Name: a battery of motor tests in a neonatal mouse model of cerebral palsy
Uploaded: 2016-11-03T12:30:00.000+00:00
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Description: Watch this Scientific Journal Video about A Battery of Motor Tests in a Neonatal Mouse Model of Cerebral Palsy at JoVE.com

We would like to thank everyone at Shriners Hospital Pediatric Research Center, in particular Dr. Mickey Seltzer, of whom without his support, this work would not have been funded. In addition, we would like to thank Isha Srivastava, who contributed to early data collection and Amy He, who helped with the figures. This study was funded by Shriners Hospitals for Children. No funding source played a role in experimental design or decision to submit the paper for publication.

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The authors declare that they have no competing financial interests.

El uso de modelos animales para estudiar enfermedades humanas sólo es relevante si existe una superposición entre la respuesta celular y molecular entre humanos y roedores, y que las pruebas de comportamiento realizadas tienen relación directa con los síntomas humanos. Uno de los principales problemas con los estudios de enfermedades pediátricas es que muchos investigadores utilizan roedores adultos para crear el modelo, así como la evaluación del comportamiento de roedores adultos, sin tener en cuenta las diferencias de desarrollo que pueden ser importantes para el proceso de la enfermedad. Debido a estos problemas, es importante que la investigación sobre el uso de enfermedad pediátrica no sólo el ajustada desarrollo puntos de tiempo apropiado (por ejemplo, el desarrollo de CNS humano a las 28 - 32 semanas es equivalente a un día post-natal 2-7 días roedor) 7, sino también las pruebas de comportamiento que se examinarán de motor adecuada, los comportamientos del desarrollo sensorial o reflexivos. Por lo tanto, a medida que se desarrolla cada nuevo modelo de enfermedad neonatal, debe ser probado rigurosamente para asegurar que el celulary las respuestas de comportamiento proporcionarán los datos traducibles más adecuadas entre roedores y humanos. La parálisis cerebral es un trastorno motor, que persisten en la edad adulta. Un problema con muchos de los modelos disponibles en la actualidad la parálisis cerebral es la falta de pruebas de motor repetible, estandarizada que puede correlacionarse con los déficits observados en los pacientes pediátricos. En este nuevo modelo, que combina la hipoxia, la isquemia y la inflamación en un ratón neonatal, el comportamiento motor se evaluó usando una batería de pruebas específicas para ratones neonatales. Con el fin de disminuir la subjetividad y aumentar la información cuantitativa, varias pruebas han sido modificados para incluir muy específico, pero fácil de evaluar las medidas que pueden ser estandarizados. Además, las evaluaciones delanteros y los miembros posteriores se pueden realizar por separado, y las diferencias de izquierda / derecha se pueden determinar. Esta batería de pruebas es específica para ratones recién nacidos hasta dos semanas de edad. Este modelo demuestra CPdificultad para caminar (ambulación, ángulo del pie de las extremidades posteriores), así como la debilidad de las extremidades específica (suspensión de cuatro extremidades, suspensión posterior), y los déficit en los reflejos del desarrollo (agarrar reflejo). Aunque en este estudio se examinó sólo un punto de tiempo, estos déficits pueden ser rastreados a través del tiempo. Hay otras series de pruebas que pueden utilizarse en el recién nacido, tales como la batería del Fox de pruebas o evaluación de hitos del desarrollo de 15 Heyser. Sin embargo, estas pruebas comparan el recién nacido al adulto, cuyas respuestas no son necesariamente lo mismo porque el recién nacido aún está en desarrollo. La batería de Fox y pruebas Assement de Heyser se basan en información subjetiva de observación con evaluación dicotómica (sí o no), en lugar de los datos objetivos (ángulo de la postura, sobre la base de la fuerza, etc). Debido a la subjetividad de estas pruebas, muchos científicos se han adaptado, añadido o eliminado criterios, con lo que sus resultados incomparables a los demás y límiteing la utilidad de los datos en términos de establecer un déficit línea de base para una enfermedad o trastorno particular. Mediante el establecimiento de un conjunto de pruebas de motor estandarizados que son cualitativa y específicamente diseñado para probar los recién nacidos, los resultados de los grupos de investigación individuales pueden ser precisa y fiable y se comparan.

