Este trabajo está financiado por la Organización Holandesa para la Investigación Científica (NWO) (sin Veni. 916.86.125), otorgado a L. Reneman. El donante no tenía ningún papel en el diseño del estudio, la recogida y análisis de datos, la decisión de publicar, o la preparación del manuscrito. No hay conflictos de intereses.

<ol>
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No tenemos nada que revelar.

5-HT phMRI es una herramienta prometedora para evaluar la función del neurotransmisor en animales in vivo. Se visualiza la respuesta cerebral a un reto de 5-HT, con la RM funcional. RM tiene la gran ventaja de tener una alta resolución espacial y no necesita la inyección de contraste, agente o moléculas marcadas por radio, evitando así la exposición repetitiva a las radiaciones ionizantes. Esta técnica es aplicable en los sujetos humanos y animales y por lo tanto muy adecuado para la investigación traslacional de los sistemas neurotransmisores y trastornos psiquiátricos. Su aplicación es por supuesto no se limita a la vía de la 5-HT y ya ha sido ampliamente utilizado para evaluar los efectos de los fármacos dopaminérgicos en ambos 5,15 animales y seres humanos 22. Sin embargo, phMRI en pequeños animales sigue siendo un reto, como ya se ha señalado en los artículos de revisión por parte de Martín y 11 de Sibson y Steward 20. Uno de estos retos es el mantenimiento of parámetros estables fisiológicos durante la adquisición de la imagen. La mayoría de los anestésicos pueden alterar la función cardiovascular y teniendo en cuenta que phMRI es críticamente dependiente de los parámetros cardiovasculares / hemodinámico es esencial para asegurar que los cambios hemodinámicos son imputables exclusivamente al desafío fármaco determinado. Por consiguiente, es de vital importancia que pCO 2 niveles se mantienen constantes durante la adquisición de línea de base. La ventilación mecánica puede ayudar a asegurar la estabilidad fisiológica, y se utiliza a menudo en este tipo de experimentos. Nosotros, sin embargo optó por utilizar de libre respiración de los animales para dejar abierta la posibilidad de realizar estudios longitudinales en el futuro. En su lugar, ampliamente monitoreado (y alterado) la tasa de respiración y los valores de gases en sangre para asegurar la estabilidad fisiológica dentro de los intervalos normales antes de iniciar la exploración funcional y de esta manera para preservar la reactividad vascular estable y, por tanto T2 * / señal en T2. La literatura sobre los efectos de los anestésicos generales sobre la hemodinámica cerebral y metabolism es abundante 20 y más allá del alcance de este manuscrito. Se optó por utilizar anestesia con gas, con el ± 2% de isoflurano en este protocolo específico, ya que con anestésicos inhalatorios, la profundidad de la anestesia puede ser rápida y fácil de controlar. Esto es importante en nuestra configuración para garantizar normales rango estable pCO 2 niveles antes del comienzo de la adquisición de la imagen. El isoflurano es el más comúnmente utilizado hoy anestésico por inhalación y permite una rápida inducción y recuperación, lo cual es importante para estudios longitudinales. También produce una mínima depresión cardiovascular y respiratoria y provoca una buena relajación de los músculos esqueléticos. En segundo lugar, la administración intravenosa del fármaco estimulante es más complicado en animales pequeños que en los seres humanos. La cirugía que se necesita para la canalización de la arteria y la vena femoral requiere personal bien capacitado y con experiencia. Debido a estos procedimientos invasivos es en este momento utiliza principalmente en los procedimientos de terminales. Sin embargo, la monitorización no invasiva de la homeostasis de sangre y la inyección de vena de la cola podría ser utilizado para estudios longitudinales 23. Además, hay algunas limitaciones más generales a la técnica, que no son específicos para los animales phMRI. Además, como se ha señalado por Martin y Sibson 11, un potencial de confundir todos los estudios fMRI es que se supone que los cambios en la actividad cerebral evocadas por el desafío reflejar cambios en la actividad neuronal en lugar de periféricos efectos