A atual pesquisa é financiada pela Fundação Solorz-Zak Research.

Todos os procedimentos cirúrgicos foram submetidos à revisão ética e aprovado pelo governo da Alta Baviera (número de referência: 55.2-1-54-2532.3-13-13 e -2532-136-11). Os animais são macho C57BL / 6 com um peso corporal de aproximadamente 25 g. 1. Preparação animal <ol><li> Induzir a anestesia, colocando o mouse em uma câmara. Lave a câmara com 5% de isoflurano até que o animal perde a consciência. </li><li> Injectar anestésicos pré-misturados por via intraperitoneal: o fentanil (0,05 mg / kg), o midazolam (5 mg / kg) e medetomidina (0,5 mg / kg). Verifique reflexos antes e regularmente durante o procedimento. Reinject um terço do volume inicial por hora para manter a anestesia. </li><li> Entubar orotracheally com um tubo feito a partir de um cateter venoso 20 L 11. Para intubação fixar o animal em uma plataforma inclinada (30 °), retrair a língua com uma pinça dobrados, visualizar as cordas vocais sob um microscópio de operação e inserir o tubo na traquéiadurante a inspiração. </li><li> Posicione o mouse em uma posição propensa e verificar o correto posicionamento do tubo com um pedaço de algodão ou um microcapnograph. </li><li> Ligue o tubo de intubação com o respirador. Ventile o mouse com ar ambiente suplementado com 25% de oxigênio com uma frequência de 180-220 ciclos / min e um volume de ejeção de 200-250 mL. </li><li> Ligue o tubo de intubação com a microcapnograph. Manter o final da expiração de pCO 2 a 30 mmHg, ajustando a frequência de ventilação. </li><li> Insira uma sonda de temperatura rectal e colocar o animal numa almofada de aquecimento, a fim de manter a 37 ° C de temperatura corporal central. </li><li> Aplicar o sensor Pulsoximeter anelar na pata traseira direita. </li><li> Abra a pele acima do crânio com um par de tesouras. A incisão deve ser de aproximadamente 0,5 cm de comprimento e entre olhos e ouvidos. </li><li> Dissecar o músculo temporal esquerdo com um bisturi do osso temporal. </li><li> Cole o fluxômetro laser Doppler (LDF) da sonda emo osso temporal esquerdo. Segurar a sonda numa posição fixa até que a cola endurece. </li><li> Faça um furo de aproximadamente 1,5 mm de diâmetro com uma broca de dentista dentro do osso temporal esquerdo. Arrefecer o osso com uma solução salina para evitar danos causados ​​pelo calor. </li><li> Insira a sonda de ICP na cavidade craniana. Empurrar para a frente como dorsalmente quanto possível para impedir danos no tecido cerebral e hemorragia. </li><li> Se a sonda estiver na posição certa, corrigir e selá-lo com cimento. Deixe secar cimento por 5 min. </li><li> Vire o mouse com cuidado para a posição supina. </li><li> Para monitorização contínua da pressão arterial, cateterização da artéria femoral esquerda. </li><li> Conectar o cateter femoral para o dispositivo de monitorização da pressão arterial. </li></ol> 2. SAH Indução <ol><li> Abra a pele com um par de tesouras de esterno de queixo (2 cm). Dissecar o tecido conjuntivo sem rodeios e empurre as glândulas salivares de lado. </li><li> Exponha a artéria carótida comum esquerda (CCA) e mobilizá-la. Preserve onervo vago, que corre na mesma bainha do tecido conjuntivo, como o CCA. Mova craniana e expor e mobilizar a ICA ea ECA usando a mesma técnica. </li><li> Ligadura do ECA, tanto cranial possível. </li><li> Prearrange mais duas ligaduras para o filamento ao redor do ECA. </li><li> Ocluir o CCA ea ICA temporariamente com microclips. Posicione os microclips com um aplicador microclipe. Certifique-se que os grampos são aplicados corretamente, puxando-os para trás. </li><li> Corte um buraco para a inserção de filamento no ECA com uma tesoura dos vasos. </li><li> Insira um filamento Prolene 5-0 com 12 mm de comprimento no ECA. </li><li> Feche o local de inserção com uma ligadura premeditado. </li><li> Retire os microclips com um aplicador microclipe do CCA e da ACI. </li><li> Avançar o filamento com uma pinça para o ICA até o ICP sobe. Um aumento repentino do ICP indica indução sangramento. </li><li> Retire o filamento imediatamente e ligadura do ECA, fechando tanto prligaduras earranged consecutivamente. Isso evita que o sangramento fora do local de inserção. </li><li> Suturar o ferimento da pele. </li><li> Monitorar os parâmetros fisiológicos do animal por mais 20 min. </li><li> Remover ICP e sondas FDL e suturar o ferimento da pele. </li></ol> 3. Fim da Experiência <ol><li> Perfundir o animal transcardíaca com 20 ml de solução salina (temperatura ambiente), seguido de 20 ml de 4% de PFA em PBS (4 ° C). </li><li> Dissecar o cérebro fora do crânio. Cortar o crânio, na linha média e entre as cavidades orbitais. Em seguida, descascar o osso a partir do cérebro. </li><li> Avaliar a distribuição de sangue no espaço subaracnóide. </li></ol> 4. Considerações Em caso de cirurgia de sobrevivência (não mostrado no vídeo) <ol><li> Injetar carprofeno (4 mg / kg por via subcutânea) para analgesia pós-operatória imediatamente após a indução da anestesia. Durante o período de observação pós-operatória carprofeno (4 mg / kg por via subcutânea) é injectada a cada 24 horas. </li&gt; <li> Em vez de medição de pressão arterial invasiva fazer uso de um sistema de monitoramento de pressão arterial não invasiva. </li><li> Para a interrupção da anestesia injectar agentes antagonizar subcutaneamente: naloxona (1,2 mg / kg), flumazenil (0,5 mg / kg) e atipamezol (2,5 mg / kg). </li><li> Extubar o animal. </li><li> Depois de recuperar de reflexos colocar o animal em uma câmara de pré-aquecido. Manter o animal em cerca de 32 ° C durante 24 hr. </li><li> Os animais são verificados regularmente para a respiração espontânea e seu estado geral durante as primeiras horas após a cirurgia. Se o tronco cerebral é afectada animais têm problemas respiratórios e deve ser sacrificados. </li><li> Os animais são controlados diariamente para a sua condição geral e do peso corporal, assim como para os défices neurológicos e sensoriais 15. </li></ol>

