This work has been supported by NIH HL-1034.

O Comitê de Cuidados e Uso de Animais da Universidade Johns Hopkins aprovado todos os protocolos de animais. 1. Equipamentos O sistema composto definido acima, pronta para medir a curva de PV é mostrado na Figura 1. <ol><li> Medição de volume: <ol><li> Gerar uma taxa constante de inflação e deflação, utilizando uma bomba de seringa com um interruptor que permite que o usuário para reverter rapidamente a bomba depois de atingir os limites de pressão. Para as curvas do mouse PV, use um muito levemente untada 5 ml seringa de vidro com o volume inicial (antes da inflação), fixado em 3 ml de ar. 3 ml é suficientemente grande para medir volumes em quase todas as curvas do mouse PV. </li><li> Medir o volume entregue pela bomba anexando um transformador diferencial linear para o alojamento da bomba, com uma pequena haste do sensor ligado ao êmbolo da seringa movendo. Nota: Um meio empíricos para corrigir para a compressão de gás no sistema é descrito sob o cu PVsecção de gravação rve. </li></ol></li><li> Medição de pressão: <ol><li> Usar um medidor de pressão barato padrão com um intervalo de 0-60 cm de H 2 O (0-1 psi). </li></ol></li><li> Gravação de medição: <ol><li> Para gravar a curva PV usar qualquer gravador digital com recursos de XY (por exemplo, PowerLab). Defina um canal para gravar o sinal de volume corrigido e outro canal para registrar a pressão transpulmonar (PTP), a fim de elaborar o gráfico da curva PV. Usar um pré-amplificador de ponte que liga ao Powerlab principal para medir a pressão. Calibrar o canal de pressão 0-40 cm H 2 O, e calibrar o canal de volume de 0-3 ml. </li></ol></li></ol> 2. Correção de Compressão de Gás Nota: Este é um passo inicial importante na configuração, uma vez que como a pressão aumenta, a diminuição do volume de gás, e, assim, o volume de ar fornecido para o rato irá ser cada vez menor do que o deslocamento do syrInge barril. <ol><li> Fechar a torneira que vai ligar o sistema de PV para os pulmões, para que o gás não possa deixar o sistema. Iniciar a infusão e observar se o canal Volume corrigido no gravador mostra todas as mudanças mensuráveis, como a pressão aumenta para cerca de 40 cm H 2 O. Se assim for, então correcta como nos passos seguintes. <ol><li> Correto para compressão de gás empiricamente subtraindo a partir da medição de deslocamento êmbolo (ou seja, o volume não corrigido) um termo proporcional à pressão de inflação. Faça isso em um canal Powerlab (chamado Vc) para mostrar o sinal de volume, menos um coeficiente vezes a pressão. </li><li> Determine o coeficiente na equação. Primeiro, faça uma estimativa inicial, vire o registro gráfico, e ligar a bomba. Uma vez que o tubo de inflação é selada, ajustar o coeficiente multiplicador de pressão para fazer o canal Vc ler zero como a pressão aumenta, 0-40 