Parte 1: La creación y la preparación de la piel <ol><li> Configure los siguientes sensores de respiración cinturón, EMG, ECG, conductancia de la piel, temperatura, suelo; </li><li> Prepare la piel frotando suavemente con una toallita de alcohol en los siguientes lugares: frente, arriba de los ojos de EMG, en el interior de las muñecas para un ECG, dorso de la mano izquierda para el suelo (ubicación de los electrodos de tierra puede variar dependiendo del equipo utilizado); </li><li> Los sensores conectados a la piel en los siguientes lugares: la respiración cinturón alrededor de la cintura, EMG en la frente por encima de cada ojo, ECG en el interior de cada muñeca, conductancia de la piel en el dedo medio de la mano izquierda, un termómetro en el puntero de la mano izquierda, baja en la parte posterior de la mano izquierda; </li><li> Seleccione la sesión de respiración VFC +; </li><li> Asegúrese de que impedancia es aceptable para todos los sensores y los sensores que responden al movimiento de clientes y produciendo lecturas razonables. </li></ol> Parte 2: Grabación <ol><li> Explicar el procedimiento al cliente. </li><li> (Min 0-2) Instruya a los clientes a sentarse en silencio con los ojos abiertos. Registro de una base de dos minutos. </li><li> (Min 2-4) Al cabo de dos minutos, comienza un factor de estrés de dos minutos cognitivo - problemas aritméticos aumentando gradualmente en dificultad para ser resuelto sin el uso de lápiz y papel o calculadora (si la aritmética es particularmente difícil para el cliente, pídales para restar siete veces a partir de 100, es decir, &quot;siete de serie&quot;). Dar las siguientes instrucciones para el cliente: &quot;En los próximos 2 minutos, se le pedirá que resolver algunos problemas de matemáticas. Por favor, trabajar lo más pronto posible y me dicen que la respuesta &quot;. No proporcione información sobre si la respuesta del cliente es correcta o incorrecta, así como alentar mínima (por ejemplo, &quot;Haz lo mejor que puede&quot;) a fin de no eliminar las propiedades de la inducción de estrés leve de este ejercicio. </li><li> (Min 04.06) Que el cliente sepa que esta parte de la evaluación es más y pida que él / ella se sientan en silencio con los ojos abiertos durante 2 minutos. </li><li> (Min 6-8) Comienza estrés físico: a su vez sobre el ruido nocivo fuertes (los bebés que lloran, los accidentes de coche, etc) y pedir al cliente que escuchar los sonidos sin desconcentra. </li><li> (Min 8-10) Apague el ruido, y pedir al cliente que se siente tranquilamente con los ojos abiertos durante 2 minutos. </li><li> (Min 12.10) Comience estrés emocional: pedir al cliente que habla de un incidente estresante que recuerdo muy bien, son los detalles del evento, lo que estaban pensando y sintiendo. Proporcionar una respuesta empática mínima para no reducir la respuesta al estrés. </li><li> (Min 12.14) Interrumpir la historia del cliente, incluso si él / ella no ha terminado, y enseñar al paciente a sentarse en silencio con los ojos abiertos. </li><li> Detener la grabación, guardar los datos, y dejar que el cliente sepa que la evaluación se lleva a cabo. </li></ol> Parte 3: Informe y la planificación del tratamiento <ol><li> Pida al cliente acerca de su experiencia durante la evaluación. ¿Qué partes fueron muy tensas y por qué? Él / ella siente las alteraciones fisiológicas pasando? </li><li> Revisión de datos de acuerdo a los criterios descritos en el anexo I. </li><li> Elija los parámetros fisiológicos que deben abordarse sobre la base de análisis anteriores. Si la lectura del sensor en la línea de base es mayor o menor de lo normal, el objetivo del tratamiento es ayudar al cliente a lograr que los parámetros fisiológicos a la normalidad en un ambiente no estresante. Si la