Gracias a Jessica Ottmar sugerencias protocolo y las contribuciones a los datos representativos. Un agradecimiento especial a Sabrina Blackledge, Kohoutek Lauren y Shelton Kerisa para demostrar este protocolo.

1. Configurar <ol><li> Todas las investigaciones deben ser aprobados por la Junta de Revisión Institucional o del comité apropiado humanos revisión de sujetos antes de la recolección de datos. </li><li> Idealmente, dos habitaciones deben estar disponibles para llevar a cabo la TSST. Una cómoda sala de espera debe estar disponible para los participantes antes del comienzo del TSST y durante los períodos de recuperación. Una sala de entrevistas por separado se debe utilizar durante la preparación del discurso, el rendimiento del habla y las porciones de matemáticas de la TSST para introducir la novedad y la uncontrolability. </li><li> Organizar la sala de entrevistas para que el participante se enfrentará a los entrevistadores de la Confederación y de vídeo de posición o equipo de grabación de audio en el campo visual de los participantes. Las batas de laboratorio debe estar disponible para los entrevistadores para usar durante el TSST para aumentar la tensión durante la TSST. Instruir a los entrevistadores para mantener el contacto visual con los participantes y abstenerse de hacer expresiones faciales emocionales durante la TSST. </li><li> Considere cuidadosamente ªe pregunta de investigación (s) al seleccionar las mediciones dependientes. Directrices para el análisis y la publicación de varias medidas de respuesta al estrés son proporcionados por diversas fuentes (directrices para la publicación y análisis de las medidas psicofisiológicas son proporcionados por la Sociedad para la Investigación psicofisiológicos: <a href="http://www.sprweb.org/journal/index.cfm#guidelines" target="_blank">http://www.sprweb.org/journal/index. pcm # guidelines</a> ). 5 Consulta de estos recursos en el diseño de experimentos es recomendable. </li><li> Configurar todos los equipos necesarios para registrar las respuestas fisiológicas, psicológicas o endocrino a la TSST. Este equipo puede incluir (pero no limitados a) los electrodos y equipos electrónicos y / o software para registrar la frecuencia cardíaca, la reactividad cardiovascular, conductancia de la piel o la respuesta, abrir y cerrar los ojos, la contracción muscular / tono, la frecuencia respiratoria, papel y lápiz o las versiones electrónicas de instrumentos psicológicos como la ansiedad Estado-Rasgo Inventory, 6 tubos y otros materiales de recolección de muestras de orina, sangre o saliva;. o equipos de grabación de vídeo Para esta demostración, vamos a utilizar tres derivaciones del electrocardiograma (ECG) electrodos para medir la frecuencia cardíaca durante el TSST, y la administración repetida de la Inventario de Ansiedad Estado-Rasgo para medir la respuesta psicológica antes y después del factor estresante. Vamos a utilizar un sistema de registro básico diseñado para medir las respuestas fisiológicas (Biopac sistema MP36) para registrar la frecuencia cardíaca. </li><li> Si las muestras de sangre (en lugar de saliva) se recogerán durante el experimento, los esfuerzos para minimizar el dolor y la angustia durante la recolección debe hacerse. Los procedimientos de recolección de sangre se puede introducir una carga adicional física o psicológica, la respuesta de la confusión a la TSST. </li><li> Cuando el participante llega, les damos la bienvenida al laboratorio. Obtener el consentimiento informado y el asiento del participante cómodamente. </li><li> Recopilar información demográfica, la información de investigación o additional llegada medidas que son requeridas por el experimento. Verificar que el participante no ha consumido alimentos o bebidas (excepto agua) en la última hora si recoger las mediciones de cortisol. Un breve período de ayuno se recomienda si va a recoger muestras de saliva para el análisis. Cambios en el pH asociados a los alimentos ácidos o bebéis, o la contaminación visible puede interferir con la medición precisa de los marcadores salivales. Examen de toxicología de orina puede ser necesario si las medidas de interés pueden estar contaminados por la medicación u otros metabolitos. Información sobre los factores que pueden afectar la respuesta al estrés (por ejemplo, los medicamentos, la enfermedad, el sueño, los últimos acontecimientos estresantes, la fobia social) también debe ser considerado cuando los participantes de detección. </li></ol> 2. Las mediciones pre-tensión <ol><li> Instruir a los participantes a esperar cómodamente durante 45 minutos en la sala de espera. Usted puede optar por ofrecer al participante con material de lectura, tales como revistas que contienen emotradicionalmente neutral en el contenido, o cualquier otra actividad emocional-neutral durante este período. </li><li> Después del período de 45 minutos de espera, para administrar el Estado-Rasgo de Ansiedad para evaluar el estrés pre-ansiedad. 6 Por otra parte, otros instrumentos que están diseñados para evaluar los cambios de estado de estrés, la ansiedad o la relajación pueden ser administrados (por ejemplo, las escalas analógicas visuales). </li><li> Se recoge una &quot;pre-tensión&quot; muestra de saliva. Instruir a los participantes a enjuagarse la boca con agua. Darle al participante una sección de 5-10 cm de paja para beber y un lugar limpio, desechable 2,0 ml vial de polipropileno. Instruir a los participantes a inclinar la cabeza hacia delante para permitir la saliva a la piscina en el piso de la boca y pasar la saliva a través de la paja en el vial. Recoger aproximadamente 1,5-2,0 ml de saliva de los participantes. Por otra parte, la saliva puede ser recogida mediante el uso de mecha absorbente dental (u otro material similar) e instruir a los participantes para masticar o para saturar el material con la saliva. Si posposible, salir de la habitación para proporcionar al participante en la vida privada. Etiqueta y guardar todas las muestras de saliva a -20 ° C tan pronto como sea posible. </li><li> Registro de 5 minutos de pre-tensión mediciones fisiológicas. </li></ol> 3. La TSST <ol><li> Si usted va a utilizar una sala de entrevistas por separado (para aumentar los efectos del estrés de la TSST), llevar el participante de la sala de espera en la sala de entrevistas. </li><li> Para comenzar el período de preparación TSST discurso, leyó el guión siguiente a la participante: &quot;Esta es la parte de preparación del discurso de la tarea, usted debe prepararse mentalmente un discurso de cinco minutos que describe por qué sería un buen candidato para su trabajo ideal. Su discurso será grabado y revisado por un panel de jueces entrenados para hablar en público. Tienes diez minutos para preparar y el tiempo comienza ahora &quot;. </li><li> Registro de las mediciones fisiológicas durante el período discurso de 10 minutos de preparación. Establecer un temporizador digital para 10 minutos y salir de la habitación. </li&gt; <li> Después de 10 minutos, volver a la sala vistiendo una bata de laboratorio. Lea el siguiente script para el participante: &quot;Esta es la parte del habla de la tarea que está a pronunciar un discurso que explique por qué usted sería un buen candidato para su trabajo ideal Usted debe hablar para todo el período de tiempo de cinco minutos... Su tiempo comienza ahora &quot;. La cámara de vídeo apoyo debe ser activado ahora, para aumentar la evaluación calidad / estrés. </li><li> Registro de las mediciones fisiológicas durante el período discurso de 5 minutos el rendimiento. Si el participante deja de hablar durante el discurso, permite que él o ella a permanecer en silencio durante 20 segundos. Si él o ella no volver a hablar, pedir al participante a seguir hablando enseñándoles: &quot;Todavía hay tiempo restante.