sistémicos. Especialmente en las estructuras cerebrales más profundas, una comprensión relativamente pobre de acoplamiento neurovascular (relación entre los cambios de actividad neuronal y los cambios hemodinámicos) se mantiene. Estudios del tipo realizado por Logothetis 10 para determinar acoplamiento neurovascular en la corteza todavía no se han realizado en otras partes del cerebro. Por lo tanto, se desconoce lo que un aumento en la señal BOLD con estructuras importantes como el estriado o la amígdala es telling nosotros acerca de la actividad neuronal. Lo mejor que podría decir en este momento es que la región del cerebro reacciona ante el desafío dado y que, dependiendo del tratamiento y / o las condiciones, podemos observar los cambios significativos de la reactividad del cerebro. Este gran medida puede ser verificada por mirando a la vez los datos de resonancia magnética y las respuestas fisiológicas. El patrón general de la activación cerebral debe ser región específica y restringida a las áreas con, en este caso, una inervación de alta de 5-HT, y no tanto una respuesta vascular en general. Además, un perfil diferente temporal entre los cambios vasculares y hemodinámicas que se espera. Considerando que los cambios de presión arterial volver a sus valores basales dentro de varios minutos, el efecto del fármaco sobre la activación BOLD es en el caso de fluoxetina visible hasta el final de la adquisición de imágenes y corresponden a las propiedades farmacocinéticas conocen de este fármaco. Finalmente, las respuestas fisiológicas de todos los animales deben ser similares a fin de hacer comparaciones entre sujetos. NonetheleSS, se sabe que una regulación neurogénica del flujo sanguíneo local por 5-HT existen 4. Por lo tanto, no puede excluirse que los cambios locales de señal BOLD puede atribuir a los cambios vasculares debido a la liberación de 5-HT en la proximidad de los vasos. Aunque estos efectos no están asociados a la activación neuronal local y por lo tanto se puede considerar como resultados positivos falsos, es también un índice de la función general específica del sistema de 5-HT (véase también 3). Los pasos críticos de esta técnica son por lo tanto, para controlar las respuestas fisiológicas y extensamente para asegurarse de que las condiciones fisiológicas del animal son estables antes y durante la adquisición de la imagen. También las condiciones del escáner debe ser tan estable como sea posible y exactamente el mismo para cada animal. Estabilidad de la señal de la secuencia deben ser verificadas y confirmadas antes del inicio de su experimento. Además, asegúrese de tener siempre gran potencia estadística suficiente, incluso con grou sujeto pequeñops. Para una buena revisión de las consideraciones experimentales de animales phMRI en general, vea Steward 20 y para ver un ejemplo adicional de un protocolo experimental para la resonancia magnética funcional farmacológica en ratas y ratones, ver a Ferrari 5. Las posibles modificaciones de la técnica que se describen aquí son numerosos. Uno podría: <ol><li> usando un medicamento diferente para el 5-HT reto, como otro ISRS o receptor 5-HT (hormiga) agonistas 16,13,18,7 o incluso a un doble desafío con el fin de revelar los mecanismos subyacentes de la acción del fármaco 6,19; </li><li> utilizar una configuración diferente experimental, tales como un régimen diferente de anestesia, ventilación mecánica, la piscina de sangre en lugar de agentes de contraste BOLD 15, los estudios longitudinales (animal necesita para mantenerse vivo, por lo que la sangre no invasiva de la presión / de sangre de medición de gas y / o ventilación mecánica es posible), o incluso combinaciones con otros (invasivo) los métodos, como la grabación de la actividad neuronal con la RM compatible con elementosctrodes 10 o PET / SPECT 4; </li><li> utilizar diferentes métodos de análisis de datos de resonancia magnética como método de la &quot;p-bloque&quot; de McKie 12 o el análisis de la conectividad funcional 15. </li></ol> ¿Qué decisiones que tomas en el montaje experimental es altamente dependiente de las posibilidades y / o la experiencia dentro de su laboratorio y el tipo de pregunta de investigación que le gustaría contestar.