lectura del sensor durante el estrés es mayor o menor de lo normal, el objetivo del tratamiento es ayudar al cliente a identificar los factores de estrés y ser capaz de mantener el nivel normal de funcionamiento fisiológico de estrés. Si la lectura del sensor durante el tiempo de abajo indica la falta de recuperación, el objetivo del tratamiento es ayudar al cliente a lograr que los parámetros fisiológicos a un nivel normal rápidamente después de un estresante. Cualquiera o todos estos objetivos pueden ser incluidos en el plan de tratamiento. </li></ol> Apéndice I Directrices para la evaluación de estrés interpretación (Todos los &quot;audaces&quot; respuestas indican las áreas a tratar) <table border="0"><tbody><tr><td colspan="3"> EMG (lectura normal &lt;3 microvoltios) </td></tr><tr><td> 1. EMG es elevado al inicio del estudio? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 2. EMG es elevada más allá de la línea de base en cualquiera de los tres factores de estrés? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 3. ¿Existe la recuperación del GGA a nivel de línea de base durante los periodos de descanso? </td><td> Sí _ </td><td> no _ </td></tr><tr><td colspan="3"> La frecuencia cardiaca (normal 60 a 80 latidos por minuto) </td></tr><tr><td> 1. Es la frecuencia cardíaca elevada al inicio del estudio? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 2. Es elevado ritmo cardíaco más allá de la línea de base en cualquiera de los tres factores de estrés? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 3. ¿Existe la recuperación de la frecuencia cardiaca a nivel de línea de base durante los periodos de descanso? </td><td> Sí _ </td><td> no _ </td></tr><tr><td colspan="3"> La frecuencia respiratoria (tarifa normal &lt;12 respiraciones por minuto) </td></tr><tr><td> 1. Es la frecuencia respiratoria elevada al inicio del estudio? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 2. Tasa de respiración se eleva más allá de la línea de base para cualquier factor estresante? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 3. ¿Existe la recuperación de la frecuencia respiratoria de base durante los periodos de descanso? </td><td> Sí _ </td><td> no _ </td></tr><tr><td colspan="3"> Conductancia de la piel </td></tr><tr><td> 1. Conductancia de la piel es elevada al inicio del estudio? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 2. Conductancia de la piel se eleva más allá de la línea de base para cualquier factor estresante? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 3. ¿Existe la recuperación de la conductancia de la piel a la línea de base durante los periodos de descanso? </td><td> Sí _ </td><td> no _ </td></tr><tr><td colspan="3"> Los dedos de temperatura (lectura normal&gt; 90 ° F) </td></tr><tr><td> 1. Es la temperatura del dedo bajo al inicio del estudio? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 2. Se reduce la temperatura dedo más allá de la línea de base para cualquier factor estresante? </td><td> y es _ </td><td> no _ </td></tr><tr><td> 3. ¿Existe la recuperación de la temperatura de los dedos a la línea base durante periodos de descanso? </td><td> Sí _ </td><td> no _ </td></tr><tr><td colspan="3"> Variabilidad del ritmo cardíaco (normas varían según la edad) </td></tr><tr><td> 1. Es la variabilidad de la frecuencia cardíaca baja al inicio del estudio? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 2. Hace disminuir la variabilidad del ritmo cardíaco más allá de la línea de base para cualquier factor estresante? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 3. ¿Existe la recuperación de la variabilidad del ritmo cardíaco durante los periodos de descanso? </td><td> Sí _ </td><td> no _ </td></tr></tbody></table> Apéndice I El estrés interpretación de evaluación: <table border="0"><tbody><tr><td colspan="3"> EMG (lectura normal &lt;3 microvoltios) </td></tr><tr><td> 1. EMG es elevado al inicio del estudio? