Name: intrauterine telemetry to measure mouse contractile pressure in vivo
Uploaded: 2015-04-06T15:00:00.000+00:00
Duration: 7 min 3 s
Description: Watch this Scientific Journal Video about Intrauterine Telemetry to Measure Mouse Contractile Pressure In Vivo at JoVE.com

This work was supported by funding from the March of Dimes FY12-133 (S.K.E.) and the National Institutes of Health R01HD037831 (S.K.E.). We would like to thank Dr. Deborah Frank for critical review of the manuscript.

NOTA: Todos los animales procedimientos conforma a las directrices para el cuidado y uso de animales establecidos por los Institutos Nacionales de Salud. Todos los protocolos fueron aprobados por la Comisión del Estudio de animales en la Universidad de Washington en St. Louis. NOTA: Los materiales y equipos específicos para este protocolo se enumeran Materiales y Equipos mesa. 1. inseminación a tiempo fijo de ratones hembra <ol><li> Raza hembras adultas entre dos y seis meses de edad en una ventana de tiempo de 2 horas con los varones. Hr específico de cría no son necesarios, siempre y cuando se mantengan consistentes para la totalidad del experimento. </li><li> Confirmar el embarazo por la presencia de un tapón de copulador. Designar el día y la hora en la que el tapón se ve por primera vez como días post-coito 0 (dpc0). Recuerde la hora específica cuando se inicia la grabación de la presión. </li><li> Realizar cirugías en dpc8-13. El DPC exacta dependerá de la cepa de ratón. </li><li> Para determinar si un ratón está embarazada antes de la cirugía, peimágenes de ultrasonido rforma del abdomen con una sonda de matriz lineal 40 MHz acoplado a un sistema de formación de imágenes antes de administrar la anestesia. Esto permitirá la visualización de sacos gestacionales. Si un sistema de ultrasonidos no está disponible, utilice el peso del ratón. NOTA: Un aumento de peso de al menos 2 g entre dpc0 y dpc8 y distensión del abdomen es un buen indicador de embarazo. </li></ol> 2. Telemetría Cirugía Prep <ol><li> Utilice un autoclave para esterilizar los instrumentos, gasas, torundas de algodón, paños quirúrgicos, y puntas de pipeta dentro de 24 horas antes de la cirugía. </li><li> Superficie cirugía limpia con etanol al 70%. </li><li> Encienda el cojín eléctrico (especializado para roedores) y esterilizador de cuentas. </li></ol> 3. Telemetría Cirugía <ol><li> Para la anestesia, utilizar 4-5% de isoflurano con una tasa de flujo de oxígeno de 1-2 L / min. </li><li> Espere hasta que el ratón muestra signos de anestesia y luego inyectar por vía subcutánea con buprenorfina (analgésico: 0,1 ml por 20 g). </li><li> Afeitar el abdomen del ratón desde la parte inferior de las costillas hacia abajo a la vejiga. </li><li> Traiga el ratón a la mesa de cirugía en el cojín de calefacción pre-calentado, y cubrir los ojos del ratón con pomada oftálmica para evitar el secado. </li><li> Comience técnica estéril. Póngase los guantes estériles y mantener la envoltura como una superficie estéril en el que colocar los matorrales de yodo y etanol. </li><li> Retire el telémetro del paquete o solución salina y utilizar pinzas estériles para colocarlo sobre una gasa estéril. <ol><li> Utilice la jeringa (proporcionado por el proveedor telémetro) para llenar la punta del catéter con gel. Apriete suavemente el catéter crear suficiente presión negativa para extraer el gel en el catéter. </li></ol></li><li> Coloque una punta estéril sobre la pipeta. Pipetear 2 l de pegamento quirúrgico y establecer la pipeta sobre una superficie estéril. </li><li> Para limpiar el sitio quirúrgico, limpie el área con yodo y luego con etanol al 70%. </li><li> Cortar un agujero en la sábana quirúrgica lo suficientemente grandepara el área necesaria para la incisión y el lugar sobre el ratón. </li><li> Realice una pizca pie (la respuesta del pedal) para asegurar que el ratón está completamente anestesiado antes de hacer una incisión. </li><li> Hacer una pequeña incisión en la línea media vertical. A continuación, hacer una incisión similar a través de la pared muscular del cuerpo subyacente. </li><li> Tire suavemente del útero de la cavidad del cuerpo y localice el cuerno uterino que contiene el mayor número de crías que sean viables. <ol><li> Opcional - Para añadir un miRNA, inyecte vector viral en la capa muscular del útero antes de la inserción del telémetro. </li></ol></li><li> Hacer una pequeña incisión en la punta del cuerno uterino con 3 mm de corte tijeras de primavera de borde. </li><li> Pase el catéter, sin apretar, entre la pared uterina y sacos fetales. Inserte el catéter pasado varios sacos de reducir su eliminación accidental. Asegúrese de que los sacos fetales no se vean perturbadas por la inserción. </li><li> Pipetear el pegamento quirúrgico para el sitio de inserción telémetro para adherir el catéter a la ucuerno terine y evitar que el catéter se deslice fuera del cuerno uterino. Espere unos segundos para que el pegamento se vuelven rígidas y coloque cuidadosamente el útero hacia el interior de la cavidad corporal. </li><li> Coloque el telémetro en la cavidad del cuerpo en el lado opuesto del sitio de inserción del catéter. </li><li> Suturar la cavidad del cuerpo se cerró con 6,0 suturas absorbibles. En primer lugar, hacer una puntada bolso y luego tres y cincuenta y siete puntadas individuales. </li><li> Suturar la piel mediante el uso de la misma técnica, pero con 6-0 sutura no absorbible. </li><li> Dibucaina Swab (analgésico tópico) ungüento sobre el lugar de la incisión. </li><li> Inyectar 0,3 a 0,4 ml de solución salina estéril por vía subcutánea para ayudar a rehidratar al animal. </li><li> Monitorear el animal después de la cirugía hasta que esté completamente despierto. No coloque el ratón de nuevo en su jaula hasta que sea capaz de sostener libremente su cabeza por encima de la mesa, como ciertos tipos de ropa de cama pueden sofocar un ratón muy anestesiado. Para un ratón que ha sido anestesiado durante 30-45 min, uno debería esperar ªe ratón para ser completamente despierto por 1 hora y 15 min. </li><li> Si la realización de cirugías adicionales, limpie instrumentos con etanol al 70% y el talón esterilizar antes de su reutilización. </li></ol> 4. Después de la cirugía de recuperación <ol><li> Supervisar diariamente el progreso después de la cirugía en un registro. Catálogo aumento de peso y la pérdida y el reloj para letargo, sangrado o cierre incompleto de la incisión quirúrgica. <ol><li> Alimente el ratón chow suavizado ratón (~ la consistencia de puré de manzana) o especializada alimento de la dieta de recuperación. </li><li> Examinar las suturas y asegurarse de que no se sacan, pero trate de no manejar el ratón en exceso. </li></ol></li></ol> 5. La telemetría de grabación <ol><li> En el día de interés (por lo menos de dos a tres días después el ratón se ha recuperado de la cirugía), colocar un imán cerca del ratón para encender el telémetro. Coloque la jaula del ratón en el receptor. La luz en el receptor debería encenderse si se activa el telémetro. </li><li> Calibrar nueva Teleme marca<html> tros con calibraciones específicas del fabricante siempre. Especificaciones de calibración telémetro reutilizados serán guardados en el software. </li><li> Dentro del software, iniciar la grabación haciendo clic derecho sobre &quot;animal&quot; y haga clic en &quot;iniciar el muestreo, continua&quot;, que permite el muestreo a una velocidad de hasta 500 Hz. Asegúrese de que el software está configurado para guardar y rastrear. NOTA: Cuando el icono de muestreo del ratón cambia a verde, se están grabando las presiones. Perturbar el ratón durante el parto puede interrumpir o alterar el trabajo, así que observar con cautela. </li><li> Después del parto o periodo de tiempo de interés, detenga la grabación haciendo clic derecho sobre &quot;animal&quot; y seleccionando &quot;detener las grabaciones, todo&quot;. Utilice un imán para apagar telémetro para salvar la vida de la batería, ya que la batería telémetro normalmente sólo tendrá una duración de 1,5 meses de grabación continua. </li><li> La eutanasia el ratón por CO 2 sobredosis y recuperar el telémetro. Los cachorros son sacrificados por decapitación. </li></ol>tle &quot;&gt; 6. telémetro Esterilización <ol><li> Coloque el telémetro en una solución de 1% de enzima detergente durante la noche en agua desionizada a temperatura ambiente. </li><li> Enjuague el telémetro en agua desionizada y eliminar todo el tejido y el pegamento, teniendo cuidado de no apretar el catéter. </li><li> Incubar el telémetro en glutaraldehído al 2% durante 24 horas a temperatura ambiente para esterilizar. </li><li> Enjuague el telémetro con solución salina estéril y almacenarlo en una solución salina estéril hasta la próxima implantación o para ser enviado de nuevo a fabricante a ser reformado. Telémetros deben ser reformadas si el tejido se encuentra en el interior del catéter después de la esterilización. Normalmente telémetros se pueden utilizar 4-5 veces antes de necesitar ser renovado. </li></ol>