El desarrollo de nuevos modelos de lesión o enfermedad pediátrica usando roedores es a menudo difícil debido a la sorprendente capacidad de ratas y ratones a recuperarse rápidamente de la lesión neurológica. Por lo tanto, con el fin de validar cualquier nuevo modelo de enfermedad pediátrica, examinando a fondo los cambios celulares y moleculares deben ir mano a mano con los resultados conductuales. En muchos sentidos, la recuperación funcional de la conducta puede ser más importante que los cambios celulares subyacentes en términos de relevancia terapéutica o de traslación. A medida que los investigadores aprenden más sobre lesiones en el adulto y el recién nacido, es evidente que sus respuestas son muy diferentes y no se pueden extrapolar entre los dos. Por ejemplo, los ratones neonatales muestran diferentes niveles de factor de crecimiento nervioso, factor neurotrófico derivado del cerebro, neurotrofina-3 y el factor neurotrófico derivado de la línea celular glial después de una lesión de la médula espinal 1,2. Además, los recién nacidos tienen significativa permeabilidad de la barrera hematoencefálica después del accidente cerebrovascular 3, demonstrate reordenamiento neurona cortical después de la lesión del nervio periférico 4, y tienen una astrogliosis retrasado o desacelerado después de una lesión de la médula espinal y la hipoxia-isquemia 5,6. Por lo tanto, es importante que la investigación pediátrica de traslación utilizar modelos de desarrollo equivalentes y que esos modelos se evalúan para los cambios moleculares / celulares y pruebas de comportamiento apropiadas para la edad. La parálisis cerebral (PC) es un trastorno motor que afecta a 3: 1000 nacidos vivos al año (NIH). Los niños con CP presentan una serie de síntomas y condiciones comórbidas, dependiendo de la gravedad de la enfermedad. Dificultad con el movimiento y la coordinación son los signos más comunes, junto con los retrasos en alcanzar los hitos del desarrollo motor. Otros signos incluyen tono muscular anormal (ya sea aumentado o disminuido), la reducción de las habilidades motoras finas, dificultad para caminar, tragar y babeo excesivo y retrasos en el habla (NIH). La causa subyacente de CP se cree que esuna falta de oxígeno y / o el flujo de sangre al cerebro durante el período de pre- o periparto, o hasta un año después del parto. Además, la inflamación es ahora cree que es un componente clave en el desarrollo de CP. La mayoría de los casos de CP están asociados con daño de la sustancia blanca alrededor de los ventrículos, conocidas como leucomalacia periventricular (PVL). Este sello neurológica sugiere que la lesión inicial que conduce a CP se produce durante el período de desarrollo del cerebro cuando los oligodendrocitos son más vulnerables a los insultos. El período de crecimiento rápido de los oligodendrocitos en un ser humano, también el período en que los oligodendrocitos son los más susceptibles a las lesiones, es entre 24 - 32 semanas de gestación. En los roedores, el período equivalente es días postnatales 2 - 7 7, y es cuando se induce CP en este modelo. El modelo de ratón neonatal de CP que se utilizó para realizar las pruebas descritos aquí combina la hipoxia e isquemia con la inflamación para crear un injUry que mejor imita la neurodegeneración ven en CP humano. Este modelo aborda algunas de las principales deficiencias observadas en otros modelos animales de CP, que carecen de los déficits distintos de motor que se asemeja a los pacientes con PC humanos, así como distintos daño de la materia blanca. Estudios previos realizados por un colaborador con el mismo modelo han demostrado que la adición de la inflamación aumenta el daño de la sustancia blanca, por lo tanto emular mejor el PVL visto en niños con parálisis cerebral 8. Sobre la base de los datos anteriores, este trabajo presenta una batería de pruebas de motor neonatales con el fin de evaluar los cambios en la conducta motora como las edades de los animales.