cm de H 2 O. Se ele vai para cima ou para baixo, basta ajustar o fator de correção até quepermanece estável ao longo da faixa de pressão. Este factor de correcção será sempre o mesmo, se as mesmas a partir de 3 ml de volume na seringa não é alterado. </li></ol></li></ol> 3. testes experimentais em camundongos <ol><li> Processo para a medição da curva de PV em ratinhos. Todos os protocolos de animais foram aprovados pelo Comitê de Johns Hopkins University Animal Care e Use. <ol><li> Anestesiar ratos (C57BL / 6 ratos às 6-12 semanas de idade) com cetamina (90 mg / kg) e xilazina (15 mg / kg), e confirmar a anestesia pela ausência do reflexo do movimento. Nota: A curva PV pode ser completada em ratos anestesiados, em menos de 10 min e é um procedimento terminal. </li><li> Tracheostomize os ratos com uma agulha 18 G stub cânula. Fazer isso através de uma pequena incisão na pele que recobre a traqueia, localizando a traqueia, em seguida, fazendo um pequeno corte na traqueia, onde o topo da agulha pode ser inserida. Fixe a cânula, amarrando com rosca. </li><li> Permitir que os ratos para respiração 100% de oxigênio para, pelo menos, 4 min. Isso pode ser feito através de respiração espontânea de um saco ou com um ventilador nominalmente definido com um volume corrente de 0,2 ml a 150 ciclos / min. </li><li> Fechar a cânula traqueal e permitir 3-4 min para o rato para absorver todo o oxigénio. Este procedimento de absorção de oxigénio resulta a morte dos animais e de uma desgaseificação quase completa do pulmão 11. Confirme morte do rato através da medição da cessação dos batimentos cardíacos com eletrodos de ECG ou observação direta. </li><li> Uma vez que a desgaseificação do pulmão está completa e o volume do pulmão é zero, começar inflar os pulmões com ar ambiente na bomba de seringa a uma taxa de 3 ml / min. Monitorar o traçado da pressão sobre o gravador digital, e quando chega a 35 cm de H 2 O, inverter a bomba. </li><li> Siga a curva de deflação até que a pressão atinge negativa de 10 cm H 2 O, época em que as vias aéreas entraram em colapso, prendendo o ar nos alvéolos que impedem uma maior redução de volume. Reverter imediatamente tele bombear novamente, permitindo que o pulmão de reinsuflação como as vias aéreas colapsadas abrir. Esta abertura heterogénea é normalmente aparente pelo membro inflação barulhento olhando para a parte inicial desta segunda inflação. </li><li> Quando a pressão novamente chega a 35 cm H 2 O, inverter a direção da bomba, e continuar a desinflar o pulmão até esta segunda parte deflação chega a 0 cm H 2 O. Em seguida, parar a bomba. </li><li> Veja o registro gráfico PowerLab de pressão e fluxo ea curva PV. Em seguida, analisar a curva PV para detectar alterações fenotípicas em parênquima pulmonar que ocorrem com diferentes patologias pulmonares. </li></ol></li></ol>