<table border="1"><tbody><tr><td> Nombre del reactivo / equipo </td><td> Empresa </td><td> Número de catálogo </td><td> Comentarios </td></tr><tr><td> El isoflurano </td><td> ABBOTT <a href="http://www.abbott.co.uk/" target="_blank">Abbott Laboratories Ltd., Maidenhead, Reino Unido</a> </td><td> No B506 </td><td> Mezcla con el aire médica </td></tr><tr><td> Médico de aire </td><td> <a href="http://www.bochealthcare.co.uk/" target="_blank">BOC Salud</a> </td><td></td><td></td></tr><tr><td> Almohadilla eléctrica </td><td> <a href="http://www.harvardapparatus.co.uk/" target="_blank">Harvard aparatos</a> </td><td> 507223F <a href="http://www.harvardapparatus.co.uk/webapp/wcs/stores/servlet/haisku3_10001_11555_39108_-1_HAUK_ProductDetail_N_37610_37611_37613" target="_blank">Sistema completo Manta homeotérmicos con sonda flexible, medio, 230 VAC, 50 Hz</a> </td><td></td></tr><tr><td> L de la Seda<html> igature </td><td> <a href="http://www.harvardapparatus.com/" target="_blank">http://www.harvardapparatus.com/</a> </td><td> <a href="http://www.harvardapparatus.com/webapp/wcs/stores/servlet/product_11051_10001_43051_-1_HAI_ProductDetail" target="_blank">2-0 de seda negro trenzado no absorbible</a> Cat Num 51-7631 </td><td></td></tr><tr><td> PE-50 Cánula </td><td> <a href="http://www.scientificlabs.co.uk/" target="_blank">http://www.scientificlabs.co.uk/~~V</a> </td><td> <a href="http://www.scientificlabs.co.uk/product/TUB3662" target="_blank">Portex tubería PE 0.58x0.96mm</a> </td><td> 0.58 ID 0.96 mm OD </td></tr><tr><td> Heparina sódica </td><td> <a href="http://www.leo-pharma.com/Home/Products/List-of-LEO-Pharma-products.aspx" target="_blank">Leo Laboratories Ltd, Bucks., Reino Unido</a> </td><td> 1000IU/ml Heparina sódica </td><td> 15 U / ml </td></tr><tr><td> Adhesivo tisular Vetbond </td><td> <a href="http://solutions.3m.co.uk" target="_blank">3M</a> </td><td>ttp :/ / solutions.3m.co.uk/wps/portal/3M/en_GB/HealthCare/Home/ProdInfo/VetProducts o &quot;target =&quot; _blank &quot;&gt; M adhesivo tisular Vetbond </td><td></td></tr><tr><td> Sistema de seguimiento </td><td> <a href="http://www.i4sa.com/" target="_blank">Instrumentos SA</a> </td><td> <a href="http://www.i4sa.com" target="_blank">http://www.i4sa.com</a> <a href="http://69.20.42.253/web_app/main/defaultProduct.aspx?ID=16&amp;PT=3" target="_blank">Modelo de sistema de monitoreo 1025L</a> </td><td> Los monitores de respiración y la temperatura </td></tr><tr><td> Transductor de presión </td><td> <a href="http://www.biopac.com/" target="_blank">Biopac Systems Corp</a> </td><td> <a href="http://www.biopac.com/blood-pressure-transducer" target="_blank">TRANSDUCTOR DE PRESION ARTERIAL - TSD104A</a> <a href="http://www.biopac.com/data-acquisition-analysis-system-mp150-system-windows" target="_blank">SISTEMA DE ADQUISICIÓN DE DATOS MP150 - WIN - MP150WSW</a> </td><td> Monitores de presión arterial </td></tr><tr><td> Analizador de gases de la sangre Rapidlab </td><td> <a href="http://www.medical.siemens.com/" target="_blank">Siemens Medical Solutions Diagnostics</a> </td><td> <a href="http://www.medical.siemens.com/webapp/wcs/stores/servlet/ProductDisplay~q_catalogId~e_-111~a_catTree~e_100001,1023069,1016115~a_langId~e_-111~a_productId~e_172940~a_storeId~e_10001.htm" target="_blank">Rapidlab 248/348 Sistemas</a> </td><td></td></tr><tr><td> Escáner de animales 4.7T </td><td> Agilent Technologies (anteriormente Magnex) Frecuencia de 199.845 MHz 4.7T </td><td></td><td></td></tr><tr><td> MR cama estereotáctica compatibles </td><td> <a href="http://www.m2mimaging.com/products/preclinical.html" target="_blank">M2M Imaging Corp</a> </td><td> Cama de la rata: el elemento PA multi AHS 50-72-1003/100 </td><td></td></tr><tr><td> Bobina </td><td> <a href="http://www.m2mimaging.com/products/preclinical.html" target="_blank">M2M Imaging Corp</a> </td><td> Volumen TH / Rx RQD1 72/112 200 </td><td></td></tr><tr><td> Clorhidrato de Fluoxetina </td><td> Sigma-Aldrich </td><td> F-132 </td><td> 5mg/kg en solución salina </td></tr></tbody></table>