&quot; </li><li> Al final del período discurso de 5 minutos el rendimiento, lea el siguiente script para el participante: &quot;Durante la porción de matemáticas últimos cinco minutos de esta tarea, se le pedirá a restar de forma secuencial el número 13 de 1022 Usted verbal.mente informe sus respuestas en voz alta, y se le pedirá que empezar de nuevo desde 1022, si se comete un error. . El tiempo empieza ahora &quot;Si el participante comete un error, le informarán con:&quot; Eso es incorrecto, por favor, empezar de nuevo desde 1022 &quot;Establecer un temporizador digital por 5 minutos.. </li><li> Registro de las mediciones fisiológicas durante el período de matemáticas de 5 minutos el rendimiento. </li><li> Al final del periodo de rendimiento en matemáticas de la TSST, se recoge una &quot;post-estrés&quot; muestra de saliva. </li><li> Administrar el estrés post-Estado-Rasgo de Ansiedad. </li></ol> 4. Estrés post-Medidas de recuperación <ol><li> Volver a los participantes a la sala de espera inicial. Durante el período de recuperación en primer lugar, instruir a los participantes a esperar cómodamente durante 20 minutos. Establecer un temporizador digital para 20 minutos y salir de la habitación. </li><li> Volver a la sala de espera después de 20 minutos y se recoge una &quot;recuperación de una&quot; muestra de saliva. </li><li> Durante el período de recuperación en segundo lugar, instruir a los participantes que esperar de nuevo cómodamente durante 20 minutos. Establecer un temporizador digital para 20 minutos y salir de la habitación. </li><li> Volver a la sala de espera después de 20 minutos y se recoge &quot;una recuperación de 2&quot; muestra de saliva. </li><li> Instruir a los participantes para completar la &quot;recuperación&quot; Estado-Rasgo de Ansiedad. </li><li> Interrogar al participante como a la verdadera naturaleza del experimento. Informará al participante que su desempeño no fue registrado y que ningún análisis de su discurso o el rendimiento en matemáticas se llevará a cabo. Explique a los participantes que las tareas con las que se presentaron eran demasiado difíciles y no reflexionar sobre la aptitud del participante o de la capacidad. </li></ol> 5. Análisis de Datos <ol><li> Para analizar las medidas psicológicas, la puntuación de cada Estado-Rasgo de Ansiedad de la tensión pre-, post-estrés y los puntos de tiempo de recuperación 6. </li><li> Para analizar las medidas fisiológicas, calcular una puntuación media de frecuencia cardiaca para cada uno de los cinco minutos de &quot;estrés pre&quot;, &quot;preparación del discurso&quot;, &quot;el rendimiento del habla&quot;, y m &quot;ATH &quot;períodos de la TSST. Por otra parte, calcular las puntuaciones medias de cada minuto de la TSST o la frecuencia cardíaca medida en puntos discretos de tiempo durante el TSST. </li><li> Para analizar las medidas neuroendocrinas, las muestras de saliva para la prueba de la concentración de hormona utilizando kits EIA disponibles comercialmente. </li></ol> 6 Opcional:. Ensayo adicional de cortisol salival Protocolo de información (que se reproduce a partir de insertar Salimetrics cortisol salival kit de ensayo) <ol><li> Lleve todos los reactivos de ensayo salivales kit a temperatura ambiente. </li><li> Prepare el tampón de lavado de acuerdo a las instrucciones del kit. </li><li> Determinar el diseño de placas y pipeta de 25 l de normas y controles en los pozos apropiados por duplicado. </li><li> Descongele las muestras de saliva, vortex y centrifugar a 1500 XG (@ 3000 RPM) durante 15 minutos. </li><li> Pipeta de 25 l de líquido claro de las muestras se centrifugaron en los pozos apropiados en la placa (por duplicado). </li><li> Prepare la enzima conjugada y la pipeta 200 l en todos los pocillos. </li&gt; <li> Mezcla de la placa durante 5 minutos a 500 rpm y se incuba durante otros 55 minutos. </li><li> Lave y seque la placa 4 veces con tampón de lavado. </li><li> Añadir 200 l tetrametilbenzidina (TMB) a cada pocillo utilizando una pipeta multicanal. </li><li> Mezcla de la placa durante 5 minutos a 500 rpm y se incuban en la oscuridad durante 25 minutos adicionales. </li><li> Añadir 50 l de solución de parada a cada pocillo y mezclar durante 3 minutos a 500 rpm para detener la reacción. </li><li> Limpie la parte inferior del plato limpio y leer la placa en un lector de placas a 450 nm a los 10 minutos de la adición de la solución de parada. </li><li> Para calcular la concentración de cortisol, calcular la densidad óptica media para todos los pocillos. Resta la densidad óptica media de los pozos de unión no específica de la densidad óptica media del cero, el control estándar, y los pozos de la muestra. </li><li> Calcular el porcentaje límite (B / Bo) para cada estándar, control y muestra dividiendo la densidad óptica media de estos pozos (B) por el promedioedad de densidad óptica de los pozos de cero (Bo). Determinar la concentración de los controles y las muestras mediante interpolación utilizando un software capaz de logística. </li></ol> 7. Los resultados representativos: Los datos representativos que aquí se presenta es una compilación de revisión de la literatura y dos estudios que se realizaron en nuestro laboratorio durante la tarde con los participantes adultos sanos de sexo masculino. Estos datos están destinadas a representar aproximadamente los resultados que se pueden encontrar a través del protocolo presentado en este artículo. Este protocolo TSST (Figura 1) induce un incremento de auto-reporte de la ansiedad (medida por el Inventario de Ansiedad Estado Rasgo, Figura 2), la frecuencia cardiaca (Figura 3), y las concentraciones de cortisol en saliva (Figura 4). El protocolo experimental se ilustra en la Figura 1 tiene por objeto proporcionar un ejemplo de cómo las diferentes medidas pueden ser hechas durante una sesión de recogida de datos con la TSST. El momento de la MEDIDAts y la inclusión de diferentes medidas será dictado por la pregunta de investigación específica. Si se utiliza una intervención que tiene por objeto reducir la ansiedad y / o respuesta al estrés, aumenta desapercibida o atenuada en estas medidas se prevé. <img alt="Figura 1" src="/files/ftp_upload/3238/3238fig1.jpg" /> Figura 1. Protocolo experimental Nota: Prueba de Estrés Social Trier (TSST); Estado-Rasgo de Ansiedad (STAI), 5 minutos en período de estrés pre (PS), el período de estrés anticipatorio de 10 minutos (preparación del discurso) (AS), 5 minutos en período de voz. (S), 5 minutos período de Matemáticas (M). <img alt="Figura 2" src="/files/ftp_upload/3238/3238fig2.jpg" /> Figura 2. Representante de las puntuaciones de ansiedad estado. Nota: puntuación de Ansiedad Estado calcula a partir de 20 Iten medir la ansiedad estado de Inventario de Ansiedad Estado Rasgo (STAI) Representante resultados reflejan los datos recogidos en nuestro laboratorio y publicados en la literatura.. Figura 3. Representante de datos de frecuencia cardiaca Nota:.. La frecuencia cardíaca en latidos por minuto (BPM) Valores representativos reflejan los datos recogidos en nuestro laboratorio y publicados en la literatura. <img alt="Figura 4" src="/files/ftp_upload/3238/3238fig4.jpg" /> Figura 4. Datos representativos cortisol salival Nota:.. La concentración de cortisol en UL / DL valores representativos reflejan los datos recogidos en nuestro laboratorio y publicados en la literatura.