<table><tbody><tr><td></td><td></td><td></td><td></td></tr><tr><td>C57BL/6 mice</td><td>Charles River Laboratories</td><td>STRAIN CODE: 027&amp;nbsp;</td><td>C57BL/6NCrl is the exact strain we use</td></tr><tr><td>Anesthesia Dish, PYREX&amp;trade; Crystallizing Dish</td><td>Corning Life Sciences Glass&amp;nbsp;</td><td>3140125</td><td>Capacity: 25.03 oz. (740ml); Dia. x H: 4.92 x 2.55 in. (125 x 65mm). However, any small round glass container will work. A 2 cup capacity pyrex food storage bowl with flat bottom will also work and is much cheaper (Pyrex model number: 6017399).</td></tr><tr><td>Covered lead ring</td><td>Fisher Scientific</td><td>S90139C</td><td>Lead ring for stablizing flasks in a water bath. It is used inside the anesthesia dish.</td></tr><tr><td>Scalpel Blade #11</td><td>World Precision Instrucments, Inc.</td><td>500240</td><td></td></tr><tr><td>Small Vessel Cauterizer</td><td>Fine Science Tools</td><td>18000-00</td><td></td></tr><tr><td>Micro Hook</td><td>Fine Science Tools</td><td>10064-14</td><td></td></tr><tr><td>Vetbond Suture Glue</td><td>3M</td><td>1469SB</td><td>n-butyl cyanoacrylate adhesive</td></tr><tr><td>Lipopolysaccharide</td><td>Sigma Life Science</td><td>L4391</td><td>Lipopolysaccaride from E.coli 0111:B4, gamma irradiated</td></tr><tr><td>12 x 12 inch opaque box</td><td>Acrylic Display Manufacturing: A division of Piasa Plastics</td><td>C4022</td><td>Colored Acrylic 5-Sided Cube, 3/16&quot; Colored Acrylic, 12&quot;W x 12&quot;D x 12&quot;H;&amp;nbsp; http://www.acrylicdisplaymfg.com/html/cubes_19.html</td></tr><tr><td>Camera/camcorder</td><td>JVC</td><td>GC-PX100BUS</td><td>Any camcorder that works well in low light and can be imported and edited. We use the JVC GC-PX100 Full HD Everio Camcorder.</td></tr><tr><td>Covidien Tendersorb&amp;trade; Underpads</td><td>Kendall Healthcare Products Co</td><td>7174</td><td></td></tr><tr><td>WypAll L40</td><td>Kimberly-Clark Professional</td><td>5600</td><td>Any surface with moderate grip will do</td></tr><tr><td>Surface at 45 degree incline</td><td></td><td></td><td>We use a cardboard box.</td></tr><tr><td>Thin wire from a pipe cleaner</td><td>Creatology</td><td>M10314420</td><td>Any pipe cleaner from any craft store will work.</td></tr><tr><td>50mL conical tube</td><td>Falcon</td><td>352070</td><td></td></tr><tr><td>Fiberglass Screen Wire</td><td>New York Wire&amp;nbsp; www.lowes.com</td><td>14436</td><td>Any supplier can be used as long as their screen is 16 x 16 or 18 x 16</td></tr><tr><td>Razor blade</td><td>Fisherbrand</td><td>12-640</td><td>A wooden stick applicator or wooden part of a cotton-tipped swab will also work.</td></tr><tr><td>OPTIX 24-in x 4-ft x 0.22-in Clear Acrylic Sheet to make Clear Acrylic Walkway</td><td>PLASKOLITE INC</td><td>1AG2196A</td><td>Clear acrylic (1/8&quot; thick) with sides and a top to limit exploration. We bought a sheet of acrylic from a local hardware store and had them cut it to size. (2) 2&quot; x 2&quot;; (3) 2&quot; x 18&quot;; (1) 2&quot; x 15.5&quot;; (1) 2&quot; x 3&quot;. Using clear tape, tape all sides together, with the 15.5&quot; piece on top. Tape the 3&quot; piece to the end of the 15.5&quot; piece to create a flap/entryway for the mice. Alternatively, part or all of the walkway can be glued together, and only taping on the top pieces. This design will allow for the walkway to be opened for easy cleaning.</td></tr><tr><td>Protractor</td><td>Westscott</td><td>ACM14371</td><td></td></tr></tbody></table>

a battery of motor tests in a neonatal mouse model of cerebral palsy

As the sheer number of transgenic mice strains grow and rodent models of pediatric disease increase, there is an expanding need for a comprehensive, standardized battery of neonatal mouse motor tests. These tests can validate injury or disease models, determine treatment efficacy and/or assess motor behaviors in new transgenic strains. This paper presents a series of neonatal motor tests to evaluate general motor function, including ambulation, hindlimb foot angle, surface righting, negative geotaxis, front- and hindlimb suspension, grasping reflex, four limb grip strength and cliff aversion. Mice between the ages of post-natal day 2 to 14 can be used. In addition, these tests can be used for a wide range of neurological and neuromuscular pathologies, including cerebral palsy, hypoxic-ischemic encephalopathy, traumatic brain injury, spinal cord injury, neurodegenerative diseases, and neuromuscular disorders. These tests can also be used to determine the effects of pharmacological agents, as well as other types of therapeutic interventions. In this paper, motor deficits were evaluated in a novel neonatal mouse model of cerebral palsy that combines hypoxia, ischemia and inflammation. Forty-eight hours after injury, five tests out of the nine showed significant motor deficits: ambulation, hindlimb angle, hindlimb suspension, four limb grip strength, and grasping reflex. These tests revealed weakness in the hindlimbs, as well as fine motor skills such as grasping, which are similar to the motor deficits seen in human cerebral palsy patients.