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	<li>Slattery, M. M., Morrison, J. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preterm+delivery.">Preterm delivery.</a> Lancet. 360, 1489-1497 (2002).</li><li>Goldenberg, R. L., Culhane, J. F., Iams, J. D., Romero, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epidemiology+and+causes+of+preterm+birth.">Epidemiology and causes of preterm birth.</a> Lancet. 371, 75-84 (2008).</li><li>Keelan, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pharmacological+inhibition+of+inflammatory+pathways+for+the+prevention+of+preterm+birth.">Pharmacological inhibition of inflammatory pathways for the prevention of preterm birth.</a> Journal Of Reproductive Immunology. 88, 176-184 (2011).</li><li>Voltolini, C., 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=Understanding+spontaneous+preterm+birth:+from+underlying+mechanisms+to+predictive+and+preventive+interventions.">Understanding spontaneous preterm birth: from underlying mechanisms to predictive and preventive interventions.</a> Reproductive Sciences. 20, 1274-1292 (2013).</li><li>Ratajczak, C. K., Muglia, L. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Insights+into+parturition+biology+from+genetically+altered+mice.">Insights into parturition biology from genetically altered mice.</a> Pediatric research. 64, 581-589 (2008).</li><li>Rajakumar, 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=Placental+HIF-1+alpha,+HIF-2+alpha,+membrane+and+soluble+VEGF+receptor-1+proteins+are+not+increased+in+normotensive+pregnancies+complicated+by+late-onset+intrauterine+growth+restriction.">Placental HIF-1 alpha, HIF-2 alpha, membrane and soluble VEGF receptor-1 proteins are not increased in normotensive pregnancies complicated by late-onset intrauterine growth restriction.</a> American Journal Of Physiology. Regulatory, Integrative And Comparative Physiology. 293, (2007).</li><li>Whitesall, S. E., Hoff, J. B., Vollmer, A. P., D'Alecy, L. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+simultaneous+measurement+of+mouse+systolic+arterial+blood+pressure+by+radiotelemetry+and+tail-cuff+methods.+American+journal+of+physiology.">Comparison of simultaneous measurement of mouse systolic arterial blood pressure by radiotelemetry and tail-cuff methods. American journal of physiology.</a> Heart And Circulatory Physiology. 286, H2408-H2415 (2004).</li><li>Mackay, L. B., Shi, L. B., Maul, H. .., Maner, W. L., Garfield, R. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+bilateral+pelvic+neurectomy+on+cervical+ripening+in+pregnant+rats.">The effect of bilateral pelvic neurectomy on cervical ripening in pregnant rats.</a> Journal Of Perinatal Medicine. 37, 263-269 (2009).</li><li>Pierce, S. L., Kutschke, W., Cabeza, R., England, S. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+measurement+of+intrauterine+pressure+by+telemetry:+a+new+approach+for+studying+parturition+in+mouse+models.">In vivo measurement of intrauterine pressure by telemetry: a new approach for studying parturition in mouse models.</a> Physiol Genomics. 42, 310-316 (2010).</li><li>Imamura, T., Luedke, C. E., Vogt, S. K., Muglia, L. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oxytocin+modulates+the+onset+of+murine+parturition+by+competing+ovarian+and+uterine+effects.">Oxytocin modulates the onset of murine parturition by competing ovarian and uterine effects.</a> American Journal Of Physiology. Regulatory, Integrative And Comparative Physiology. 279, R1061-R1067 (2000).</li><li>McCloskey, C., 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=The+inwardly+rectifying+K++channel+KIR7.1+controls+uterine+excitability+throughout+pregnancy.">The inwardly rectifying K+ channel KIR7.1 controls uterine excitability throughout pregnancy.</a> EMBO Mol Med. 6 (9), 1161-1174 (2014).</li></ol>