<ol>
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None of the authors have any financial interests that would be in conflict with the material presented in this paper.

Neste papel um método reprodutível simples tem sido descrito para medir em ratinhos um método clássico de fenotipagem elasticidade pulmonar, a curva total PV pulmão. Essas curvas foram fundamentais para a descoberta de surfactante pulmonar e sua importância no fornecimento de estabilidade de pulmão. Aqui mostra-se como a curva PV também é útil ao proporcionar um meio para medir diversas variáveis ​​relacionadas com a elasticidade pulmonar no pulmão adulto do mouse. Houve mudanças muito significativas em...

O rato é agora o modelo animal primária de uma variedade de doenças pulmonares. Em modelos de enfisema ou fibrose, as alterações fenotípicas essenciais são melhor avaliadas medindo as alterações na elasticidade do pulmão. Embora existam muitas maneiras de medir a elasticidade, o método clássico é que a curva de pressão-volume total (PV) medido a partir do volume residual (VR) a capacidade pulmonar total (CPT). Essa medição foi feita no pulmão adulto de quase todas as espécies de mamíferos que remonta quase 100 anos 1-3. Tais curvas PV também desempenhou um papel importante na descoberta e compreensão da função do surfactante pulmonar em desenvolvimento pulmonar fetal 4-7. Apesar de a importância da curva de PV como uma medição do fenótipo do pulmão, não houve nenhuma maneira padronizada de efectuar esta medição. Foi feito simplesmente inflar e desinflar o pulmão com passos discretos (esperando um tempo variável de equilíbrio após cada) ou com bombas quepode continuamente inflar e desinflar o pulmão. A curva PV é feito muitas vezes ao longo de um intervalo de volume entre zero e alguma capacidade pulmonar definir pelo usuário, mas o tempo de duração de cada ciclo de volume pressão relatada por diferentes laboratórios tem sido extremamente variável, variando de alguns segundos de 8 a 2 hr. Alguns pesquisadores se referem a esta curva PV pulmonar total como estático ou quasiestático, mas estes são termos qualitativos que oferecem pouco de visão, e eles não são utilizados aqui. Além disso, a curva de PV não tem sido amplamente relatado no ratinho, apesar do facto de que pode fornecer informação útil sobre os efeitos de alterações estruturais macroscópicos nos pulmões. Várias questões resultaram em variabilidade na aquisição da curva PV incluindo: 1) a taxa de inflação e deflação; 2) as excursões de pressão para a inflação e deflação; e 3) os meios para determinar uma medida absoluta do volume pulmonar. No método aqui presente, uma taxa de 3 ml / min foi escolhido como um compromise, não sendo demasiado curto para refletir a elasticidade dinâmica associada à ventilação normal e não é demasiado lenta para fazer a medição impraticável, particularmente quando se estuda grandes grupos. Uma vez que uma capacidade pulmonar total nominal em um / 6 rato saudável C57BL é da ordem de 1,2 ml 9, esta taxa normalmente permite que dois completo fechado PV loops para ser feito em cerca de 1,5 min. Na literatura extensas onde as curvas PV foram relatados, a pressão de enchimento pico utilizado tem sido extremamente variável, variando de tão baixo como 20 a mais de 40 cm de H 2 O. Parte dessa variabilidade pode estar relacionada à espécie, mas o objetivo principal de estabelecer o limite de pressão superior para curvas PV é inflar o pulmão a capacidade pulmonar total (CPT), ou volume pulmonar máxima. A TLC em seres humanos é definida pelo esforço voluntária máxima de um indivíduo pode fazer, mas, infelizmente, esta nunca poderá ser duplicada em qualquer modelo animal. Assim, o volume máximo em curvas PV experimentais é dissuadirminado por uma pressão máxima arbitrariamente definida pelo investigador. O objectivo é estabelecer uma pressão onde a curva PV é plana, mas, infelizmente, o membro de inflação de uma curva PV pulmão de mamífero é nunca plana. Assim, a maioria dos investigadores definir uma pressão onde a curva começa a inflação para achatar substancialmente, tipicamente de 30 cm H 2 O. No rato, no entanto, a curva PV é ainda mais complexo, com uma corcunda dupla no membro inflação, e onde esta parte da inflação é muitas vezes continua a aumentar acentuadamente a 30 cm H 2 O 10, portanto 30 não é um bom ponto final para a curva PV. Por esta razão, é utilizado 35 centímetros H2O como o limite da pressão para a curva de rato PV, que é uma pressão à qual os membros de enchimento de todas as estirpes que examinamos começam a achatar. Uma vez que a própria curva PV é muito linear, o aparecimento de um loop PV irá depender do volume a partir de onde a curva começa. Alguns ventiladores comerciais permitem que os usuários façam grandes laços PV, a partir de FRC, mas se o volume da FRC é desconhecido, então é possível interpretar tais mudanças na curva de PV com qualquer patologia, uma vez que essas modificações podem simplesmente resultar de uma alteração no volume de partida, e não alterações estruturais no pulmão. Assim, sem uma medição de volume absoluto, curvas PV são quase impossíveis de interpretar e, portanto, têm pouca utilidade. Apesar de, há várias maneiras de medir volumes pulmonares, estes são muitas vezes complicado e requer equipamento especial. Na abordagem simples descrita aqui, a curva PV começa no volume zero após um processo de desgaseificação in vivo. Em resumo, este trabalho demonstra um método simples para padronizar medição curva PV de pulmão no pulmão do rato, e define várias métricas que podem ser calculados a partir da curva que estão ligados à estrutura do pulmão. A curva PV, portanto, fornece um teste de função pulmonar, que tem aplicação direta em ser capaz de detectar mudanças estruturais fenotípicas em camundongos com commem patologias pulmonares como enfisema e fibrose.