the use of pharmacological-challenge fmri in pre-clinical research: application to the 5-ht system

Farmacológico resonancia magnética (phMRI) es un método nuevo y prometedor para estudiar los efectos de las sustancias sobre las funciones cerebrales que a la larga pueden ser utilizados para desentrañar los mecanismos neurobiológicos subyacentes detrás de la acción de drogas y los trastornos relacionados con los neurotransmisores, como la depresión y el TDAH. Como la mayoría de los métodos de imagen (PET, SPECT, CT), que representa un avance en la investigación de los trastornos del cerebro y la función relacionada de las vías de neurotransmisores en forma no invasiva con respecto a la conectividad neuronal global. Por otra parte, también ofrece la herramienta ideal para la traducción a las investigaciones clínicas. Resonancia magnética, mientras que todavía detrás de las estrategias de imagen molecular en comparación con el PET y SPECT, tiene la gran ventaja de tener una alta resolución espacial y no hay necesidad de que la inyección de un contraste agente o moléculas marcadas por radio, evitando así la exposición repetitiva a las radiaciones ionizantes . Resonancia magnética funcional (fMRI) es ampliamente utilizado en la fijación de la investigación y la clínica, donde es gn general combinado con una tarea psico-motor. phMRI es una adaptación de resonancia magnética funcional que permite la investigación de un sistema neurotransmisor específico, como la serotonina (5-HT), bajo condiciones fisiológicas o patológicas después de la activación a través de la administración de un fármaco estimulante específico. El objetivo del método descrito aquí es evaluar el cerebro de 5-HT función en los animales libres de respiración. Al desafiar el sistema 5-HT, mientras que al mismo tiempo la adquisición de las imágenes de RM funcionales a través del tiempo, la respuesta del cerebro a este desafío puede ser visualizada. Varios estudios en animales han demostrado ya que la droga-inducida por el aumento en los niveles extracelulares de por ejemplo 5-HT (agentes liberadores, selectivos de la recaptación de los bloqueadores, etc) evocan específicos de la región los cambios en el nivel de oxigenación de la sangre que dependen de señales de resonancia magnética (negrita) (señal debido a un cambio de los niveles de hemoglobina oxigenados / desoxigenada que ocurren durante la activación del cerebro a través de un aumento del suministro de sangre para suministrar el oxígeno y glucose a las neuronas más exigentes) que proporciona un índice de la función del neurotransmisor. También se ha demostrado que estos efectos pueden ser revertidos por los tratamientos que disminuyen la disponibilidad de 5-HT 16,13,18,7. En ratas adultas, los cambios de señal BOLD siguientes aguda administración ISRS han sido descritos en varias de 5-HT regiones del cerebro relacionadas, es decir, áreas corticales, el hipocampo, el hipotálamo y el tálamo 9,16,15. La estimulación del sistema 5-HT y su respuesta a este reto se puede por lo tanto utilizarse como una medida de su función en animales y seres humanos 2,11.