The authors have nothing to disclose.

Las investigaciones de la contractilidad del miometrio se han basado en las mediciones in vivo ex de la tensión muscular. Esta metodología puede ser útil para realizar pruebas en etapa temprana de fármacos uterotónicos recientemente desarrollados que no pueden ser administrados a los animales vivos. Sin embargo, los enfoques in vivo son necesarios para comprender longitudinalmente la progresión de la mano de obra. Las mediciones in vivo sirven para varios propósitos. En primer lugar, pe...

El nacimiento prematuro es la principal causa de morbilidad y mortalidad perinatal en los países desarrollados; que es responsable del 50% de la morbilidad perinatal y el 75% de los 1,2 mortalidad perinatal. El parto prematuro es multifacética y puede ser idiopática. Aunque muchas investigaciones han surgido en las vías moleculares y eléctricas que transforman el miometrio de un tejido quiescente en un contráctil uno, la fisiopatología exacta del trabajo de parto prematuro sigue siendo difícil de alcanzar. Endocrino, inflamatoria, y la regulación de genes han sido relacionados con el parto prematuro 3,4. Sin embargo, las preocupaciones éticas limitan la capacidad para llevar a cabo la investigación sobre los mecanismos de trabajo de parto prematuro en los seres humanos. Dadas estas limitaciones, muchos investigadores han recurrido a los ratones como sistema modelo con el que el estudio de la fisiología del parto. Los ratones tienen gestaciones corta duración aproximadamente tres semanas y pueden ser fácilmente manipulable genéticamente. Además, varios modelos genéticos de ratón tienenha desarrollado para determinar las vías de señalización que son esenciales para el trabajo 5. A pesar de las diferencias clave entre el ratón y el parto humano, los ratones y los humanos comparten muchos de los mismos mecanismos que son esenciales para el trabajo, incluyendo la inflamación y la infección intrauterina 6. De este modo, los ratones sirven como una valiosa herramienta para examinar la actividad uterina. Hasta la fecha, el estándar de oro para medir la contractilidad uterina en ratones ha sido ex vivo grabaciones tensión isométrica; sin embargo, esto está limitado a un punto de tiempo gestacional por experimento y requiere la extirpación del útero a partir de su entorno fisiológico. Otro factor importante es el gran número de animales necesarios para estas investigaciones. Por último, esta metodología no permite estudios longitudinales que examinan la inducción del trabajo de parto y parto prematuro fisiopatología. Los recientes avances realizados en los dispositivos de telemetría radiométricas utilizadas para estudiar la presión arterial cambia 7 en ratones y intpresión rauterine en ratas 8 plantea la cuestión de si la misma tecnología podría usarse para estudiar los cambios en la presión intrauterina en ratones durante el embarazo. Después de la resolución de problemas inicial, se desarrolló un método con éxito para medir la inducción del parto y la progresión en un ratón. Este enfoque in vivo puede medir la transición desde el estado de baja presión del útero de reposo al estado de alta presión indicativa de fuertes contracciones del parto. Este método también ha sido capaz de detectar diferencias de presión significativas entre los ratones transgénicos con el parto comprometido y ratones de tipo salvaje, lo que demuestra el impacto fisiológico de la expresión génica 9. Este protocolo proporciona un tiempo real, en el método in vivo para estudiar la presión uterina ratón durante el embarazo.