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 2. EMG es elevada más allá de la línea de base en cualquiera de los tres factores de estrés? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 3. ¿Existe la recuperación del GGA a nivel de línea de base durante los periodos de descanso? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> La frecuencia cardiaca (normal 60 a 80 latidos por minuto) </td></tr><tr><td> 1. Es la frecuencia cardíaca elevada al inicio del estudio? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 2. Es elevado ritmo cardíaco más allá de la línea de base en cualquiera de los tres factores de estrés? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 3. ¿Existe la recuperación de la frecuencia cardiaca a nivel de línea de base durante los periodos de descanso? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> La frecuencia respiratoria (tarifa normal &lt;12 respiraciones por minuto) </td></tr><tr><td> 1. Es la frecuencia respiratoria elevada al inicio del estudio? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 2. Tasa de respiración se eleva más allá de la línea de base para cualquier factor estresante? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 3. ¿Existe la recuperación de la frecuencia respiratoria de base durante los periodos de descanso? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> Conductancia de la piel (normal de lectura &lt;5 ohmios micro) </td></tr><tr><td> 1. Conductancia de la piel es elevada al inicio del estudio? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 2. Conductancia de la piel se eleva más allá de la línea de base para cualquier factor estresante? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 3. ¿Existe la recuperación de la conductancia de la piel a la línea de base durante los periodos de descanso? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> Los dedos de temperatura (lectura normal&gt; 90 ° F) </td></tr><tr><td> 1. Es la temperatura del dedo bajo al inicio del estudio? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 2. Se reduce la temperatura dedo más allá de la línea de base para cualquier factor estresante? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 3. ¿Existe la recuperación de la temperatura de los dedos a la línea base durante periodos de descanso? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> Variabilidad del ritmo cardíaco (normas varían según la edad) </td></tr><tr><td> 1. Es la variabilidad de la frecuencia cardíaca baja al inicio del estudio? </td><td> Sí _ </td><td> no _ </td></tr><tr><td> 2. Hace disminuir la variabilidad del ritmo cardíaco más allá de la línea de base para cualquier factor estresante? </td><td> Sí _ </td><td> no _ </td></tr><tr> &lt;td&gt; 3. ¿Existe la recuperación de la variabilidad del ritmo cardíaco durante los periodos de descanso? </td><td> Sí _ </td><td> no _ </td></tr></tbody></table> La planificación del tratamiento: Si la lectura es más alta o más baja de lo normal al inicio del estudio, el objetivo del tratamiento es ayudar a los clientes llevar ese parámetro fisiológico a la normalidad en todas las circunstancias Si la lectura es más alta o más baja de lo normal durante el estrés, el objetivo del tratamiento es ayudar al cliente a identificar los factores de estrés y ser capaz de mantener el nivel normal de funcionamiento fisiológico de estrés Si la lectura indica la falta de recuperación durante el tiempo de inactividad, el objetivo del tratamiento es ayudar a los clientes llevar ese parámetro fisiológico a un nivel normal rápidamente después de un estresante.