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			<td height="21" style="height:21px;width:244px;">Name of Material/ Equipment</td>
			<td style="width:127px;">Company</td>
			<td style="width:155px;">Catalog Number</td>
			<td style="width:167px;">Comments/Description</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:244px;"></td>
			<td style="width:127px;"></td>
			<td style="width:155px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:244px;"></td>
			<td style="width:127px;"></td>
			<td style="width:155px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:244px;">&#160;Syringe Pump</td>
			<td style="width:127px;">Harvard Apparatus</td>
			<td>55-2226</td>
			<td>Infuse/Withdraw syringe pump</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;">Pump 22 Reversing Switch&#160;</td>
			<td style="width:127px;">Harvard Apparatus</td>
			<td>552217</td>
			<td>&#160;included with pump</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;"></td>
			<td style="width:127px;"></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:244px;">Linear displacement transformer</td>
			<td style="width:127px;">Trans-Tek, Inc.</td>
			<td style="width:155px;">0244-0000</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:244px;"></td>
			<td style="width:127px;"></td>
			<td style="width:155px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:244px;">5 mL glass syringe</td>
			<td style="width:127px;">Becton Dickenson</td>
			<td style="width:155px;"></td>
			<td>Several other possible vendors</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:244px;"></td>
			<td style="width:127px;"></td>
			<td style="width:155px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:244px;">Digital recorder</td>
			<td style="width:127px;">ADInstruments</td>
			<td>PL3504</td>
			<td>Several other possible vendors</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Bridge Amp Signal Conditioner</td>
			<td style="width:127px;">ADInstruments</td>
			<td>FE221</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;"></td>
			<td style="width:127px;"></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:244px;">Gas tank,100% oxygen</td>
			<td style="width:127px;">Airgas, Inc</td>
			<td style="width:155px;"></td>
			<td>Any supplier or hospital source will work</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:244px;"></td>
			<td style="width:127px;"></td>
			<td style="width:155px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Pressure Transducer - 0-1psi&#160; millivolt output</td>
			<td>Omega Engineering</td>
			<td>PX-137</td>
			<td>Range: &#8776;0-60 cmH2O</td>
		</tr>
	</tbody>
</table>

measurement of the pressure-volume curve in mouse lungs

Nas últimas décadas, o ratinho tornou-se o modelo animal primária de uma variedade de doenças pulmonares. Em modelos de enfisema ou fibrose, as alterações fenotípicas essenciais são melhor avaliada por medição das alterações na elasticidade do pulmão. Para melhor compreender os mecanismos específicos subjacentes a essas patologias em camundongos, é essencial para fazer medições funcionais que podem refletir a patologia em desenvolvimento. Embora existam muitas maneiras de medir a elasticidade, o método clássico é o do pulmão pressão-volume total (PV) curva feito ao longo de toda a gama de volumes pulmonares. Essa medição foi feita no pulmão adulto de quase todas as espécies de mamíferos que remonta quase 100 anos, e essas curvas PV também desempenhou um papel importante na descoberta e compreensão da função do surfactante pulmonar em desenvolvimento pulmonar fetal. Infelizmente, tais curvas totais PV não têm sido amplamente relatado na rato, apesar do fato de que eles podem fornecer informações úteis sobre o macroscOPIC efeitos de mudanças estruturais no pulmão. Embora curvas PV parciais para avaliar apenas as variações no volume do pulmão são, por vezes relatados, sem uma medida de volume absoluto, a natureza não-linear da curva total PV faz com que essas parciais muito difícil de interpretar. No presente estudo, descrevemos uma maneira padronizada para medir a curva total PV. Estamos, então, testaram a capacidade dessas curvas para detectar mudanças na estrutura do pulmão do rato em duas patologias pulmonares comum, enfisema e fibrose. Os resultados mostraram alterações significativas em diversas variáveis ​​consistentes com as mudanças estruturais esperadas com estas patologias. Esta medição da curva de PV de pulmão em ratinhos proporciona assim um meio simples para controlar a progressão das alterações fisiopatológicas ao longo do tempo e o efeito potencial de procedimentos terapêuticos.

Embora o procedimento para as curvas PV é demonstrado no vídeo apenas para controlo de ratos saudáveis, foi examinada a capacidade da curva PV para detectar alterações funcionais e patológicos em ratinhos com duas patologias comuns diferentes, enfisema e fibrose. Os detalhes destes modelos tradicionais descrito em outros lugares 12,13. Muito brevemente, após anestesia com isoflurano a 3% foi enfisema causado por 3 ou 6 L elastase pancreática porcina instilado na traqueia e estudou 3 semanas mais tarde...

Watch this Scientific Journal Video about Measurement of the Pressure-volume Curve in Mouse Lungs at JoVE.com

Measurement of the Pressure-volume Curve in Mouse Lungs

Here we present a protocol to simply and reliably measure the lung pressure-volume curve in mice, showing that it is sufficiently sensitive to detect phenotypic parenchymal changes in two common lung pathologies, pulmonary fibrosis and emphysema. This metric provides a means to quantify the lung&#8217;s structural changes with developing pathology.