<table><tbody><tr><td>Surgical Glue</td><td>3M Vetbond</td><td>1469SB</td><td>Tissue Adhesive--stored in Drierite</td></tr><tr><td>Absorbable 6-0 Sutures</td><td>Ethicon</td><td>J212H</td><td>Vicryl</td></tr><tr><td>Non-absorbable 6-0 Sutures</td><td>Ethicon</td><td>8706H</td><td>Prolene</td></tr><tr><td>Water Jacket Blanket + Heating Pad</td><td>Gaymer</td><td>T/Pump PN 11184-000 Blanket-66N111CC</td><td>Specialized for rodents</td></tr><tr><td>Bead sterilizer</td><td>Keller</td><td>Z378577</td><td>Steri 250 Sterilizer</td></tr><tr><td>Disecting Microscope</td><td>Nikon</td><td>SMZ754</td><td>Fibre optic gooseneck external light source</td></tr><tr><td>Sterile Surgical Gloves--Latex&amp;nbsp;</td><td>Cardinal Health Triflex</td><td>2D7253</td><td></td></tr><tr><td>PhysioTel PA-C10 Pressure Transmitter</td><td>&amp;nbsp;Data Sciences International</td><td>270-0135-001</td><td>TA11PA-C10</td></tr><tr><td>Telemeter Reciever</td><td>&amp;nbsp;Data Sciences International</td><td>272-6001-001</td><td>RPC-1</td></tr><tr><td>Dataquest ART 4.3.2 Analysis Platinum</td><td>&amp;nbsp;Data Sciences International</td><td>271-0147-141</td><td>Analysis software</td></tr><tr><td>Diet Recovery Gel</td><td>Clear H2O</td><td>72-01-5022</td><td>Purified Soft Diet for Rodents</td></tr><tr><td>Tergazyme</td><td>Sigma-Aldrich</td><td>Z273287</td><td>Enzyme detergent</td></tr></tbody></table>

intrauterine telemetry to measure mouse contractile pressure in vivo

Se necesita una integración compleja de señales moleculares y eléctricos para transformar un útero en reposo en un órgano contráctil al final del embarazo. A pesar del descubrimiento de los principales reguladores de la contractilidad uterina, este proceso todavía no se comprende totalmente. Los ratones transgénicos proporcionan un modelo ideal en el que estudiar el parto. Anteriormente, el único método para estudiar la contractilidad uterina en el ratón era ex vivo grabaciones de tensión isométrica, que son subóptima por varias razones. El útero debe ser retirado de su entorno fisiológico, un curso de tiempo limitado de investigación es posible, y los ratones debe ser sacrificada. El reciente desarrollo de telemetría radiométrica ha permitido, mediciones en tiempo real longitudinales de la presión intrauterina in vivo en ratones. Aquí, la implantación de un telémetro intrauterino para medir los cambios de presión en el útero de ratón a partir de la mitad del embarazo hasta que se describe entrega. Mediante la comparación de las diferencias en la prensaUres entre tipo salvaje y ratones transgénicos, el impacto fisiológico de un gen de interés puede ser dilucidado. Esta técnica debería acelerar el desarrollo de terapias que se utilizan para tratar los trastornos del miometrio durante el embarazo, incluyendo el parto prematuro.

En tiempo real de las presiones intrauterinas in vivo se puede grabar mediante el uso de un sistema de adquisición de telemetría. Muestreo de presión junto con la grabación de vídeo simultánea del ratón se utiliza para capturar el momento exacto de la entrega de cada cachorro y correlacionar el tiempo de entrega a la presión intrauterina de la madre. Todos los puntos de datos pueden entonces ser trazados a lo largo de la grabación para generar una parcela (Figura 1A) de presi?...