Los ratones se pusieron a prueba de P7 (24 hr después de la cirugía) para P13 (1 semana después de la cirugía), utilizando ratones diferentes para cada punto de tiempo de manera que el aprendizaje de un paradigma de pruebas no era una variable de confusión. P8 fue seleccionado como resultados representativos, como los ratones mostraron los mayores déficits en este punto temporal. La transición de gatear a caminar es retardado en ratones recién nacidos CP los pacientes con PC humanos tienen alteraciones de la marcha, que van desde caminar de puntillas a una marcha en tijera. Como este modelo CP pantallas de la marcha y los déficits similares a los humanos, se evaluó la deambulación. Los ratones se obtuvieron de la uniformidad de la marcha y el movimiento de las extremidades-pata recta durante un paseo. A las 48 horas después de la cirugía (PND 8), los ratones tenían menos CP movimiento de las extremidades simétricas y un &quot;rastreo&quot; de la marcha, en comparación con sus homólogos sham (puntuación media deambulación: CP 1.083 ±0,6337, n = 12 vs simulacro 1,639 ± 0,4859, n = 9; p &lt;0,05, Figura 10). Por una semana, tanto CP y ratones sham han hecho la transición a caminar (datos no mostrados). <img alt="Figura 10" src="/files/ftp_upload/53569/53569fig10.jpg" /> Figura 10. CP de ratón Los cachorros no Ambulate, así como Shams. Ratones Sham (barra de color negro) tienen una puntuación media de 1.639 ± 0.4859 (n = 9), es decir, su desarrollo se sitúa entre ambulatoria movimiento de las extremidades asimétricas y rastreo lento. ratones CP (barra gris) reciben una puntuación media de 1.083 ± 0.6337 (n = 12), es decir, su deambulación está menos desarrollada y tienden a tener movimiento de las extremidades asimétrica. Los datos se expresan como media ± SEM; * Es p &lt;0,05. <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/53569/53569fig10large.jpg" target="_blank">Haga clic aquí para ver una versión más grande de esta figura.</a> Hindlimb Pie de ángulo se incrementa en CP Además de la deambulación, se evaluó extremidad posterior del pie de ángulo. Ocho días de edad, los cachorros ratón del impostor caminar con sus patas traseras hacia adelante, en comparación con los ratones HIL, que han extendidos patas traseras al caminar en una línea recta (Figura 2; ángulo medio: CP 77,48 ± 9,848, n = 9, vs simulacro 54.54 ± 8,043, n = 11; p &lt;0,0001, Figura 11). Este aumento del ángulo se correlaciona con la inestabilidad de la marcha, en la que los cachorros necesitan para aumentar el ángulo de sus patas traseras con el fin de estabilizar su marcha y ayudar con el equilibrio y la coordinación. <img alt="Figura 11" src="/files/ftp_upload/53569/53569fig11.jpg" /> Figura 11. CP ratón Los cachorros Splay sus patas traseras al caminar. Ratones CP (barras negras) tienen un ángulo medio entre sus miembros traseros de 77,48 ± 8,043 (n = 11), mientras que los ratones sham (b gris<html> ARS) tienen un ángulo promedio de 54,54 ± 9,848 (n = 9). Los datos se expresan como media ± SEM; **** Es p &lt;0,0001. <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/53569/53569fig11large.jpg" target="_blank">Haga clic aquí para ver una versión más grande de esta figura.