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S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessment+of+salivary+hormones.">Assessment of salivary hormones.</a> Methods in Social Neuroscience. , 17-44 (2009).</li><li>Kudielka, B. M., Hellhammer, D. H., W&#252;st, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Why+do+we+respond+so+differently?+Reviewing+determinants+of+human+salivary+cortisol+responses+to+challenge.">Why do we respond so differently? Reviewing determinants of human salivary cortisol responses to challenge.</a> Psychoneuroendocrinology. 34, 2-18 (2009).</li><li>Buske-Kirschbaum, A., Jobst, S., Wustmans, A., Kirschbaum, C., Rauh, W., Hellhammer, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Attenuated+free+cortisol+response+to+psychosocial+stress+in+children+with+atopic+dermatitis.">Attenuated free cortisol response to psychosocial stress in children with atopic dermatitis.</a> Psychosom. Med. 59, 419-426 (1997).</li></ol>

No hay conflictos de interés declarado.

En este artículo, hemos demostrado cómo llevar a cabo la prueba de Trier estrés social en un voluntario sano. El TSST es un factor estresante de laboratorio estandarizadas social que induce un incremento robusto y fiable de las medidas psicológicas, fisiológicas y neuroendocrinos. El TSST es una alternativa útil a los estresores físicos, en la prueba de prensa-frío o caminata en cinta rodante, y se reproduce la tensión psicológica más naturalista de rendimiento en presencia de una audiencia de evaluación. </...