<ol>
	<li>Arena, J., Schwartz, M. S., Schwartz, M. S., Andrasik, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Psychophysiological+assessment+and+biofeedback+baselines.">Psychophysiological assessment and biofeedback baselines.</a> A Practitioners Guide. , 128-158 (2005).</li><li>Schwartz, N. M., Schwartz, M. S., Schwartz, M. S., Andrasik, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Definitons+of+biofeedback+and+applied+psychophysiology.">Definitons of biofeedback and applied psychophysiology.</a> A Practitioners Guide. , 27-42 (2005).</li></ol>

The authors have nothing to disclose.

Psicofisiológicos completado en la evaluación de estrés Apéndices de un profesional de la biorretroalimentación proporciona la información necesaria para la formulación de un plan de tratamiento. Esta evaluación permite obtener información tanto estereotipia la respuesta del individuo, o la uniformidad de ciertas respuestas fisiológicas a todos los factores de estrés, y la especificidad de estímulo-respuesta, o únicas respuestas fisiológicas del individuo a diferentes tipos de estrés. Con esta informació...

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psychophysiological stress assessment using biofeedback 

En el último medio siglo, la investigación en biofeedback ha demostrado la medida en que la mente humana puede influir en el funcionamiento del sistema nervioso autónomo, que anteriormente se consideraban fuera del control consciente. Al permitir que las personas observan las señales de su propio cuerpo, el biofeedback les permite desarrollar una mayor conciencia de sus reacciones fisiológicas y psicológicas, tales como el estrés, y para aprender a modificar estas reacciones. Los profesionales de la biorretroalimentación puede facilitar este proceso mediante la evaluación de las reacciones de la gente s a los eventos estresantes y ligeramente la formulación de un plan de tratamiento basado en biofeedback. Durante la evaluación de la tensión el primer practicante registra una línea de base para las lecturas fisiológicas, y luego presenta al cliente con varios factores de estrés leve, como un factor de estrés cognitivo, físico y emocional. Variedad de factores de estrés se presenta con el fin de determinar de una persona de estímulo-respuesta especificidad, o las diferencias en la reacción de cada persona a los estímulos cualitativamente diferente. Este vídeo demuestra el proceso de evaluación de estrés psicofisiológico con las directrices generales de biofeedback y el presente para planificar el tratamiento.

Watch this Scientific Journal Video about Psychophysiological Stress Assessment Using Biofeedback at JoVE.com

Psychophysiological Stress Assessment Using Biofeedback

En este video se describe un método para evaluar la reacción psicofisiológica persona al estrés utilizando biofeedback. También presentará las directrices generales para la planificación del tratamiento.

Evaluación psicofisiológica de estrés uso de Biofeedback

In the last half century, research in biofeedback has shown the extent to which the human mind can influence the functioning of the autonomic nervous ...

psychophysiological-stress-assessment-using-biofeedback

Research

JoVE Journal

Biology

Psychophysiological Stress Assessment Using Biofeedback 

13.2K vistas.  Cambridge Health Alliance. En este video se describe un método para evaluar la reacción psicofisiológica persona al estrés utilizando biofeedback. También presentará las directrices generales para la planificación del tratamiento.

Video: Psychophysiological Stress Assessment Using Biofeedback

Monitoring Plant Hormones During Stress Responses

Plant hormones and related signaling compounds play an important role in the regulation of plant responses to various environmental stimuli and stresses. Among the most severe stresses are insect herbivory, pathogen infection, and drought stress. For each of these stresses a specific set of hormones and/or combinations thereof are known to fine-tune the responses, thereby ensuring the plant's survival. The major hormones involved in the regulation of these responses are jasmonic acid (JA), salicylic acid (SA), and abscisic acid (ABA). To better understand the role of individual hormones as well as their potential interaction during these responses it is necessary to monitor changes in their abundance in a temporal as well as in a spatial fashion. For the easy, sensitive, and reproducible quantification of these and other signaling compounds we developed a method based on vapor phase extraction and gas chromatography/mass spectrometry (GC/MS) analysis (1, 2, 3, 4). After extracting these compounds from the plant tissue by acidic aqueous 1-propanol mixed with dichloromethane the carboxylic acid-containing compounds are methylated, volatilized under heat, and collected on a polymeric absorbent. After elution into a sample vial the analytes are separated by gas chromatography and detected by chemical ionization mass spectrometry. The use of appropriate internal standards then allows for the simple quantification by relating the peak areas of analyte and internal standard.

A simple method is provided that allows for the rapid extraction and analysis of multiple plant hormones from small tissue samples. The procedure uses vapor phase extraction as the solemn purification step. Samples are analyzed by GC/MS with chemical ionization that produces mainly (M+1)+ ions.

Monitoreo hormonas vegetales en las respuestas de estrés

Plant hormones and related signaling compounds play an important role in the regulation of plant responses to various environmental stimuli and stresses. ...

Investigación

Biología

Microsurgical Clip Obliteration of Middle Cerebral Aneurysm Using Intraoperative Flow Assessment

Cerebral aneurysms are abnormal widening or ballooning of a localized segment of an intracranial blood vessel.  Surgical clipping is an important treatment for aneurysms which attempts to exclude blood from flowing into the aneurysmal segment of the vessel while preserving blood flow in a normal fashion.  Improper clip placement may result in residual aneurysm with the potential for subsequent aneurysm rupture or partial or full occlusion of distal arteries resulting in cerebral infarction.  Here we describe the use of an ultrasonic flow probe to provide quantitative evaluation of arterial flow before and after microsurgical clip placement at the base of a middle cerebral artery aneurysm.  This information helps ensure adequate aneurysm reconstruction with preservation of normal distal blood flow.