Medição da curva pressão-volume no mouse Lungs

In recent decades the mouse has become the primary animal model of a variety of lung diseases. In models of emphysema or fibrosis, the essential ...

measurement-of-the-pressure-volume-curve-in-mouse-lungs

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18.5K visualizações.  Johns Hopkins University. Here we present a protocol to simply and reliably measure the lung pressure-volume curve in mice, showing that it is sufficiently sensitive to detect phenotypic parenchymal changes in two common lung pathologies, pulmonary fibrosis and emphysema. This metric provides a means to quantify the lung’s structural changes with developing pathology.

Vídeo: Measurement of the Pressure-volume Curve in Mouse Lungs

The Measurement and Treatment of Suppression in Amblyopia

Amblyopia, a developmental disorder of the visual cortex, is one of the leading causes of visual dysfunction in the working age population. Current estimates put the prevalence of amblyopia at approximately 1-3%1-3, the majority of cases being monocular2. Amblyopia is most frequently caused by ocular misalignment (strabismus), blur induced by unequal refractive error (anisometropia), and in some cases by form deprivation.
Although amblyopia is initially caused by abnormal visual input in infancy, once established, the visual deficit often remains when normal visual input has been restored using surgery and/or refractive correction. This is because amblyopia is the result of abnormal visual cortex development rather than a problem with the amblyopic eye itself4,5 . Amblyopia is characterized by both monocular and binocular deficits6,7 which include impaired visual acuity and poor or absent stereopsis respectively. The visual dysfunction in amblyopia is often associated with a strong suppression of the inputs from the amblyopic eye under binocular viewing conditions8. Recent work has indicated that suppression may play a central role in both the monocular and binocular deficits associated with amblyopia9,10 .
Current clinical tests for suppression tend to verify the presence or absence of suppression rather than giving a quantitative measurement of the degree of suppression. Here we describe a technique for measuring amblyopic suppression with a compact, portable device11,12 . The device consists of a laptop computer connected to a pair of virtual reality goggles. The novelty of the technique lies in the way we present visual stimuli to measure suppression. Stimuli are shown to the amblyopic eye at high contrast while the contrast of the stimuli shown to the non-amblyopic eye are varied. Patients perform a simple signal/noise task that allows for a precise measurement of the strength of excitatory binocular interactions. The contrast offset at which neither eye has a performance advantage is a measure of the "balance point" and is a direct measure of suppression. This technique has been validated psychophysically both in control13,14 and patient6,9,11 populations.
In addition to measuring suppression this technique also forms the basis of a novel form of treatment to decrease suppression over time and improve binocular and often monocular function in adult patients with amblyopia12,15,16 . This new treatment approach can be deployed either on the goggle system described above or on a specially modified iPod touch device15.

Amblyopia is a developmental disorder of the visual cortex that is often accompanied by strong suppression of one eye. We present a new technique for measuring and treating interocular suppression in patients with amblyopia that can be deployed using virtual reality goggles or a portable iPod Touch device.

A Medição e Tratamento de Supressão de ambliopia

Amblyopia, a developmental disorder of the visual cortex, is one of the leading causes of visual dysfunction in the working age population. Current ...

Medicina

In vivo Measurement of the Mouse Pulmonary Endothelial Surface Layer

The endothelial glycocalyx is a layer of proteoglycans and associated glycosaminoglycans lining the vascular lumen. In vivo, the glycocalyx is highly hydrated, forming a substantial endothelial surface layer (ESL) that contributes to the maintenance of endothelial function. As the endothelial glycocalyx is often aberrant in vitro and is lost during standard tissue fixation techniques, study of the ESL requires use of intravital microscopy. To best approximate the complex physiology of the alveolar microvasculature, pulmonary intravital imaging is ideally performed on a freely-moving lung. These preparations, however, typically suffer from extensive motion artifact. We demonstrate how closed-chest intravital microscopy of a freely-moving mouse lung can be used to measure glycocalyx integrity via ESL exclusion of fluorescently-labeled high molecular weight dextrans from the endothelial surface. This non-recovery surgical technique, which requires simultaneous brightfield and fluorescent imaging of the mouse lung, allows for longitudinal observation of the subpleural microvasculature without evidence of inducing confounding lung injury.