<table border="1"><tbody><tr><th> Nombre del equipo / suministro </th><th> Empresa </th><th> Número de catálogo </th></tr><tr><td> BSL Psicofisiología del sistema, incluyendo Mac OS MP36 Unidad de Adquisición de Datos y tres electrodos de electrocardiograma </td><td> Biopac Systems Inc. </td><td> BSLPSY-M </td></tr><tr><td> Inventario de Ansiedad Estado-Rasgo </td><td> Mente Jardín, Inc. </td><td> Conservador-B </td></tr><tr><td> Enzima de cortisol en saliva Inmunoensayo Kit </td><td> Salimetrics </td><td> 1-3002 </td></tr><tr><td> 2,0 ml de polipropileno viales </td><td> Fisher Scientific </td><td> 05 40B 146 </td></tr><tr><td> Lector de microplacas BioRad, modelo 680, con software de gestión de microplacas </td><td> BioRad </td><td> Lector de Placas: 168 1000 Software: 1706800 </td></tr></tbody></table>

the trier social stress test protocol for inducing psychological stress

En este artículo muestra un protocolo de estrés psicológico para su uso en un entorno de laboratorio. Protocolos que permiten a los investigadores a estudiar los procesos biológicos de la respuesta al estrés en la salud y la enfermedad son fundamentales para el progreso de la investigación en el estrés y la ansiedad. 1 Aunque existen numerosos protocolos para la inducción de la respuesta al estrés en el laboratorio, el abandono muchas de proporcionar un contexto naturalista o para incorporar los aspectos de la tensión social y psicológico. De los protocolos de estrés psicológico, el meta-análisis sugiere que la prueba de estrés social Trier (TSST) es el protocolo estándar más apropiado y útil para los estudios de la reactividad de la hormona del estrés. 2 En la descripción original de la TSST, los investigadores trataron de diseñar y evaluar un procedimiento de capaz de inducir una respuesta de estrés fiable en la mayoría de los voluntarios sanos. 3 Estos investigadores encontraron elevaciones de la frecuencia cardiaca, presión arterial y el estrés endocrinas markers en respuesta a la TSST (un factor de estrés psicológico) en comparación con una inyección de solución salina (un factor de estrés físico). 3 Aunque la TSST ha sido modificado para satisfacer las necesidades de diversos grupos de investigación, por lo general consiste en un período de espera a la llegada, el discurso de anticipación preparación, el habla de rendimiento, y la aritmética verbal períodos de desempeño, seguido de uno o más períodos de recuperación. La TSST exige a los participantes para preparar y pronunciar un discurso, verbal y responder a un problema de aritmética difícil en presencia de un público socialmente evaluación. 3 Evaluación social y falta de control han sido identificados como componentes clave de la inducción de estrés por la TSST. 4 En el uso de más de una década, el objetivo de la TSST es inducir una respuesta de estrés de manera sistemática con el fin de medir las diferencias en la reactividad, la ansiedad y la activación del eje hipotálamo-pituitario-adrenal (HPA) o simpático-adrenal medular (SAM), el eje en la tarea 1. Researcella en general, evaluar los cambios en el auto-reporte de la ansiedad, las medidas fisiológicas (por ejemplo, tasa cardiaca), y / o índices neuroendocrinos (por ejemplo, la hormona del estrés cortisol) en respuesta a la TSST. Muchos investigadores han adoptado muestras salivales de los marcadores de estrés como el cortisol y la alfa-amilasa (un marcador de activación del sistema nervioso autónomo) como alternativa a la extracción de sangre para reducir el estrés de confusión de las técnicas de recogida de sangre. Además de los cambios experimentados por un individuo de completar la TSST, los investigadores pueden comparar los cambios entre diferentes grupos de tratamiento (por ejemplo, las muestras clínicas en comparación con controles sanos) o la eficacia de las intervenciones para reducir el estrés 1.

Watch this Scientific Journal Video about The Trier Social Stress Test Protocol for Inducing Psychological Stress at JoVE.com

The Trier Social Stress Test Protocol for Inducing Psychological Stress

En este artículo se describe un protocolo para la inducción de estrés psicológico en los participantes, lo que permite a los investigadores para medir las respuestas psicológicas, fisiológicas y neuroendocrina al estrés en los participantes o entre grupos.

El estrés social Trier Protocolo de prueba para la inducción de estrés psicológico

This article demonstrates a psychological stress protocol for use in a laboratory setting.  Protocols that allow researchers to study the biological ...

the-trier-social-stress-test-protocol-for-inducing-psychological

Research

JoVE Journal

Medicine

55.6K vistas.  Northern Arizona University. En este artículo se describe un protocolo para la inducción de estrés psicológico en los participantes, lo que permite a los investigadores para medir las respuestas psicológicas, fisiológicas y neuroendocrina al estrés en los participantes o entre grupos.

Video: The Trier Social Stress Test Protocol for Inducing Psychological Stress

Using Visual and Narrative Methods to Achieve Fair Process in Clinical Care

The Institute of Medicine has targeted patient-centeredness as an important area of quality improvement. A major dimension of patient-centeredness is respect for patient's values, preferences, and expressed needs. Yet specific approaches to gaining this understanding and translating it to quality care in the clinical setting are lacking. From a patient perspective quality is not a simple concept but is best understood in terms of five dimensions: technical outcomes; decision-making efficiency; amenities and convenience; information and emotional support; and overall patient satisfaction. Failure to consider quality from this five-pronged perspective results in a focus on medical outcomes, without considering the processes central to quality from the patient's perspective and vital to achieving good outcomes. In this paper, we argue for applying the concept of fair process in clinical settings. Fair process involves using a collaborative approach to exploring diagnostic issues and treatments with patients, explaining the rationale for decisions, setting expectations about roles and responsibilities, and implementing a core plan and ongoing evaluation. Fair process opens the door to bringing patient expertise into the clinical setting and the work of developing health care goals and strategies. This paper provides a step by step illustration of an innovative visual approach, called photovoice or photo-elicitation, to achieve fair process in clinical work with acquired brain injury survivors and others living with chronic health conditions. Applying this visual tool and methodology in the clinical setting will enhance patient-provider communication; engage patients as partners in identifying challenges, strengths, goals, and strategies; and support evaluation of progress over time. Asking patients to bring visuals of their lives into the clinical interaction can help to illuminate gaps in clinical knowledge, forge better therapeutic relationships with patients living with chronic conditions such as brain injury, and identify patient-centered goals and possibilities for healing. The process illustrated here can be used by clinicians, (primary care physicians, rehabilitation therapists, neurologists, neuropsychologists, psychologists, and others) working with people living with chronic conditions such as acquired brain injury, mental illness, physical disabilities, HIV/AIDS, substance abuse, or post-traumatic stress, and by leaders of support groups for the types of patients described above and their family members or caregivers.