Watch this Scientific Journal Video about Intrauterine Telemetry to Measure Mouse Contractile Pressure In Vivo at JoVE.com

Intrauterine Telemetry to Measure Mouse Contractile Pressure In Vivo

This manuscript provides a protocol for implanting telemeters in the mouse for the purpose of measuring intrauterine pressures during pregnancy.

Telemetría intrauterina para medir la presión del ratón contráctil<em&gt; En Vivo</em

A complex integration of molecular and electrical signals is needed to transform a quiescent uterus into a contractile organ at the end of pregnancy. ...

intrauterine-telemetry-to-measure-mouse-contractile-pressure-in-vivo

Research

JoVE Journal

Medicine

Intrauterine Telemetry to Measure Mouse Contractile Pressure In Vivo

9.6K vistas.  Washington University in St. Louis. This manuscript provides a protocol for implanting telemeters in the mouse for the purpose of measuring intrauterine pressures during pregnancy. 

Video: Intrauterine Telemetry to Measure Mouse Contractile Pressure In Vivo

Measuring Left Ventricular Pressure in Late Embryonic and Neonatal Mice

Blood pressure increases significantly during embryonic and postnatal development in vertebrate animals. In the mouse, blood flow is first detectable around embryonic day (E) 8.51. Systolic left ventricular (LV) pressure is 2 mmHg at E9.5 and 11 mmHg at E14.52. At these mid-embryonic stages, the LV is clearly visible through the chest wall for invasive pressure measurements because the ribs and skin are not fully developed. Between E14.5 and birth (approximately E21) imaging methods must be used to view the LV. After birth, mean arterial pressure increases from 30 - 70 mmHg from postnatal day (P) 2 - 353. Beyond P20, arterial pressure can be measured with solid-state catheters (i.e. Millar or Scisense). Before P20, these catheters are too big for developing mouse arteries and arterial pressure must be measured with custom pulled plastic catheters attached to fluid-filled pressure transducers3 or glass micropipettes attached to servo null pressure transducers4.
Our recent work has shown that the greatest increase in blood pressure occurs during the late embryonic to early postnatal period in mice5-7. This large increase in blood pressure may influence smooth muscle cell (SMC) phenotype in developing arteries and trigger important mechanotransduction events. In human disease, where the mechanical properties of developing arteries are compromised by defects in extracellular matrix proteins (i.e. Marfan's Syndrome8 and Supravalvular Aortic Stenosis9) the rapid changes in blood pressure during this period may contribute to disease phenotype and severity through alterations in mechanotransduction signals. Therefore, it is important to be able to measure blood pressure changes during late embryonic and neonatal periods in mouse models of human disease.
We describe a method for measuring LV pressure in late embryonic (E18) and early postnatal (P1 - 20) mice. A needle attached to a fluid-filled pressure transducer is inserted into the LV under ultrasound guidance. Care is taken to maintain normal cardiac function during the experimental protocol, especially for the embryonic mice. Representative data are presented and limitations of the protocol are discussed.

Measuring left ventricular pressure (LV) in embryonic and neonatal mice is described. Pressure is measured by inserting a needle connected to a fluid-filled transducer into the LV under ultrasound guidance. Care must be taken to maintain normal cardiac function during the experimental protocol.

Medición de la presión del ventrículo izquierdo a finales de embriones de ratones y Neonatal

Blood pressure increases significantly during embryonic and postnatal development in vertebrate animals. In the mouse, blood flow is first detectable ...

Investigación

Bioingeniería

Implantation of Radiotelemetry Transmitters Yielding Data on ECG, Heart Rate, Core Body Temperature and Activity in Free-moving Laboratory Mice

The laboratory mouse is the animal species of choice for most biomedical research, in both the academic sphere and the pharmaceutical industry. Mice are a manageable size and relatively easy to house. These factors, together with the availability of a wealth of spontaneous and experimentally induced mutants, make laboratory mice ideally suited to a wide variety of research areas.
In cardiovascular, pharmacological and toxicological research, accurate measurement of parameters relating to the circulatory system of laboratory animals is often required. Determination of heart rate, heart rate variability, and duration of PQ and QT intervals are based on electrocardiogram (ECG) recordings. However, obtaining reliable ECG curves as well as physiological data such as core body temperature in mice can be difficult using conventional measurement techniques, which require connecting sensors and lead wires to a restrained, tethered, or even anaesthetized animal. Data obtained in this fashion must be interpreted with caution, as it is well known that restraining and anesthesia can have a major artifactual influence on physiological parameters1, 2.
Radiotelemetry enables data to be collected from conscious and untethered animals. Measurements can be conducted even in freely moving animals, and without requiring the investigator to be in the proximity of the animal. Thus, known sources of artifacts are avoided, and accurate and reliable measurements are assured. This methodology also reduces interanimal variability, thus reducing the number of animals used, rendering this technology the most humane method of monitoring physiological parameters in laboratory animals3, 4. Constant advancements in data acquisition technology and implant miniaturization mean that it is now possible to record physiological parameters and locomotor activity continuously and in realtime over longer periods such as hours, days or even weeks3, 5.
Here, we describe a surgical technique for implantation of a commercially available telemetry transmitter used for continuous measurements of core body temperature, locomotor activity and biopotential (i.e. onelead ECG), from which heart rate, heart rate variability, and PQ and QT intervals can be established in freeroaming, untethered mice. We also present pre-operative procedures and protocols for post-operative intensive care and pain treatment that improve recovery, well-being and survival rates in implanted mice5, 6.