In vivo Measurement of the Mouse Pulmonary Endothelial Surface Layer

The endothelial glycocalyx/endothelial surface layer is ideally studied using intravital microscopy. Intravital microscopy is technically challenging in a moving organ such as the lung. We demonstrate how simultaneous brightfield and fluorescent microscopy may be used to estimate endothelial surface layer thickness in a freely-moving in vivo mouse lung.

Na medição in vivo da camada superficial do rato pulmonar endotelial

The endothelial glycocalyx is a layer of proteoglycans and associated glycosaminoglycans lining the vascular lumen. In vivo, the glycocalyx is highly ...

Technical Aspects of the Mouse Aortocaval Fistula

Technical aspects of creating an arteriovenous fistula in the mouse are discussed. Under general anesthesia, an abdominal incision is made, and the aorta and inferior vena cava (IVC) are exposed. The proximal infrarenal aorta and the distal aorta are dissected for clamp placement and needle puncture, respectively. Special attention is paid to avoid dissection between the aorta and the IVC. After clamping the aorta, a 25 G needle is used to puncture both walls of the aorta into the IVC. The surrounding connective tissue is used for hemostatic compression. Successful creation of the AVF will show pulsatile arterial blood flow in the IVC. Further confirmation of successful AVF can be achieved by post-operative Doppler ultrasound.

The goal is to produce an arteriovenous fistula that is simple and reproducible. This method does not use sutures or glue adhesive. Therefore the samples can be used with the least amount of foreign materials for analysis.

Aspectos técnicos da artocava Fístula Rato

Technical aspects of creating an arteriovenous fistula in the mouse are discussed. Under general anesthesia, an abdominal incision is made, and the aorta ...

Myocardial Infarction and Functional Outcome Assessment in Pigs

Introduction of newly discovered cardiovascular therapeutics into first-in-man trials depends on a strictly regulated ethical and legal roadmap. One important prerequisite is a good understanding of all safety and efficacy aspects obtained in a large animal model that validly reflect the human scenario of myocardial infarction (MI). Pigs are widely used in this regard since their cardiac size, hemodynamics, and coronary anatomy are close to that of humans. Here, we present an effective protocol for using the porcine MI model using a closed-chest coronary balloon occlusion of the left anterior descending artery (LAD), followed by reperfusion. This approach is based on 90 min of myocardial ischemia, inducing large left ventricle infarction of the anterior, septal and inferoseptal walls. Furthermore, we present protocols for various measures of outcome that provide a wide range of information on the heart, such as cardiac systolic and diastolic function, hemodynamics, coronary flow velocity, microvascular resistance, and infarct size. This protocol can be easily tailored to meet study specific requirements for the validation of novel cardioregenerative biologics at different stages (i.e. directly after the acute ischemic insult, in the subacute setting or even in the chronic MI once scar formation has been completed). This model therefore provides a useful translational tool to study MI, subsequent adverse remodeling, and the potential of novel cardioregenerative agents.

This protocol describes the porcine myocardial infarction (MI) model using a 90 min closed-chest coronary balloon occlusion of the left anterior descending artery (LAD), followed by reperfusion. Furthermore, the protocol for several outcome parameters, such as cardiac function, hemodynamics, microvascular resistance, and infarct size, are also presented.

Infarto do Miocárdio e Avaliação de Resultados Funcional em porcos

Introduction of newly discovered cardiovascular therapeutics into first-in-man trials depends on a strictly regulated ethical and legal roadmap. One ...