Description of the surgical obliteration of a cerebral aneurysm utilizing an ultrasonic flow probe to assess arterial flow prior to and after aneurysm clip placement.

La obliteración de microcirugía Clip de Aneurisma Cerebral Media uso de la evaluación intraoperatoria de flujo

Cerebral aneurysms are abnormal widening or ballooning of a localized segment of an intracranial blood vessel.  Surgical clipping is an important ...

A Quantitative Assessment of The Yeast Lipidome using Electrospray Ionization Mass Spectrometry

Lipids are one of the major classes of biomolecules and play important roles membrane dynamics, energy storage, and signalling1-4. The budding yeast Saccharomyces cerevisiae, a genetically and biochemically manipulable unicellular eukaryote with annotated genome and very simple lipidome, is a valuable model for studying biological functions of various lipid species in multicellular eukaryotes2,3,5. S. cerevisiae has 10 major classes of lipids with chain lengths mainly of 16 or 18 carbon atoms and either zero or one degree of unsaturation6,7. Existing methods for lipid identification and quantification - such as high performance liquid chromatography, thin-layer chromatography, fluorescence microscopy, and gas chromatography followed by MS - are well established but have low sensitivity, insufficiently separate various molecular forms of lipids, require lipid derivitization prior to analysis, or can be quite time consuming. Here we present a detailed description of our experimental approach to solve these inherent limitations by using survey-scan ESI/MS for the identification and quantification of the entire complement of lipids in yeast cells. The described method does not require chromatographic separation of complex lipid mixtures recovered from yeast cells, thereby greatly accelerating the process of data acquisition. This method enables lipid identification and quantification at the concentrations as low as g/ml and has been successfully applied to assessing lipidomes of whole yeast cells and their purified organelles. Lipids extraction from whole yeast cells for using this method of lipid analysis takes two to three hours. It takes only five to ten minutes to run each sample of extracted and dried lipids on a Q-TOF mass spectrometer equipped with a nano-electrospray source.

We describe a new quantitative lipidomics method for identifying numerous lipid species in yeast using survey-scan electrospray ionization mass spectrometry (ESI/MS). This method exceeds currently available methods for lipid identification and quantification in the ability to resolve various molecular forms of lipids, sensitivity, and speed.

Una evaluación cuantitativa de la Lipidome levadura utilizando la espectrometría de masas de ionización por electrospray

Lipids are one of the major classes of biomolecules and play important roles membrane dynamics, energy storage, and signalling1-4. The budding yeast ...

The use of Biofeedback in Clinical Virtual Reality: The INTREPID Project

Generalized anxiety disorder (GAD) is a psychiatric disorder characterized by a constant and unspecific anxiety that interferes with daily-life activities. Its high prevalence in general population and the severe limitations it causes, point out the necessity to find new efficient strategies to treat it. Together with the cognitive-behavioral treatments, relaxation represents a useful approach for the treatment of GAD, but it has the limitation that it is hard to be learned. The INTREPID project is aimed to implement a new instrument to treat anxiety-related disorders and to test its clinical efficacy in reducing anxiety-related symptoms. The innovation of this approach is the combination of virtual reality and biofeedback, so that the first one is directly modified by the output of the second one. In this way, the patient is made aware of his or her reactions through the modification of some features of the VR environment in real time. Using mental exercises the patient learns to control these physiological parameters and using the feedback provided by the virtual environment is able to gauge his or her success. The supplemental use of portable devices, such as PDA or smart-phones, allows the patient to perform at home, individually and autonomously, the same exercises experienced in therapist's office. The goal is to anchor the learned protocol in a real life context, so enhancing the patients' ability to deal with their symptoms. The expected result is a better and faster learning of relaxation techniques, and thus an increased effectiveness of the treatment if compared with traditional clinical protocols.