Watch this Scientific Journal Video about The Use of Pharmacological-challenge fMRI in Pre-clinical Research: Application to the 5-HT System at JoVE.com

The Use of Pharmacological-challenge fMRI in Pre-clinical Research: Application to the 5-HT System

El objetivo de esta técnica consiste en evaluar la serotonina (5-HT) la función de neurotransmisor en el animal vivo y sin respiración, con imágenes de resonancia magnética farmacológica (phMRI) y un desafío por vía intravenosa con un inhibidor selectivo de la recaptación de serotonina (ISRS), fluoxetina.

El uso de la fMRI farmacológico-reto en la investigación preclínica: la aplicación al sistema de 5-HT

Pharmacological MRI (phMRI) is a new and promising method to study the effects of substances on brain function that can ultimately be used to unravel ...

the-use-pharmacological-challenge-fmri-pre-clinical-research

Research

JoVE Journal

Medicine

15.3K vistas.  Academic Medical Center Amsterdam. El objetivo de esta técnica consiste en evaluar la serotonina (5-HT) la función de neurotransmisor en el animal vivo y sin respiración, con imágenes de resonancia magnética farmacológica (phMRI) y un desafío por vía intravenosa con un inhibidor selectivo de la recaptación de serotonina (ISRS), fluoxetina.

Video: The Use of Pharmacological-challenge fMRI in Pre-clinical Research: Application to the 5-HT System

Use of Animal Model of Sepsis to Evaluate Novel Herbal Therapies

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection. It has been routinely simulated in animals by several techniques, including infusion of exogenous bacterial toxin (endotoxemia) or bacteria (bacteremia), as well as surgical perforation of the cecum by cecal ligation and puncture (CLP)1-3. CLP allows bacteria spillage and fecal contamination of the peritoneal cavity, mimicking the human clinical disease of perforated appendicitis or diverticulitis. The severity of sepsis, as reflected by the eventual mortality rates, can be controlled surgically by varying the size of the needle used for cecal puncture2. In animals, CLP induces similar, biphasic hemodynamic cardiovascular, metabolic, and immunological responses as observed during the clinical course of human sepsis3. Thus, the CLP model is considered as one of the most clinically relevant models for experimental sepsis1-3.
Various animal models have been used to elucidate the intricate mechanisms underlying the pathogenesis of experimental sepsis. The lethal consequence of sepsis is attributable partly to an excessive accumulation of early cytokines (such as TNF, IL-1 and IFN-&gamma;)4-6 and late proinflammatory mediators (e.g., HMGB1)7. Compared with early proinflammatory cytokines, late-acting mediators have a wider therapeutic window for clinical applications. For instance, delayed administration of HMGB1-neutralizing antibodies beginning 24 hours after CLP, still rescued mice from lethality8,9, establishing HMGB1 as a late mediator of lethal sepsis. The discovery of HMGB1 as a late-acting mediator has initiated a new field of investigation for the development of sepsis therapies using Traditional Chinese Herbal Medicine. In this paper, we describe a procedure of CLP-induced sepsis, and its usage in screening herbal medicine for HMGB1-targeting therapies.

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection, and can be simulated by a surgical technique termed cecal ligation and puncture (CLP). Here we describe a method to use CLP-induced animal model to screen medicinal herbs for therapeutic agents.

El uso de modelos animales de sepsis para evaluar nuevas terapias a base de plantas

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection. It has been routinely simulated in animals by several ...

Investigación

Medicina

Handling of the Cotton Rat in Studies for the Pre-clinical Evaluation of Oncolytic Viruses

Oncolytic viruses are a novel anticancer therapy with the ability to target tumor cells, while leaving healthy cells intact. For this strategy to be successful, recent studies have shown that involvement of the host immune system is essential. Therefore, oncolytic virotherapy should be evaluated within the context of an immunocompetent model. Furthermore, the study of antitumor therapies in tolerized animal models may better recapitulate results seen in clinical trials. Cotton rats, commonly used to study respiratory viruses, are an attractive model to study oncolytic virotherapy as syngeneic models of mammary carcinoma and osteosarcoma are well established. However, there is a lack of published information on the proper handling procedure for these highly excitable rodents. The handling and capture approach outlined minimizes animal stress to facilitate experimentation. This technique hinges upon the ability of the researcher to keep calm during handling and perform procedures in a timely fashion. Finally, we describe how to prepare cotton rat mammary tumor cells for consistent subcutaneous tumor formation, and how to perform intratumoral and intraperitoneal injections. These methods can be applied to a wide range of studies furthering the development of the cotton rat as a relevant pre-clinical model to study antitumor therapy.