A surgical technique for implantation of commercially available telemetry transmitters used for continuous measurement of biopotential (one-lead ECG), heart rate, core body temperature and locomotor activity in freely moving mice is shown. Recommendations and protocols for post-operative care and pain relief, improving recovery, well being and survival rate are also presented.

Implantación de transmisores de radiotelemetría Ceder los datos en el ECG, frecuencia cardiaca, la temperatura corporal y la actividad en ratones de laboratorio de movimiento libre

The laboratory mouse is the animal species of choice for most biomedical research, in both the academic sphere and the pharmaceutical industry. Mice are a ...

Medicina

In utero Measurement of Heart Rate in Mouse by Noninvasive M-mode Echocardiography

Congenital heart disease (CHD) is the most frequent noninfectious cause of death at birth. The incidence of CHD ranges from 4 to 50/1,000 births (Disease and injury regional estimates, World Health Organization, 2004). Surgeries that often compromise the quality of life are required to correct heart defects, reminding us of the importance of finding the causes of CHD. Mutant mouse models and live imaging technology have become essential tools to study the etiology of this disease. Although advanced methods allow live imaging of abnormal hearts in embryos, the physiological and hemodynamic states of the latter are often compromised due to surgical and/or lengthy procedures. Noninvasive ultrasound imaging, however, can be used without surgically exposing the embryos, thereby maintaining their physiology. Herein, we use simple M-mode ultrasound to assess heart rates of embryos at E18.5 in utero. The detection of abnormal heart rates is indeed a good indicator of dysfunction of the heart and thus constitutes a first step in the identification of developmental defects that may lead to heart failure.

In utero Measurement of Heart Rate in Mouse by Noninvasive M-mode Echocardiography

Ultra-high frequency ultrasound is a powerful live imaging tool to examine cardiac abnormalities in small animals. Its noninvasiveness allows maintaining the physiologic state of embryos. Herein, we show the use of M-mode ultrasound to measure the heart rates of embryos at E18.5 in utero.

En el útero de medición de la frecuencia cardíaca en el ratón por no invasiva Ecocardiografía en modo M

Congenital heart disease (CHD) is the most frequent noninfectious cause of death at birth. The incidence of CHD ranges from 4 to 50/1,000 births (Disease ...

Mouse Maternal Autonomic Changes from Pregnancy to Pup-Rearing

Analysis of Electrocardiograms and Behavior in Mice from Pregnancy to Lactation Period

Changes in the mother-offspring relationship are presumably accompanied by dynamic changes in the autonomic nervous system. Although temporal measurements of autonomic activity have been performed in human mothers and infants, the analysis of long-term changes remains unexplored. Mouse mothers can form social bonds with their pups and have a short period of pregnancy and lactation, which makes them useful for the examination of physiological changes from pregnancy to pup-rearing. Therefore, a telemetry system was used for several weeks to measure the changes in the autonomic nervous system and the behavior of mouse mothers. The current results showed that an electrocardiogram (ECG) could be stably recorded regardless of the movements of mothers and parturition. ECG analysis showed that the heart rate gradually decreased from pregnancy to lactation, and sympathetic activity sharply increased as the pups developed. Furthermore, the simultaneous recording of behavior and ECG in the home cage enabled us to understand the behavior-dependent influences on the ECG, thereby revealing the characteristics of autonomic nervous activity during each behavior. Thus, the present experimental method helps to understand how the physiological characteristics of mothers change from pregnancy through pup rearing, supporting the healthy development of pups.

A simultaneous recording of autonomic activity and detailed maternal behavior of mother mice from pregnancy to lactation was achieved using a telemetry system. This method helps to understand the dynamics of the physiological and behavioral characteristics in mothers from pregnancy to weaning.

Changes in the mother-offspring relationship are presumably accompanied by dynamic changes in the autonomic nervous system. Although temporal measurements ...

Comportamiento

Long-term Blood Pressure Measurement in Freely Moving Mice Using Telemetry

During the development of new vasoactive agents, arterial blood pressure monitoring is crucial for evaluating the efficacy of the new proposed drugs. Indeed, research focusing on the discovery of new potential therapeutic targets using genetically altered mice requires a reliable, long-term assessment of the systemic arterial pressure variation. Currently, the gold standard for obtaining long-term measurements of blood pressure in ambulatory mice uses implantable radio-transmitters, which require&#160;artery cannulation. This technique eliminates the need for tethering, restraining, or anesthetizing the animals which introduce stress and artifacts during data sampling. However, arterial blood pressure monitoring in mice via catheterization can be rather challenging due to the small size of the arteries. Here we present a step-by-step guide to illustrate the crucial key passages for a successful subcutaneous implantation of radio-transmitters and carotid artery cannulation in mice. We also include examples of long-term blood pressure activity taken from freely moving mice after a period of post-surgery recovery. Following this procedure will allow reliable direct blood pressure recordings from multiple animals simultaneously.