This paper illustrates an innovative visual approach (photovoice or photo-elicitation) to achieve fair process in clinical care for patients living with chronic health conditions, illuminate gaps in clinical knowledge, forge better therapeutic relationships, and identify patient-centered goals and possibilities for healing.

Uso de métodos visuales y narrativos para lograr un proceso justo en la atención clínica

The Institute of Medicine has targeted patient-centeredness as an important area of quality improvement. A major dimension of patient-centeredness is ...

Investigación

Medicina

Development of a Unilaterally-lesioned 6-OHDA Mouse Model of Parkinson's Disease

The unilaterally lesioned 6-hyroxydopamine (6-OHDA)-lesioned rat model of Parkinson's disease (PD) has proved to be invaluable in advancing our understanding of the mechanisms underlying parkinsonian symptoms, since it recapitulates the changes in basal ganglia circuitry and pharmacology observed in parkinsonian patients1-4. However, the precise cellular and molecular changes occurring at cortico-striatal synapses of the output pathways within the striatum, which is the major input region of the basal ganglia remain elusive, and this is believed to be site where pathological abnormalities underlying parkinsonian symptoms arise3,5.
In PD, understanding the mechanisms underlying changes in basal ganglia circuitry following degeneration of the nigro-striatal pathway has been greatly advanced by the development of bacterial artificial chromosome (BAC) mice over-expressing green fluorescent proteins driven by promoters specific for the two striatal output pathways (direct pathway: eGFP-D1; indirect pathway: eGFP-D2 and eGFP-A2a)8, allowing them to be studied in isolation. For example, recent studies have suggested that there are pathological changes in synaptic plasticity in parkinsonian mice9,10. However, these studies utilised juvenile mice and acute models of parkinsonism. It is unclear whether the changes described in adult rats with stable 6-OHDA lesions also occur in these models. Other groups have attempted to generate a stable unilaterally-lesioned 6-OHDA adult mouse model of PD by lesioning the medial forebrain bundle (MFB), unfortunately, the mortality rate in this study was extremely high, with only 14% surviving the surgery for 21 days or longer11. More recent studies have generated intra-nigral lesions with both a low mortality rate &gt;80% loss of dopaminergic neurons, however expression of L-DOPA induced dyskinesia11,12,13,14 was variable in these studies. Another well established mouse model of PD is the MPTP-lesioned mouse15. Whilst this model has proven useful in the assessment of potential neuroprotective agents16, it is less suitable for understanding mechanisms underlying symptoms of PD, as this model often fails to induce motor deficits, and shows a wide variability in the extent of lesion17, 18.
Here we have developed a stable unilateral 6-OHDA-lesioned mouse model of PD by direct administration of 6-OHDA into the MFB, which consistently causes &gt;95% loss of striatal dopamine (as measured by HPLC), as well as producing the behavioural imbalances observed in the well characterised unilateral 6-OHDA-lesioned rat model of PD. This newly developed mouse model of PD will prove a valuable tool in understanding the mechanisms underlying generation of parkinsonian symptoms.

Development of a Unilaterally-lesioned 6-OHDA Mouse Model of Parkinson&#039;s Disease

A protocol for performing unilateral 6-OHDA lesions of the medial forebrain bundle in mice is described. This method has a low mortality rate (13.3 %) with 89% of the surviving animals showing &#62;95% loss of striatal dopamine and 90.63&#177;-4.02 % ipsiversive rotational bias towards the side of the lesion.

Desarrollo de una manera unilateral-lesionado 6-OHDA modelo de ratón de la enfermedad de Parkinson

The unilaterally lesioned 6-hyroxydopamine (6-OHDA)-lesioned rat model of Parkinson's disease (PD) has proved to be invaluable in advancing our ...

Implantation of a Carotid Cuff for Triggering Shear-stress Induced Atherosclerosis in Mice

It is widely accepted that alterations in vascular shear stress trigger the expression of inflammatory genes in endothelial cells and thereby induce atherosclerosis (reviewed in 1 and 2). The role of shear stress has been extensively studied in vitro investigating the influence of flow dynamics on cultured endothelial cells 1,3,4 and in vivo in larger animals and humans 1,5,6,7,8. However, highly reproducible small animal models allowing systematic investigation of the influence of shear stress on plaque development are rare. Recently, Nam et al. 9 introduced a mouse model in which the ligation of branches of the carotid artery creates a region of low and oscillatory flow. Although this model causes endothelial dysfunction and rapid formation of atherosclerotic lesions in hyperlipidemic mice, it cannot be excluded that the observed inflammatory response is, at least in part, a consequence of endothelial and/or vessel damage due to ligation.
In order to avoid such limitations, a shear stress modifying cuff has been developed based upon calculated fluid dynamics, whose cone shaped inner lumen was selected to create defined regions of low, high and oscillatory shear stress within the common carotid artery 10. By applying this model in Apolipoprotein E (ApoE) knockout mice fed a high cholesterol western type diet, vascular lesions develop upstream and downstream from the cuff. Their phenotype is correlated with the regional flow dynamics 11 as confirmed by in vivo Magnetic Resonance Imaging (MRI) 12: Low and laminar shear stress upstream of the cuff causes the formation of extensive plaques of a more vulnerable phenotype, whereas oscillatory shear stress downstream of the cuff induces stable atherosclerotic lesions 11. In those regions of high shear stress and high laminar flow within the cuff, typically no atherosclerotic plaques are observed.
 