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	<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>

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As opções de tratamento após SAH são escassos e pouco ineficaz. Portanto, a fisiopatologia da lesão cerebral pós-hemorrágica precisa ser mais bem compreendidos, a fim de identificar novos alvos terapêuticos e desenvolver novas abordagens terapêuticas. Padronizado e modelos animais bem reprodutíveis em animais geneticamente modificados, ou seja, ratos, são cruciais para tais investigações. O modelo CWP tornou-se um modelo amplamente utilizado para a HAS, uma vez que se assemelha a fisiopatologia em ...

HAS é o subtipo tempos com o resultado menos benéfica para os pacientes: 40% dos pacientes morrem dentro de um mês após o sangramento 1 e sobreviventes raramente têm um resultado clinicamente favorável. A grande maioria da SAHOS espontâneas (80%) são causados ​​pela ruptura de aneurismas intracranianos, que são na sua maioria localizadas ao longo da anterior e posterior da artéria, a artéria basilar, artéria cerebral média (MCA) 2 comunicando. Tais aneurismas são difíceis de modelar em animais e, portanto, os modelos animais de SAH são realizadas quer por injecção do sangue para dentro do espaço / ventrículos cerebrais subaracnoideos ou por perfuração de um navio de endovascular subaracnóide. Injeção de sangue autólogo na cisterna magna é de fácil execução e reprodutível, como o volume de sangue pode ser controlada diretamente 3. Infelizmente, alguns aspectos da fisiopatologia da HAS, por exemplo, olesão do vaso, não pode ser modelado por este procedimento. Outra abordagem técnica para indução de HAS é a abertura de uma veia intracisternal 4. No entanto, o CWP intraluminal no ramo MCA parece ser o processo de que os modelos da fisiopatologia em seres humanos mais estreita 5. O método foi desenvolvido e descrito pela primeira vez em ratos por Bederson e colegas, e ao mesmo tempo por Veelken e colegas 6,7. Mais tarde, o modelo da perfuração intraluminal foi adaptado para ratinhos 8,9. Um filamento é introduzido na artéria carótida externa (ECA) e avançada até à base do crânio, através da artéria carótida interna (ICA). No ponto de ramificação do MCA filamento perfura o vaso e induz a uma hemorragia no espaço subaracnóide da base do crânio. O sangue, em seguida, distribui para o espaço subaracnóide restante ao longo de fissuras e vasos sanguíneos. A hemorragia é parada pela formação de coágulos no local da perfuração, mas rebleedings, which são muitas vezes prejudiciais em pacientes de 10, pode ocorrer. Por conseguinte, o modelo de filamentos endovascular se tornou um modelo amplamente usado HAS durante os últimos anos. A desvantagem mais citada do modelo filamento perfuração é que o volume de sangramento não pode ser diretamente controlado e pode, portanto, ser variável. Esta variabilidade pode ser significativamente reduzido por um rígido controle de fisiologia animal e pós-hemorrágica ICP. Os ratos têm a grande vantagem de que um grande número de estirpes geneticamente modificados estão disponíveis. No entanto, devido ao seu pequeno tamanho procedimentos cirúrgicos tendem a ser mais complexo do que em espécies maiores, por exemplo, ratos ou coelhos. Portanto, a redução de escala de técnicas desenvolvidas para os ratos para os ratos costumam não leva aos resultados desejados, por exemplo, como os ratos têm um peso corporal e volume de sangue muito limitadas técnicas não invasivas para a pressão arterial e análise de gases sanguíneos, bem como para a saturação de hemoglobina e monitoramento da freqüência cardíacatem que ser aplicada sempre que possível. Por conseguinte, o objectivo da publicação é descrever o modelo da perfuração de filamentos para HSA em ratinhos e para demonstrar como este modelo pode ser realizado de uma forma padronizada e altamente reprodutível.