Cardiac Catheterization in Mice to Measure the Pressure Volume Relationship: Investigating the Bowditch Effect

Animal models that mimic human cardiac disorders have been created to test potential therapeutic strategies. A key component to evaluating these strategies is to examine their effects on heart function. There are several techniques to measure in vivo cardiac mechanics (e.g., echocardiography, pressure/volume relations, etc.). Compared to echocardiography, real-time left ventricular (LV) pressure/volume analysis via catheterization is more precise and insightful in assessing LV function. Additionally, LV pressure/volume analysis provides the ability to instantaneously record changes during manipulations of contractility (e.g., &#946;-adrenergic stimulation) and pathological insults (e.g., ischemia/reperfusion injury). In addition to the maximum (+dP/dt) and minimum (-dP/dt) rate of pressure change in the LV, an accurate assessment of LV function via several load-independent indexes (e.g., end systolic pressure volume relationship and preload recruitable stroke work) can be attained. Heart rate has a significant effect on LV contractility such that an increase in the heart rate is the primary mechanism to increase cardiac output (i.e., Bowditch effect). Thus, when comparing hemodynamics between experimental groups, it is necessary to have similar heart rates. Furthermore, a hallmark of many cardiomyopathy models is a decrease in contractile reserve (i.e., decreased Bowditch effect). Consequently, vital information can be obtained by determining the effects of increasing heart rate on contractility. Our and others data has demonstrated that the neuronal nitric oxide synthase (NOS1) knockout mouse has decreased contractility. Here we describe the procedure of measuring LV pressure/volume with increasing heart rates using the NOS1 knockout mouse model.

This article describes the measurement of murine left ventricular function via pressure/volume analysis at different heart rates.

O cateterismo cardíaco em camundongos para medir o volume Pressão do relacionamento: Investigando o efeito Bowditch

Animal models that mimic human cardiac disorders have been created to test potential therapeutic strategies. A key component to evaluating these ...

Preparation Steps for Measurement of Reactivity in Mouse Retinal Arterioles Ex Vivo

Vascular insufficiency and alterations in normal retinal perfusion are among the major factors for the pathogenesis of various sight-threatening ocular diseases, such as diabetic retinopathy, hypertensive retinopathy, and possibly glaucoma. Therefore, retinal microvascular preparations are pivotal tools for physiological and pharmacological studies to delineate the underlying pathophysiological mechanisms and to design therapies for the diseases. Despite the wide use of mouse models in ophthalmic research, studies on retinal vascular reactivity are scarce in this species. A major reason for this discrepancy is the challenging isolation procedures owing to the small size of these retinal blood vessels, which is ~ &#8804; 30 &#181;m in luminal diameter. To circumvent the problem of direct isolation of these retinal microvessels for functional studies, we established an isolation and preparation technique that enables ex vivo studies of mouse retinal vasoactivity under near-physiological conditions. Although the present experimental preparations will specifically refer to the mouse retinal arterioles, this methodology can readily be employed to microvessels from rats.

Preparation Steps for Measurement of Reactivity in Mouse Retinal Arterioles Ex Vivo

Many vision-threatening ocular diseases are associated with dysfunctional retinal microvessels. Therefore, the measurement of retinal arteriole responses is important to investigate the underlying pathophysiological mechanisms. This article describes a detailed protocol for mouse retinal arteriole isolation and preparation to assess the effects of vasoactive substances on vascular diameter.

Etapas de preparação para medição de reatividade no Mouse arteríolas retina Ex Vivo

Vascular insufficiency and alterations in normal retinal perfusion are among the major factors for the pathogenesis of various sight-threatening ocular ...

Real-time Pressure-volume Analysis of Acute Myocardial Infarction in Mice

Acute myocardial infarction can lead to acute heart failure and cardiogenic shock. The evaluation of hemodynamics is critical for the evaluation of any potential therapeutic approach directed against acute left ventricular (LV) dysfunction. Current imaging modalities (e.g., echocardiography and magnetic resonance imaging) have several limitations since data on LV pressure cannot directly be measured. LV catheterization in mice undergoing coronary artery occlusion could serve as a novel method for a real-time evaluation of LV function.
At the beginning of the procedure, mice were anesthetized followed by endotracheal intubation. For LV catheterization, the right carotid artery was exposed via middle-neck incision. The catheter was introduced and placed into the LV cavity. Left thoracotomy was conducted and the left main coronary artery (LCA) was ligated. To induce reperfusion, the suture was released after 45 min. Pressure-volume data was recorded at all times.
Ligation of the LCA caused a decrease in LV systolic function as evidenced by a 30% reduction in stroke volume, LV ejection fraction (EF), and cardiac output. Maximum dP/dt as a parameter for LV contractility was also significantly reduced and diastolic function was severely impaired (minimum dP/dt -40%). Reperfusion over a period of 20 min did not lead to a complete recovery of LV function.
Real-time pressure-volume analysis served as a valid procedure for monitoring cardiac function during acute myocardial infarction in mice. Maintaining stable anesthesia and a standardized surgical approach was crucial to ensure valid results. As the early phase of acute myocardial infarction is critical for morbidity and mortality, the delineated method could be beneficial for preclinical evaluation of new strategies for cardioprotection.

