39;. The compensatory reserve can be accurately evaluated with real-time measurements of changes in the features of the arterial waveform measured with the use of a high-powered computer. Lower Body Negative Pressure (LBNP) has been shown to simulate many of the physiological responses in humans associated with hemorrhage, and is used to study the compensatory response to hemorrhage. The purpose of this study is to demonstrate how compensatory reserve is assessed during progressive reductions in central blood volume with LBNP as a simulation of hemorrhage.

The purpose of this protocol is to demonstrate the techniques for measuring compensatory responses to reduced central blood volume using lower body negative pressure as a noninvasive experimental model of human hemorrhage which can be used to quantify the total integration of compensatory mechanisms to blood volume deficit in humans.

Integrierte kompensatorische Antworten in einem menschlichen Modell von Einblutung

Hemorrhage is the leading cause of trauma-related deaths, partly because early diagnosis of the severity of blood loss is difficult. Assessment of ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

Messung der isometrischen Kontraktilität der Maus mesenterialen Arterie mit Draht Myographie

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...