In conclusion, the shear stress-modifying cuff procedure is a reliable surgical approach to produce phenotypically different atherosclerotic lesions in ApoE-deficient mice.

The constricting cuff presented in this article is designed to induce atherosclerosis in the murine common carotid artery. Due to the conical shape of its inner lumen the implanted cuff generates well-defined regions of low, high and oscillatory shear stress triggering the development of atherosclerotic lesions of different inflammatory phenotypes.

Implantación de un manguito de la carótida para provocar esfuerzo cortante aterosclerosis inducida en ratones

It is widely accepted that alterations in vascular shear stress trigger the expression of inflammatory genes in endothelial cells and thereby induce ...

Biomarkers in an Animal Model for Revealing Neural, Hematologic, and Behavioral Correlates of PTSD

Identification of biomarkers representing the evolution of the pathophysiology of Post Traumatic Stress Disorder (PTSD) is vitally important, not only for objective diagnosis but also for the evaluation of therapeutic efficacy and resilience to trauma. Ongoing research is directed at identifying molecular biomarkers for PTSD, including traumatic stress induced proteins, transcriptomes, genomic variances and genetic modulators, using biologic samples from subjects' blood, saliva, urine, and postmortem brain tissues. However, the correlation of these biomarker molecules in peripheral or postmortem samples to altered brain functions associated with psychiatric symptoms in PTSD remains unresolved. Here, we present an animal model of PTSD in which both peripheral blood and central brain biomarkers, as well as behavioral phenotype, can be collected and measured, thus providing the needed correlation of the central biomarkers of PTSD, which are mechanistic and pathognomonic but cannot be collected from people, with the peripheral biomarkers and behavioral phenotypes, which can.
Our animal model of PTSD employs restraint and tail shocks repeated for three continuous days - the inescapable tail-shock model (ITS) in rats. This ITS model mimics the pathophysiology of PTSD 17, 7, 4, 10. We and others have verified that the ITS model induces behavioral and neurobiological alterations similar to those found in PTSD subjects 17, 7, 10, 9. Specifically, these stressed rats exhibit (1) a delayed and exaggerated startle response appearing several days after stressor cessation, which given the compressed time scale of the rat's life compared to a humans, corresponds to the one to three months delay of symptoms in PTSD patients (DSM-IV-TR PTSD Criterian D/E 13), (2) enhanced plasma corticosterone (CORT) for several days, indicating compromise of the hypothalamopituitary axis (HPA), and (3) retarded body weight gain after stressor cessation, indicating dysfunction of metabolic regulation.
The experimental paradigms employed for this model are: (1) a learned helplessness paradigm in the rat assayed by measurement of acoustic startle response (ASR) and a charting of body mass; (2) microdissection of the rat brain into regions and nuclei; (3) enzyme-linked immunosorbent assay (ELISA) for blood levels of CORT; (4) a gene expression microarray plus related bioinformatics tools 18. This microarray, dubbed rMNChip, focuses on mitochondrial and mitochondria-related nuclear genes in the rat so as to specifically address the neuronal bioenergetics hypothesized to be involved in PTSD.

We describe a rat model of post traumatic stress disorder (PTSD) that reveals the persistent alterations in neuroendocrine function and the delayed long-term, exaggerated fear response, characteristic of PTSD patients. The animal model and methods described here are useful for correlating biomarkers in brain nuclei, which are mechanistic but cannot be measured in patients, with biomarkers in peripheral white blood cells, which can.

Biomarcadores en un modelo animal para Revelar correlatos neurales, hematológicas y conductuales de estrés postraumático

Identification of biomarkers representing the evolution of the pathophysiology of Post Traumatic Stress Disorder (PTSD) is vitally important, not only for ...

Cardiac Stress Test Induced by Dobutamine and Monitored by Cardiac Catheterization in Mice

Dobutamine is a &beta;-adrenergic agonist with an affinity higher for receptor expressed in the heart (&beta;1) than for receptors expressed in the arteries (&beta;2). When systemically administered, it increases cardiac demand. Thus, dobutamine unmasks abnormal rhythm or ischemic areas potentially at risk of infarction. 
Monitoring of heart function during a cardiac stress test can be performed by either ecocardiography or cardiac catheterization. The latter is an invasive but more accurate and informative technique that the former.
Cardiac stress test induced by dobutamine and monitored by cardiac catheterization accomplished as described here allows, in a single experiment, the measurement of the following hemodynamic parameters: heart rate (HR), systolic pressure, diastolic pressure, end-diastolic pressure, maximal positive pressure development (dP/dtmax) and maximal negative pressure development (dP/dtmin), at baseline conditions and under increasing doses of dobutamine.
As expected, in normal mice we observed a dobutamine dose-related increase in HR, dP/dtmax and dP/dtmin. Moreover, at the highest dose tested (12 ng/g/min) the cardiac decompensation of high fat diet-induced obese mice was unmasked.