The Project on Virtual Reality Intelligent Multi-sensor System for the Treatment of Anxiety-related Disorders (INTREPID) is aimed at developing a multi-sensor context-aware virtual reality system for the treatment of anxiety-related disorders.

El uso de la biorretroalimentación en Realidad Virtual Clínica: El Proyecto de INTREPID

Generalized anxiety disorder (GAD) is a psychiatric disorder characterized by a constant and unspecific anxiety that interferes with daily-life ...

Assessment of GFP Expression and Viability Using the Tali Image-Based Cytometer

Single-cell and population information are commonly obtained either by flow cytometry or fluorescence microscopy. However, these two methods provide different information. Flow cytometry gives quantitative multi-parametric information about physical characteristics and staining or expression, but doesn't allow for visualization. Stand-alone fluorescence microscopy provides visual data, but doesn't allow for straightforward quantitative measurements1.
Image-based cytometry bridges the gap between these two methods, enabling the quick visualization and simultaneous quantitative analysis of thousands of cells in heterogeneous populations2. Here, we present a method for performing cell viability and green fluorescent protein (GFP) expression assays using the Tali Image-Based Cytometer3. The Tali instrument is a 3-channel (bright field, green fluorescence, red fluorescence) benchtop assay platform that offers several advantages over flow cytometry and fluorescence microscopy. The Tali cytometer is less expensive, takes up less bench space, requires less maintenance, and the work flow has been simplified so that the operation and analysis is much simpler and quicker. The Tali cytometer is capable of performing a range of suspension cell-based assays, including GFP and red fluorescent protein (RFP) expression, apoptosis4-6 and cell viability analysis with propidium iodide (PI)7-11.
Here, we demonstrate the use of the Tali instrument in performing a cell viability assay in cells expressing GFP. GFP-transduced cells are stained using the Tali Viability Kit - Dead Cell Red. The cells are then pipetted into a Tali Cellular Analysis Slide and loaded into the cytometer. Bright field, red fluorescence and green fluorescence images are captured and analyzed using assay specific algorithms. Histograms are then generated to display cell size, PI fluorescence intensity, and GFP fluorescence intensity. These parameters can then be thresholded to home in on a specific cell population.
A side-by side comparison of the Tali Image-Based Cytometer and traditional flow cytometry demonstrates that the two methods provide comparable data regarding cell viability and protein expression. However, the Tali instrument provides additional visual information about the cell population that cannot be obtained using a flow cytometer.

This protocol describes how to perform cell viability and fluorescence expression assays using the Tali Image-Based Cytometer.

Evaluación de la expresión de GFP y viabilidad mediante el Tali basada en imágenes citómetro

Single-cell and population information are commonly obtained either by flow cytometry or fluorescence microscopy. However, these two methods provide ...

Analysis of Oxidative Stress in Zebrafish Embryos

High levels of reactive oxygen species (ROS) may cause a change of cellular redox state towards oxidative stress condition. This situation causes oxidation of molecules (lipid, DNA, protein) and leads to cell death. Oxidative stress also impacts the progression of several pathological conditions such as diabetes, retinopathies, neurodegeneration, and cancer. Thus, it is important to define tools to investigate oxidative stress conditions not only at the level of single cells but also in the context of whole organisms. Here, we consider the zebrafish embryo as a useful in vivo system to perform such studies and present a protocol to measure in vivo oxidative stress. Taking advantage of fluorescent ROS probes and zebrafish transgenic fluorescent lines, we develop two different methods to measure oxidative stress in vivo: i) a &#8220;whole embryo ROS-detection method&#8221; for qualitative measurement of oxidative stress and ii) a &#8220;single-cell&#160;ROS detection method&#8221; for quantitative measurements of oxidative stress. Herein, we demonstrate the efficacy of these procedures by increasing oxidative stress in tissues by oxidant agents and physiological or genetic methods. This protocol is amenable for forward genetic screens and it will help address cause-effect relationships of ROS in animal models of oxidative stress-related pathologies such as neurological disorders and cancer.