Cotton rats are extremely excitable and have a strong flight-or-fight response. A handling method optimized to reduce the stress of the animals is described which will make cotton rats more accessible as a preclinical model.

Manejo de la rata del algodón en Estudios para la evaluación preclínica de virus oncolíticos

Oncolytic viruses are a novel anticancer therapy with the ability to target tumor cells, while leaving healthy cells intact.  For this strategy to be ...

Pre-clinical Orthotopic Murine Model of Human Prostate Cancer

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

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

Preclínica modelo murino ortotópico de cáncer de próstata humano

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

Use of a Low-flow Digital Anesthesia System for Mice and Rats

A traditional vaporizer depends on flowing gas and atmospheric pressure for passive anesthetic vaporization. Newly developed direct injection vaporizers utilize a syringe pump to directly administer volatile anesthetics into a gas stream. Unlike a traditional vaporizer, it can be used at very low flow rates, making it ideal for use on mice and rats. 
The equipment's capability to use low flow rates could result in a substantial cost savings due to the reduced need for anesthetic agents, compressed gas, and charcoal scavenging filters1. A lower flow rate means less waste of anesthetic gas and likely reduces the risk of anesthetic exposure to laboratory personnel. Thus, the high levels of precision and safety associated with direct injection vaporizers, along with a reduced need for anesthetic agents, compressed gas, and charcoal filters are beneficial for research requiring small animal anesthesia. 
The goal of this protocol is to demonstrate the use of a syringe-driven direct injection vaporizer as part of a digital, low-flow anesthesia system. The direct injection vaporizer is capable of accurately delivering anesthesia at very low flow rates compared to a traditional vaporizer, making it a promising alternative for controlled gas anesthetic delivery to rodents.

Here, we present a protocol to more safely and efficiently administer anesthetic gas to mice using a digital, low flow anesthesia system utilizing a syringe-driven direct injection vaporizer.

El uso de un sistema de bajo flujo Anestesia digital para ratones y ratas

A traditional vaporizer depends on flowing gas and atmospheric pressure for passive anesthetic vaporization. Newly developed direct injection vaporizers ...

Ultrahigh Resolution Mouse Optical Coherence Tomography to Aid Intraocular Injection in Retinal Gene Therapy Research

HR-SD-OCT is utilized to monitor the progression of photoreceptor degeneration in live mouse models, assess the delivery of therapeutic agents into the subretinal space, and to evaluate toxicity and efficacy in vivo. HR-SD-OCT uses near infrared light (800-880 nm) and has optics specifically designed for the unique optics of the mouse eye with sub-2-micron axial resolution. Transgenic mouse models of outer retinal (photoreceptor) degeneration and controls were imaged to assess the disease progression. Pulled glass microneedles were used to deliver sub retinal injections of adeno-associated virus (AAV) or nanoparticles (NP) via a trans-scleral and trans-choroidal approach. Careful positioning of the needle into the subretinal space was required prior to a calibrated pressure injection, which delivers fluid into the sub retinal space. Real time subretinal surgery was conducted on our retinal imaging system (RIS). HR-SD-OCT demonstrated progressive uniform retinal degeneration due to expression of a toxic mutant human mutant rhodopsin (P347S) (RHOP347S) transgene in mice. HR-SD-OCT allows rigorous quantification of all the retinal layers. Outer nuclear layer (ONL) thickness and photoreceptor outer segment length (OSL) measurements correlate with photoreceptor vitality, degeneration, or rescue. The RIS delivery system allows real-time visualization of subretinal injections in neonatal (~P10-14) or adult mice, and HR-SD-OCT immediately determines success of delivery and maps areal extent. HR-SD-OCT is a powerful tool that can evaluate the success of subretinal surgery in mice, in addition to measuring vitality of photoreceptors in vivo. HR-SD-OCT can also be used to identify uniform animal cohorts to evaluate the extent of retinal degeneration, toxicity, and therapeutic rescue in preclinical gene therapy research studies.