We describe the protocol to perform a cardiac stress test induced by dobutamine and monitored by cardiac catheterization in normal mice. Also we show its application to unmask subclinical cardiac disease in high fat diet-induced obese mice.

Prueba de esfuerzo cardíaco inducida por dobutamina y supervisado por cateterismo cardíaco en ratones

Dobutamine is a &beta;-adrenergic agonist with an affinity higher for receptor expressed in the heart (&beta;1) than for receptors expressed in the ...

Reduction in Left Ventricular Wall Stress and Improvement in Function in Failing Hearts using Algisyl-LVR

Injection of Algisyl-LVR, a treatment under clinical development, is intended to treat patients with dilated cardiomyopathy. This treatment was recently used for the first time in patients who had symptomatic heart failure. In all patients, cardiac function of the left ventricle (LV) improved significantly, as manifested by consistent reduction of the LV volume and wall stress. Here we describe this novel treatment procedure and the methods used to quantify its effects on LV wall stress and function.
Algisyl-LVR is a biopolymer gel consisting of Na+-Alginate and Ca2+-Alginate. The treatment procedure was carried out by mixing these two components and then combining them into one syringe for intramyocardial injections. This mixture was injected at 10 to 19 locations mid-way between the base and apex of the LV free wall in patients.
Magnetic resonance imaging (MRI), together with mathematical modeling, was used to quantify the effects of this treatment in patients before treatment and at various time points during recovery. The epicardial and endocardial surfaces were first digitized from the MR images to reconstruct the LV geometry at end-systole and at end-diastole. Left ventricular cavity volumes were then measured from these reconstructed surfaces.
Mathematical models of the LV were created from these MRI-reconstructed surfaces to calculate regional myofiber stress. Each LV model was constructed so that 1) it deforms according to a previously validated stress-strain relationship of the myocardium, and 2) the predicted LV cavity volume from these models matches the corresponding MRI-measured volume at end-diastole and end-systole. Diastolic filling was simulated by loading the LV endocardial surface with a prescribed end-diastolic pressure. Systolic contraction was simulated by concurrently loading the endocardial surface with a prescribed end-systolic pressure and adding active contraction in the myofiber direction. Regional myofiber stress at end-diastole and end-systole was computed from the deformed LV based on the stress-strain relationship.

This article describes procedures for implanting a novel hydrogel in failing hearts and quantifying its effect on left ventricular wall stress and function. These procedures have been successfully applied in dogs and humans.

Reducción del estrés de la pared del ventrículo izquierdo y la mejoría de la función en su defecto corazones con Algisyl LVR-

Injection of  Algisyl-LVR, a treatment under clinical development, is intended to treat  patients with dilated cardiomyopathy. This treatment was recently ...

A Mouse Tumor Model of Surgical Stress to Explore the Mechanisms of Postoperative Immunosuppression and Evaluate Novel Perioperative Immunotherapies

Surgical resection is an essential treatment for most cancer patients, but surgery induces dysfunction in the immune system and this has been linked to the development of metastatic disease in animal models and in cancer patients. Preclinical work from our group and others has demonstrated a profound suppression of innate immune function, specifically NK cells in the postoperative period and this plays a major role in the enhanced development of metastases following surgery. Relatively few animal studies and clinical trials have focused on characterizing and reversing the detrimental effects of cancer surgery. Using a rigorous animal model of spontaneously metastasizing tumors and surgical stress, the enhancement of cancer surgery on the development of lung metastases was demonstrated. In this model, 4T1 breast cancer cells are implanted in the mouse mammary fat pad. At day 14 post tumor implantation, a complete resection of the primary mammary tumor is performed in all animals. A subset of animals receives additional surgical stress in the form of an abdominal nephrectomy. At day 28, lung tumor nodules are quantified. When immunotherapy was given immediately preoperatively, a profound activation of immune cells which prevented the development of metastases following surgery was detected. While the 4T1 breast tumor surgery model allows for the simulation of the effects of abdominal surgical stress on tumor metastases, its applicability to other tumor types needs to be tested. The current challenge is to identify safe and promising immunotherapies in preclinical mouse models and to translate them into viable perioperative therapies to be given to cancer surgery patients to prevent the recurrence of metastatic disease.

A mouse tumor model of surgical stress is used to explore how postoperative immune suppression promotes metastatic disease and to evaluate immunostimulatory perioperative therapies.

Un modelo de tumor de Estrés quirúrgico para explorar los mecanismos de inmunosupresión postoperatoria y Evaluar Novel perioperatorias Inmunoterapias Ratón

Surgical resection is an essential treatment for most cancer patients, but surgery induces dysfunction in the immune system and this has been linked to ...

A Modified In vitro Invasion Assay to Determine the Potential Role of Hormones, Cytokines and/or Growth Factors in Mediating Cancer Cell Invasion

Blood serum serves as a chemoattractant towards which cancer cells migrate and invade, facilitating their intravasation into microvessels. However, the actual molecules towards which the cells migrate remain elusive. This modified invasion assay has been developed to identify targets which drive cell migration and invasion. This technique compares the invasion index under three conditions to determine whether a specific hormone, growth factor, or cytokine plays a role in mediating the invasive potential of a cancer cell. These conditions include i) normal fetal bovine serum (FBS), ii) charcoal-stripped FBS (CS-FBS), which removes hormones, growth factors, and cytokines and iii) CS-FBS + molecule (denoted &#8220;X&#8221;). A significant change in cell invasion with CS-FBS as compared to FBS, indicates the involvement of hormones, cytokines or growth factors in mediating the change. Individual molecules can then be added back to CS-FBS to assay their ability to reverse or rescue the invasion phenotype. Furthermore, two or more factors can be combined to evaluate the additive or synergistic effects of multiple molecules in driving or inhibiting invasion. Overall, this method enables the investigator to determine whether hormones, cytokines, and/or growth factors play a role in cell invasion by serving as chemoattractants or inhibitors of invasion for a particular type of cancer cell or a specific mutant. By identifying specific chemoattractants and inhibitors, this modified invasion assay may help to elucidate signaling pathways that direct cancer cell invasion.