Here we report a protocol to measure oxidative stress in living zebrafish embryos. This procedure allows reactive oxygen species (ROS) detection in both whole embryo tissues and single-cell populations. This protocol will accomplish both qualitative and quantitative analyses.

Análisis de estrés oxidativo en embriones de pez cebra

High levels of reactive oxygen species (ROS) may cause a change of cellular redox state towards oxidative stress condition. This situation causes ...

Method for the Assessment of Effects of a Range of Wavelengths and Intensities of Red/near-infrared Light Therapy on Oxidative Stress In Vitro

Red/near-infrared light therapy (R/NIR-LT), delivered by laser or light emitting diode (LED), improves functional and morphological outcomes in a range of central nervous system injuries in vivo, possibly by reducing oxidative stress. However, effects of R/NIR-LT on oxidative stress have been shown to vary depending on wavelength or intensity of irradiation. Studies comparing treatment parameters are lacking, due to absence of commercially available devices that deliver multiple wavelengths or intensities, suitable for high through-put in vitro optimization studies. This protocol describes a technique for delivery of light at a range of wavelengths and intensities to optimize therapeutic doses required for a given injury model. We hypothesized that a method of delivering light, in which wavelength and intensity parameters could easily be altered, could facilitate determination of an optimal dose of R/NIR-LT for reducing reactive oxygen species (ROS) in vitro.
Non-coherent Xenon light was filtered through narrow-band interference filters to deliver varying wavelengths (center wavelengths of 440, 550, 670 and 810nm) and fluences (8.5 x 10-3 to 3.8 x 10-1 J/cm2) of light to cultured cells. Light output from the apparatus was calibrated to emit therapeutically relevant, equal quantal doses of light at each wavelength. Reactive species were detected in glutamate stressed cells treated with the light, using DCFH-DA and H2O2 sensitive fluorescent dyes.&#160;
We successfully delivered light at a range of physiologically and therapeutically relevant wavelengths and intensities, to cultured cells exposed to glutamate as a model of CNS injury. While the fluences of R/NIR-LT used in the current study did not exert an effect on ROS generated by the cultured cells, the method of light delivery is applicable to other systems including isolated mitochondria or more physiologically relevant organotypic slice culture models, and could be used to assess effects on a range of outcome measures of oxidative metabolism.

Method for the Assessment of Effects of a Range of Wavelengths and Intensities of Red/near-infrared Light Therapy on Oxidative Stress In Vitro

Non-coherent Xenon light was passed through narrow-band interference and neutral density filters to deliver light of varying wavelength and intensity to cultured cells. This protocol was used to assess the effects of red/near-infrared light therapy on production of reactive species in vitro: no effects were observed using the tested parameters.

Método para la Evaluación de los efectos de un rango de longitudes de onda e intensidades de / Terapia de luz del infrarrojo cercano Red sobre el estrés oxidativo<em&gt; In Vitro</em

Red/near-infrared light therapy (R/NIR-LT), delivered by laser or light emitting diode (LED), improves functional and morphological outcomes in a range of ...

A Protocol for Using Gene Set Enrichment Analysis to Identify the Appropriate Animal Model for Translational Research

Recent studies that compared transcriptomic datasets of human diseases with datasets from mouse models using traditional gene-to-gene comparison techniques resulted in contradictory conclusions regarding the relevance of animal models for translational research. A major reason for the discrepancies between different gene expression analyses is the arbitrary filtering of differentially expressed genes. Furthermore, the comparison of single genes between different species and platforms often is limited by technical variance, leading to misinterpretation of the con/discordance between data from human and animal models. Thus, standardized approaches for systematic data analysis are needed. To overcome subjective gene filtering and ineffective gene-to-gene comparisons, we recently demonstrated that gene set enrichment analysis (GSEA) has the potential to avoid these problems. Therefore, we developed a standardized protocol for the use of GSEA to distinguish between appropriate and inappropriate animal models for translational research. This protocol is not suitable to predict how to design new model systems a-priori, as it requires existing experimental omics data. However, the protocol describes how to interpret existing data in a standardized manner in order to select the most suitable animal model, thus avoiding unnecessary animal experiments and misleading translational studies.