Here we demonstrate a novel approach to using high resolution spectral-domain optical coherence tomography (HR-SD-OCT) to assist delivery of gene therapy agents into the subretinal space, assess its areal coverage, and characterize photoreceptor vitality.

Tomografía de coherencia óptica ultraalta resolución ratón para facilitar la inyección Intraocular en investigación de retina la terapia génica

HR-SD-OCT is utilized to monitor the progression of photoreceptor degeneration in live mouse models, assess the delivery of therapeutic agents into the ...

A Pre-Clinical Porcine Model of Orthotopic Heart Transplantation

Fifty-years following the first successful report, cardiac transplantation remains the gold-standard treatment for eligible patients with advanced heart failure. Multiple small-animal models of heart transplantation have been used to study the acute and long-term effects of novel therapies. However, few are tested and demonstrated success in clinical trials. It is of critical importance to evaluate new therapies in a clinically relevant large-animal model for efficient and reliable translation of basic studies&#39;&#160;findings. Here, we describe a pre-clinical large-animal (porcine) model of orthotopic heart transplantation that has been firmly established and previously used to investigate novel cardioprotective strategies. This procedure focuses on acute ischemia-reperfusion injury and is a reliable method to investigate novel interventions which have been tested and validated in smaller experimental models, such as the murine model. We demonstrate its usefulness in assessing cardiac performance during the early post-transplantation period and other potential possibilities enabled by the model.

Here, we describe a pre-clinical large-animal (porcine) model of orthotopic heart transplantation that has been firmly established and utilized to investigate novel cardioprotective strategies.

Un modelo preclínico porcino de trasplante ortotópico de corazón

Fifty-years following the first successful report, cardiac transplantation remains the gold-standard treatment for eligible patients with advanced heart ...

Pre-clinical Model of Cardiac Donation after Circulatory Death

Cardiac transplantation demand is on the rise; nevertheless, organ availability is limited due to a paucity of suitable donors. Organ donation after circulatory death (DCD) is a solution to address this limited availability, but due to a period of prolonged warm ischemia and the risk of tissue injury, its routine use in cardiac transplantation is seldom seen. In this manuscript we provide a detailed protocol closely mimicking current clinical practices in the context of DCD with continuous monitoring of heart function, allowing for the evaluation of novel cardioprotective strategies and interventions to decrease ischemia-reperfusion injury.
In this model, the DCD protocol is initiated in anesthetized Lewis rats by stopping ventilation to induce circulatory death. When systolic blood pressure drops below 30 mmHg, the warm ischemic time is initiated. After a pre-set warm ischemic period, hearts are flushed with a normothermic cardioplegic solution, procured, and mounted onto a Langendorff ex vivo heart perfusion system. Following 10 min of initial reperfusion and stabilization, cardiac reconditioning is continuously evaluated for 60 min using intraventricular pressure monitoring. A heart injury is assessed by measuring cardiac troponin T and the infarct size is quantified by histological staining. The warm ischemic time can be modulated and tailored to develop the desired amount of structural and functional damage. This simple protocol allows for the evaluation of different cardioprotective conditioning strategies introduced at the moment of cardioplegia, initial reperfusion and/or during ex vivo perfusion. Findings obtained from this protocol can be reproduced in large models, facilitating clinical translation.

This protocol shows a simple and flexible approach for the evaluation of new conditioning agents or strategies to increase the feasibility of cardiac donation after circulatory death.

Modelo preclínico de donación cardíaca después de la muerte circulatoria

Cardiac transplantation demand is on the rise; nevertheless, organ availability is limited due to a paucity of suitable donors. Organ donation after ...