Acute myocardial infarction in mice induces acute but incompletely characterized changes in left ventricular (LV) function. LV catheterization in mice undergoing coronary artery occlusion serves as a novel method for a real-time evaluation of LV function.

Análise em tempo real de pressão-volume de infarto agudo do miocárdio em ratos

Acute myocardial infarction can lead to acute heart failure and cardiogenic shock. The evaluation of hemodynamics is critical for the evaluation of any ...

Echocardiographic Measurement of Right Ventricular Diastolic Parameters in Mouse

Diastolic dysfunction is a prominent feature of right ventricular (RV) remodeling associated with conditions of pressure overload. However, the RV diastolic function is rarely quantified in experimental studies. This might be due to technical difficulties in the visualization of the RV in the apical four-chamber view in rodents. Here we describe two positions facilitating the visualization of the apical four-chamber view in mice to assess the RV diastolic function.
The apical four-chamber view is enabled by tilting the mouse fixation platform to the left and caudally (LeCa) or to the right and cranially (RiCr). Both positions provide images of comparable quality. The results of the RV diastolic function obtained from two positions are not significantly different. Both positions are comparably easy to perform. This protocol can be incorporated into published protocols and enables detailed investigations of the RV function.

Here we describe and compare two positions for obtaining the apical four-chamber view in mice. These positions enable the quantification of the right ventricular function, provide comparable results, and can be used interchangeably.

Mensuração ecocardiográfica dos parâmetros diastólicos do ventrículo direito no mouse

Diastolic dysfunction is a prominent feature of right ventricular (RV) remodeling associated with conditions of pressure overload. However, the RV ...

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 ...

Biventricular Assessment of Cardiac Function and Pressure-Volume Loops by Closed-Chest Catheterization in Mice

Assessment of cardiac function is essential to conduct cardiovascular and pulmonary-vascular preclinical research. Pressure-volume loops (PV loops) generated by recording both pressure and volume during cardiac catheterization are vital when assessing both systolic and diastolic cardiac function. Left and right heart function are closely related, reflected in ventricular interdependence. Thus, recording biventricular function in the same animal is important to get a complete assessment of cardiac function. In this protocol, a closed chest approach to cardiac catheterization consistent with the way catheterization is performed in patients is adopted in mice. While challenging, the closed chest strategy is a more physiological approach, because opening the chest results in major changes in preload and afterload that create artifacts, most notably a fall in systemic blood pressure. While high-resolution echocardiography is used to assess rodents, cardiac catheterization is invaluable, particularly when assessing diastolic pressures in both ventricles.
Described here is a procedure to perform invasive, closed chest, sequential left and right ventricular pressure-volume (PV) loops in the same animal. PV loops are acquired using&#160;admittance technology with a mouse pressure-volume catheter and pressure-volume system acquisition. The procedure is described, beginning with the neck dissection, which is required to access the right jugular vein and the right carotid artery, to the insertion and positioning of the catheter, and finally the data acquisition. Then, the criteria required to ensure the acquisition of high-quality PV loops are discussed. Finally, the analysis of the left and right ventricular PV loops and the different hemodynamic parameters available to quantify systolic and diastolic ventricular function are briefly described.

Presented here is a protocol to assess biventricular heart function in mice by generating pressure-volume (PV) loops from the right and left ventricle in the same animal using closed chest catheterization. The focus is on the technical aspect of surgery and data acquisition.

Avaliação Biventricular da Função Cardíaca e Alças Pressão-Volume por Cateterismo Fechado de Tórax em Camundongos

Assessment of cardiac function is essential to conduct cardiovascular and pulmonary-vascular preclinical research. Pressure-volume loops (PV loops) ...