The goal of this protocol is to assess systemic blood pressure in conscious freely moving mice using implantable radio-telemetry devices.

A largo plazo de medida de la presión arterial en ratones con libertad de movimientos mediante telemetría

During the development of new vasoactive agents, arterial blood pressure monitoring is crucial for evaluating the efficacy of the new proposed drugs. ...

Application of Consistent Massage-Like Perturbations on Mouse Calves and Monitoring the Resulting Intramuscular Pressure Changes

Massage is generally recognized to be beneficial for relieving pain and inflammation. Although previous studies have reported anti-inflammatory effects of massage on skeletal muscles, the molecular mechanisms behind are poorly understood. We have recently developed a simple device to apply local cyclical compression (LCC), which can generate intramuscular pressure waves with varying amplitudes. Using this device, we have demonstrated that LCC modulates inflammatory responses of macrophages in situ and alleviates immobilization-induced muscle atrophy. Here, we describe protocols for the optimization and application of LCC as a massage-like intervention against immobilization-induced inflammation and atrophy of skeletal muscles of mouse hindlimbs. The protocol that we have developed can be useful for investigating the mechanism underlying beneficial effects of physical exercise and massage. Our experimental system provides a prototype of the analytical approach to elucidate the mechanical regulation of muscle homeostasis, although further development needs to be made for more comprehensive studies.

Here we describe the protocols for applying defined mechanical loads to mouse calves and for monitoring the concomitant intramuscular pressure changes. The experimental systems that we have developed can be useful for investigating the mechanism behind the beneficial effects of physical exercise and massage.

Aplicación de Perturbaciones Consistentes Similares al Masaje en Los terneros de ratón y Monitoreo de los Cambios de Presión Intramuscular Resultantes

Massage is generally recognized to be beneficial for relieving pain and inflammation. Although previous studies have reported anti-inflammatory effects of ...

Inmunología e Infección

Implantation of Combined Telemetric ECG and Blood Pressure Transmitters to Determine Spontaneous Baroreflex Sensitivity in Conscious Mice

Blood pressure (BP) and heart rate (HR) are both controlled by the autonomic nervous system (ANS) and are closely intertwined due to reflex mechanisms. The baroreflex is a key homeostatic mechanism to counteract acute, short-term changes in arterial BP and to maintain BP in a relatively narrow physiological range. BP is sensed by baroreceptors located in the aortic arch and carotid sinus. When BP changes, signals are transmitted to the central nervous system and are then communicated to the parasympathetic and sympathetic branches of the autonomic nervous system to adjust HR. A rise in BP causes a reflex decrease in HR, a drop in BP causes a reflex increase in HR.
Baroreflex sensitivity (BRS) is the quantitative relationship between changes in arterial BP and corresponding changes in HR. Cardiovascular diseases are often associated with impaired baroreflex function. In various studies reduced BRS has been reported in e.g., heart failure, myocardial infarction, or coronary artery disease. 
Determination of BRS requires information from both BP and HR, which can be recorded simultaneously using telemetric devices. The surgical procedure is described beginning with the insertion of the pressure sensor into the left carotid artery and positioning of its tip in the aortic arch to monitor arterial pressure followed by the subcutaneous placement of the transmitter and ECG electrodes. We also describe postoperative intensive care and analgesic management. After a two-week period of post-surgery recovery long-term ECG and BP recordings are performed in conscious and unrestrained mice. Finally, we include examples of high-quality recordings and the analysis of spontaneous baroreceptor sensitivity using the sequence method.

The baroreflex is a heart-rate regulation mechanism by the autonomic nervous system in response to blood-pressure changes. We describe a surgical technique to implant telemetry transmitters for continuous and simultaneous measurement of electrocardiogram and blood pressure in mice. This can determine spontaneous baroreflex sensitivity, an important prognostic marker for cardiovascular disease.

Implantación de ECG telemétrico combinado y transmisores de presión arterial para determinar la sensibilidad barorrefleja espontánea en ratones conscientes

Blood pressure (BP) and heart rate (HR) are both controlled by the autonomic nervous system (ANS) and are closely intertwined due to reflex mechanisms. ...

Real-Time Electrocardiogram Monitoring During Treadmill Training in Mice

Regular physical exercise is a major contributor to cardiovascular health, influencing various metabolic as well as electrophysiological processes. However, in certain cardiac diseases such as inherited arrhythmia syndromes, e.g., arrhythmogenic cardiomyopathy (ACM) or myocarditis, physical exercise may have negative effects on the heart leading to a proarrhythmogenic substrate production. Currently, the underlying molecular mechanisms of exercise-related proarrhythmogenic remodeling are largely unknown, thus it remains unclear which frequency, duration, and intensity of exercise can be considered safe in the context of disease(s).
The proposed method allows to study proarrhythmic/antiarrhythmic effects of physical exercise by combining treadmill training with real-time monitoring of the ECG. Implantable telemetry devices are used to continuously record the ECG of freely moving mice over a period of up to 3 months both at rest and during treadmill training. Data acquisition software with its analysis modules is used to analyze basic ECG parameters such as heart rate, P wave duration, PR interval, QRS interval, or QT duration at rest, during and after training. Furthermore, heart rate variability (HRV) parameters and occurrence of arrhythmias are evaluated. In brief, this manuscript describes a step-by-step approach to experimentally explore exercise induced effects on cardiac electrophysiology, including potential proarrhythmogenic remodeling in mouse models.