Invasive Hemodynamic Assessment for the Right Ventricular System and Hypoxia-Induced Pulmonary Arterial Hypertension in Mice

Pulmonary arterial hypertension (PAH) is a chronic and severe cardiopulmonary disorder. Mice are a popular animal model used to mimic this disease. However, the evaluation of right ventricular pressure (RVP) and pulmonary artery pressure (PAP) remains technically challenging in mice. RVP and PAP are more difficult to measure than left ventricular pressure because of the anatomical differences between the left and right heart systems. In this paper, we describe a stable right heart hemodynamic measurement method and its validation using healthy and PAH mice. This method is based on open-chest surgery and mechanical ventilation support. It is a complicated procedure compared to closed chest procedures. While a well-trained surgeon is required for this surgery, the advantage of this procedure is that it can generate both RVP and PAP parameters at the same time, so it is a preferable procedure for the evaluation of PAH models.

Here, we present a protocol to perform an invasive hemodynamic assessment of the right ventricle and pulmonary artery in mice using an open-chest surgery approach.

Evaluación hemodinámica invasiva para el sistema ventricular derecho y hipertensión arterial pulmonar inducida por hipoxia en ratones

Pulmonary arterial hypertension (PAH) is a chronic and severe cardiopulmonary disorder. Mice are a popular animal model used to mimic this disease. ...

A Pre-clinical Rat Model for the Study of Ischemia-reperfusion Injury in Reconstructive Microsurgery

Ischemia-reperfusion injury is the main cause of flap failure in reconstructive microsurgery. The rat is the preferred preclinical animal model in many areas of biomedical research due to its cost-effectiveness and its translation to humans. This protocol describes a method to create a preclinical free skin flap model in rats with ischemia-reperfusion injury. The described 3 cm x 6 cm rat free skin flap model is easily obtained after the placement of several vascular ligatures and the section of the vascular pedicle. Then, 8 h after the ischemic insult and completion of the microsurgical anastomosis, the free skin flap develops the tissue damage. These ischemia-reperfusion injury-related damages can be studied in this model, making it a suitable model for evaluating therapeutic agents to address this pathophysiological process. Furthermore, two main monitoring techniques are described in the protocol for the assessment of this animal model: transit-time ultrasound technology and laser speckle contrast analysis.

Here, we describe a pre-clinical animal model for studying the pathophysiology of ischemia-reperfusion injury in reconstructive microsurgery. This free skin flap model based on the superficial caudal epigastric vessels in the rat may also allow for the evaluation of different therapies and compounds to counteract ischemia-reperfusion injury-related damage.

Un modelo preclínico de rata para el estudio de la lesión por isquemia-reperfusión en microcirugía reconstructiva

Ischemia-reperfusion injury is the main cause of flap failure in reconstructive microsurgery. The rat is the preferred preclinical animal model in many ...

Use of an Integrated Low-Flow Anesthetic Vaporizer, Ventilator, and Physiological Monitoring System for Rodents

Low-flow digital vaporizers commonly utilize a syringe pump to directly administer volatile anesthetics into a stream of carrier gas. Per animal welfare recommendations, animals are warmed and monitored during procedures requiring anesthesia. Common anesthesia and physiological monitoring equipment include gas tanks, anesthetic vaporizers and stands, warming controllers and pads, mechanical ventilators, and pulse oximeters. A computer is also necessary for data collection and to run equipment software. In smaller spaces or when performing field work, it can be challenging to configure all this equipment in limited space.
The goal of this protocol is to demonstrate best practices for use of a low-flow digital vaporizer using both compressed oxygen and room air, along with an integrated mechanical ventilator, pulse oximeter, and far infrared warming as an all-inclusive anesthesia and physiological monitoring suite ideal for rodents.

Here, we present a protocol to safely and effectively administer anesthetic gas to mice using a digital, low flow anesthesia system with integrated ventilator and physiological monitoring modules.

Uso de un vaporizador anestésico de bajo flujo integrado, ventilador y sistema de monitoreo fisiológico para roedores

Low-flow digital vaporizers commonly utilize a syringe pump to directly administer volatile anesthetics into a stream of carrier gas. Per animal welfare ...