<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

Nesta publicação de vídeo de um modelo padronizado rato de hemorragia subaracnóide (HSA) é apresentado. Sangramento é induzida pelo Círculo endovascular de Willis perfuração (CWP) e comprovada por pressão intracraniana (PIC) de monitoramento. Assim, uma distribuição homogênea de sangue em espaços subaracnóide em torno da circulação arterial e fissuras do cerebelo é alcançado. Fisiologia animal é mantido por entubação, ventilação mecânica, monitorização contínua e on-line de vários parâmetros fisiológicos e cardiovasculares: temperatura corporal, pressão arterial sistêmica, freqüência cardíaca e saturação de hemoglobina. Deste modo, a pressão de perfusão cerebral pode ser firmemente monitorado resultando em um volume variável de menos de sangue extravasado. Isto permite uma melhor padronização de perfuração filamento endovascular em ratos e faz todo o modelo altamente reprodutível. Assim, é facilmente disponível para os estudos farmacológicos e fisiopatológicos no tipo selvagem e genéticaly alterada ratinhos.

Mortalidade Uma vez que a técnica de cirurgia é dominado o procedimento não provocou quaisquer mortalidade intra-operatória. Além disso o sangramento pode ser conseguida em quase todos os animais. Mortalidade pós-operatória é de 30-40%, com a maior parte dos animais morrer no dia 1 após a cirurgia (Figura 5). ICP valores após SAH O ICP antes de sangramento é de cerca de 4 mmHg. Sangramento resulta em um aumento a...

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

A Murine Model of Subarachnoid Hemorrhage

Um modelo de rato padronizado de hemorragia subaracnóide por Círculo intraluminal de Willis perfuração é descrito. Perfuração de vasos e hemorragia subaracnóide são monitorados pela monitorização da pressão intracraniana. Além disso os vários parâmetros vitais são registadas e controlada para manter as condições fisiológicas.

Um modelo murino de hemorragia subaracnóide

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 visualizações.  University of Munich Medical Center.  Um modelo de rato padronizado de hemorragia subaracnóide por Círculo intraluminal de Willis perfuração é descrito. Perfuração de vasos e hemorragia subaracnóide são monitorados pela monitorização da pressão intracraniana. Além disso os vários parâmetros vitais são registadas e controlada para manter as condições fisiológicas. 

Vídeo: 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.

Um Modelo de Rato Baixa Mortalidade para Avaliar vasoespasmo cerebral adiada após hemorragia subaracnóide Experimental

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

Medicina

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.

Modelo Murino de Cicatrização

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.

Um novo modelo murino de reparação endovascular de aneurisma da aorta

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.

Injection intraestriatal de sangue autólogo ou Clostridial colagenase como modelos murinos de hemorragia intracerebral

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.

Um Modelo murino de metástases hepáticas pré-clínica

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.

O coelho Modelo Blood-shunt para o Estudo da aguda e tardia Sequelas de hemorragia subaracnóidea: Aspectos Técnicos

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.

Um modelo murino de irreversível e reversível obstrução ureteral unilateral

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

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.

Respostas compensatórias integrados em um modelo humano de hemorragia

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.

Medição de contratilidade isométrica da artéria mesentérica Mouse usando fio miografia

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

A Preclinical Controlled Cortical Impact Model for Traumatic Hemorrhage Contusion and Neuroinflammation

Cerebral contusion is a severe medical problem affecting millions of people worldwide each year. There is an urgent need to understand the pathophysiological mechanism and to develop effective therapeutic strategy for this devastating neurological disorder. Intraparenchymal hemorrhage and post-traumatic inflammatory response induced by initial physical impact can aggravate microglia/macrophage activation and neuroinflammation which subsequently worsen brain pathology. We provide here a controlled cortical impact (CCI) protocol that can reproduce experimental cortical contusion in mice by using a pneumatic impactor system to deliver mechanical force with controllable magnitude and velocity onto the dural surface. This preclinical model allows researchers to induce moderately severe focal cerebral contusion in mice and to investigate a wide range of post-traumatic pathological progressions including hemorrhage contusion, microglia/macrophage activation, iron toxicity, axonal injury, as well as short-term and long-term neurobehavioral deficits. The present protocol can be useful for exploring the long-term effects of and potential interventions for cerebral contusion.

<meta charset="utf8"></head><body>Um modelo pré-clínico de impacto cortical controlado para hemorragia traumática, contusão e neuroinflamação

Intraparenchymal hemorrhage and neuroinflammation accompanied by cerebral contusion can trigger severe secondary brain injury. This protocol details a mouse controlled cortical impact (CCI) model allowing researchers to study hemorrhage contusion and post-traumatic immune responses and explore potential therapeutics.

Methods Article

Um modelo murino de hemorragia subaracnóide