We provide a standardized protocol for the use of gene set enrichment analysis of transcriptomic data to identify an ideal mouse model for translational research. 
This protocol can be used with DNA microarray and RNA sequencing data and can further be extended to other omics data if data are available.

Un protocolo para el uso de gen Set enriquecimiento análisis para identificar el modelo Animal adecuado para la investigación traslacional

Recent studies that compared transcriptomic datasets of human diseases with datasets from mouse models using traditional gene-to-gene comparison ...

Fish Sperm Assessment Using Software and Cooling Devices

For gamete quality evaluation, there are innovative, rapid, and quantitative techniques that can provide useful data for aquaculture. Computerized systems for sperm analysis were developed to measure several parameters and one of the most commonly measured is the sperm motility.
Initially, this computer technology was designed for mammalian species, although it can also be used for fish sperm analysis. Fish have specific features that can affect sperm assessment such as a short motility time after activation and, in some cases, adaptation to lower temperatures. Thus, it is necessary to modify both software and hardware components to make motility analysis more efficient for fish sperm analysis. For mammalian sperm, the heating plate is used to maintain optimal temperatures of spermatozoa. However, for some fish species, it is advantageous to use a lower temperature to prolong the duration of motility, since the sperm remain active for less than 2 min. Therefore, cooling devices are necessary to refrigerate samples at constant temperature over the time of analysis, including on the optical microscope. This protocol describes the analysis of fish sperm motility using software for sperm analysis and new cooling devices to optimize the results.

The present protocol describes a procedure of fish sperm assessment using computer-assisted sperm analysis and cooling devices. The software gives a rapid, accurate and quantitative analysis of fish sperm quality based on spermatozoa motility, which can be a useful tool in aquaculture to improve reproduction success.

Evaluación del esperma de pescado utilizando Software y equipos de refrigeración

For gamete quality evaluation, there are innovative, rapid, and quantitative techniques that can provide useful data for aquaculture. Computerized systems ...

An Air-liquid Interface Bronchial Epithelial Model for Realistic, Repeated Inhalation Exposure to Airborne Particles for Toxicity Testing

For toxicity testing of airborne particles, air-liquid interface (ALI) exposure systems have been developed for in vitro tests in order to mimic realistic exposure conditions. This puts specific demands on the cell culture models. Many cell types are negatively affected by exposure to air (e.g., drying out) and only remain viable for a few days. This limits the exposure conditions that can be used in these models: usually relatively high concentrations are applied as a cloud (i.e., droplets containing particles, which settle down rapidly) within a short period of time. Such experimental conditions do not reflect realistic long-term exposure to low concentrations of particles. To overcome these limitations the use of a human bronchial epithelial cell line, Calu-3 was investigated. These cells can be cultured at ALI conditions for several weeks while retaining a healthy morphology and a stable monolayer with tight junctions. In addition, this bronchial model is suitable for testing the effects of repeated exposures to low, realistic concentrations of airborne particles using an ALI exposure system. This system uses a continuous airflow in contrast to other ALI exposure systems that use a single nebulization producing a cloud. Therefore, the continuous flow system is suitable for repeated and prolonged exposure to airborne particles while continuously monitoring the particle characteristics, exposure concentration, and delivered dose. Taken together, this bronchial model, in combination with the continuous flow exposure system, is able to mimic realistic, repeated inhalation exposure conditions that can be used for toxicity testing.

Described is the cell culture and exposure method of an in vitro bronchial model for realistic, repeated inhalation exposure to particles for toxicity testing.

Un modelo epitelial bronquial de interfaz aire-líquido para la exposición realista y repetida por inhalación a partículas en el aire para pruebas de toxicidad

For toxicity testing of airborne particles, air-liquid interface (ALI) exposure systems have been developed for in vitro tests in order to mimic realistic ...