A Modified In vitro Invasion Assay to Determine the Potential Role of Hormones, Cytokines and/or Growth Factors in Mediating Cancer Cell Invasion

This video article describes a modified in vitro method to examine the role of hormones, cytokines and/or growth factors in driving cancer cell invasion.

Un Modificado<em&gt; In vitro</em&gt; Invasión Ensayo para determinar el papel potencial de hormonas, citoquinas y / o factores de crecimiento en la invasión de células del cáncer de Mediación

Blood serum serves as a chemoattractant towards which cancer cells migrate and invade, facilitating their intravasation into microvessels. However, the ...

Glutamate and Hypoxia as a Stress Model for the Isolated Perfused Vertebrate Retina

Neuroprotection has been a strong field of investigation in ophthalmological research in the past decades and affects diseases such as glaucoma, retinal vascular occlusion, retinal detachment, and diabetic retinopathy. It was the object of this study to introduce a standardized stress model for future preclinical therapeutic testing.
Bovine retinas were prepared and perfused with an oxygen saturated standard solution, and the ERG was recorded. After recording stable b-waves, hypoxia (pure N2) or glutamate stress (250 &#181;m glutamate) was exerted for 45 min. To investigate the effects on photoreceptor function alone, 1 mM aspartate was added to obtain a-waves. ERG-recovery was monitored for 75 min.
For hypoxia, a decrease in a-wave amplitude of 87.0% was noted (p &#60;0.01) after an exposition time of 45 min (decrease of 36.5% after the end of the washout p = 0.03). Additionally, an initial decrease in b-wave amplitudes of 87.23% was recorded, that reached statistical significance (p &#60;0.01, decrease of 25.5% at the end of the washout, p = 0.03).
For 250 &#181;m glutamate, an initial 7.8% reduction of a-wave amplitudes (p &#62;0.05) followed by a reduction of 1.9% (p &#62;0.05). A reduction of 83.7% of b-wave amplitudes (p &#60;0.01) was noted; after a washout of 75 min the reduction was 2.3% (p = 0.62). In this study, a standardized stress model is presented that may be useful to identify possible neuroprotective effects in the future.

With this study, we introduce a standardized stress model for the isolated superfused bovine retina for future preclinical therapeutic testing. The effect of either hypoxia (pure N2) or glutamate stress (250 &#181;M glutamate) on retinal function represented by a- and b-wave amplitudes was evaluated.

El glutamato y la hipoxia como un Modelo de estrés para el perfundido aislado Vertebrados Retina

Neuroprotection has been a strong field of investigation in ophthalmological research in the past decades and affects diseases such as glaucoma, retinal ...

Induction and Analysis of Oxidative Stress in Sleeping Beauty Transposon-Transfected Human Retinal Pigment Epithelial Cells

Oxidative stress plays a critical role in several degenerative diseases, including age-related macular degeneration (AMD), a pathology that affects ~30 million patients worldwide. It leads to a decrease in retinal pigment epithelium (RPE)-synthesized neuroprotective factors, e.g., pigment epithelium-derived factor (PEDF) and granulocyte-macrophage colony-stimulating factor (GM-CSF), followed by the loss of RPE cells, and eventually photoreceptor and retinal ganglion cell (RGC) death. We hypothesize that the reconstitution of the neuroprotective and neurogenic retinal environment by the subretinal transplantation of transfected RPE cells overexpressing PEDF and GM-CSF has the potential to prevent retinal degeneration by mitigating the effects of oxidative stress, inhibiting inflammation, and supporting cell survival. Using the Sleeping Beauty transposon system (SB100X) human RPE cells have been transfected with the PEDF and GM-CSF genes and shown stable gene integration, long-term gene expression, and protein secretion using qPCR, western blot, ELISA, and immunofluorescence. To confirm the functionality and the potency of the PEDF and GM-CSF secreted by the transfected RPE cells, we have developed an in vitro assay to quantify the reduction of H2O2-induced oxidative stress on RPE cells in culture. Cell protection was evaluated by analyzing cell morphology, density, intracellular level of glutathione, UCP2 gene expression, and cell viability. Both, transfected RPE cells overexpressing PEDF and/or GM-CSF and cells non-transfected but pretreated with PEDF and/or GM-CSF (commercially available or purified from transfected cells) showed significant antioxidant cell protection compared to non-treated controls. The present H2O2-model is a simple and effective approach to evaluate the antioxidant effect of factors that may be effective to treat AMD or similar neurodegenerative diseases.

Induction and Analysis of Oxidative Stress in Sleeping Beauty Transposon-Transfected Human Retinal Pigment Epithelial Cells

We present a protocol for the development and use ofan oxidative stress-model by treating retinal pigment epithelial cells with H2O2, analyzing cell morphology, viability, density, glutathione, and UCP-2 level. It is a useful model to investigate the antioxidant effect of proteins secreted by transposon-transfected cells to treat neuroretinal degeneration.

Inducción y análisis del estrés oxidativo en células epiteliales pigmentarias de la retina humana transfectadas por transposones de la bella durmiente

Oxidative stress plays a critical role in several degenerative diseases, including age-related macular degeneration (AMD), a pathology that affects ~30 ...