Electrocardiogram (ECG) is the key variable to understanding cardiac electrophysiology. Physical exercise has beneficial effects but may also be harmful in the context of cardiovascular diseases. This manuscript provides a method of recording real-time ECG during exercise, which can serve to investigate its effects on cardiac electrophysiology in mice.

Monitoreo de electrocardiograma en tiempo real durante el entrenamiento en cinta rodante en ratones

Regular physical exercise is a major contributor to cardiovascular health, influencing various metabolic as well as electrophysiological processes. ...

Ex Vivo Method for Assessing the Mouse Reproductive Tract Spontaneous Motility and a MATLAB-based Uterus Motion Tracking Algorithm for Data Analysis

Dysmenorrhea, or painful cramping, is the most common symptom associated with menses in females and its severity can hinder women&#39;s everyday lives. Here, we present an easy and inexpensive method that would be instrumental for testing new drugs decreasing uterine contractility. This method utilizes the unique ability of the entire mouse reproductive tract to exhibit spontaneous motility when maintained ex vivo in a Petri dish containing oxygenated Krebs buffer. This spontaneous motility resembles the wave-like myometrial activity of the human uterus, referred to as endometrial waves. To demonstrate the effectiveness of the method, we employed a well-known uterine relaxant drug, epinephrine. We demonstrate that the spontaneous motility of the entire mouse reproductive tract can be quickly and reversibly inhibited by 1 &#181;M epinephrine in this Petri dish model. Documenting the changes of uterine motility can be easily done using an ordinary smart phone or a sophisticated digital camera. We developed a MATLAB-based algorithm allowing motion tracking to quantify spontaneous uterine motility changes by measuring the rate of uterine horn movements. A major advantage of this ex vivo approach is that the reproductive tract remains intact throughout the entire experiment, preserving all intrinsic intrauterine cellular interactions. The major limitation of this approach is that up to 10-20% of uteri may exhibit no spontaneous motility. Thus far, this is the first quantitative ex vivo method for assessing spontaneous uterine motility in a Petri dish model.

Uterine contractions are important for the well-being of females. However, pathologically increased contractility may result in dysmenorrhea, especially in younger females. Here, we describe a simple ex vivo preparation allowing quick assessment of the efficacy of smooth muscle relaxants that may be used for treating dysmenorrhea.

Método Ex Vivo para evaluar la motilidad espontánea del tracto reproductivo de ratón y un algoritmo de seguimiento de movimiento del útero basado en MATLAB para el análisis de datos

Dysmenorrhea, or painful cramping, is the most common symptom associated with menses in females and its severity can hinder women&#39;s everyday lives. ...

Biología

Continuous Telemetric In Utero Tracheal Pressure Measurements in Fetal Lambs

Normal in utero lung development and growth rely upon the expansion of airspaces and the controlled efflux of lung liquid into the amniotic space. Infants with congenital diaphragmatic hernia (CDH) also have lung hypoplasia due to occupation of the chest cavity by the stomach and bowel and, in the most severe cases, the liver. Balloon tracheal occlusion reduces the severity of lung hypoplasia in fetuses with CDH but increases the risk of premature birth. Understanding the optimal occlusion pressure and duration required to improve lung hypoplasia with tracheal occlusion is essential to improving in utero corrective treatments for CDH. The study reports a new method for continuous measurement of the intratracheal and amniotic pressures in an unoccluded and occluded fetal lamb surgical model of CDH. Time-pregnant Merino ewes underwent two recovery hysterotomies: the first at ~80 days of gestation to create the CDH, and the second at ~101 days of gestation to occlude the fetal trachea and implant an intratracheal and amniotic pressure measurement device. Lambs were delivered at ~142 days, and the pressure measurement device was removed and cleaned. The data were downloaded and filtered using a 6 h window. Transrespiratory pressure was calculated.

<meta charset="utf8"></head><body>Mediciones telemétricas continuas de la presión traqueal en el útero en corderos fetales

Optimization of occlusion pressure and duration required to improve lung hypoplasia with tracheal occlusion is essential to improve in utero corrective treatments for congenital diaphragmatic hernia (CDH). This study reports a new method for continuous measurement of the tracheal pressure in an occluded and unoccluded fetal lamb surgical model of CDH.

Mediciones telemétricas continuas de la presión traqueal en el útero en corderos fetales

Normal in utero lung development and growth rely upon the expansion of airspaces and the controlled efflux of lung liquid into the amniotic space. Infants ...

Telemetría intrauterina para medir la presión del ratón contráctil

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