</a> CP Los ratones no presentan déficit cuando superficial de enderezamiento Se incluyó la prueba de enderezamiento superficie, ya que algunos pacientes con PC tienen problemas de control del tronco (Heyrman et al., 2013). Además, el sistema vestibular es necesario detectar la necesidad de corregir y hay déficit vestibular en algunos pacientes CP 23. ratones CP no muestran déficits significativos cuando enderezamiento en comparación con controles simulados (datos no mostrados). Los ratones CP realizar la misma como Sham en negativo Prueba geotaxis t &quot;fo: keep-together.within-page =&quot; 1 &quot;&gt; geotaxis negativo se utiliza para probar la coordinación motora en ratones jóvenes cachorros son desafiados por ser el lugar que va cuesta abajo sobre una superficie inclinada retraso o falta de orientar hacia arriba podría indicar déficits.. en coordinación, el equilibrio, o la entrada vestibular. ratones CP no muestran déficits cuando fueron estimulados con geotaxis negativos en comparación con los ratones sham (datos no mostrados). Además, los ratones CP no mostraron una preferencia a girar hacia un lado frente a otro cuando se re-orientación . Frente-Limb suspensión de ensayo es apropiado para ratones más viejos de 10 Días los pacientes con PC han disminución del tono muscular y los déficits en las habilidades motoras finas, tales como agarrar. Para probar la debilidad en este modelo de ratón, se utilizó una prueba de suspensión delantera de las extremidades. Por otra parte, este modelo utiliza la lesión isquémica unilateral y bilateral-dad se pudo determinar el uso de esta prueba de suspensión. Esta pruebaes mejor para los ratones de más de 10 días 15. A los 8 días de edad, dos días después de la lesión, no había diferencias significativas entre CP y ratones sham (datos no mostrados). La fuerza de las extremidades posteriores se reduce en ratones CP los pacientes con PC humanos a menudo necesitan aparatos ortopédicos o dispositivos de soporte para caminar debido a la falta de control del motor y la fuerza. Con el fin de comparar el modelo CP de roedores a los seres humanos, la fuerza del miembro posterior se evaluó mediante la prueba de suspensión posterior. Cuando suspendido desde el lado de un tubo cónico, los ratones CP mostraron debilidad de las extremidades posteriores, como se demuestra por una disminución en colgando puntuación (miembro posterior puntuación colgando: CP 3,468 ± 0,5561, n = 13, vs sham 3,891 ± 0,1329, n = 13; p &lt; 0,05, Figura 12). No se observó ninguna diferencia en el tiempo de suspensión de las extremidades posteriores (datos no mostrados). Por lo tanto, similar a los pacientes CP humanos, ratones CP demuestran hindlIMB (pierna) debilidad. <img alt="Figura 12" src="/files/ftp_upload/53569/53569fig12.jpg" /> Figura 12. Los ratones Sham son ligeramente, pero significativamente más fuerte en sus patas traseras que los ratones CP. En una nota media que cuelga de 3,891 ± 0,1329 (n = 13), los ratones sham (barra de color negro) muestra una separación más extremidad posterior, y por lo tanto una postura más fuerte de las extremidades posteriores, al colgar en el borde de un tubo que los ratones CP (barra gris) con una puntuación media que cuelga de 3.468 ± 0.5561 (n = 13). Los datos se expresan como media ± SEM; * Es p &lt;0,05. <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/53569/53569fig12large.jpg" target="_blank">Haga clic aquí para ver una versión más grande de esta figura.</a> La fuerza de agarre se reduce después de una lesión CP Agarrando con las cuatro patas es importante para un roedor entérminos de escalada y correr sobre superficies irregulares. Grip requiere fuerza sostenida significativa, en lugar de destreza o fuerza lineal, principalmente en los dígitos y las patas 24. Los ratones fueron requeridos para mantener su peso corporal en una pantalla de malla de alambre invertida. Ratones CP no fueron capaces de mantener su agarre y estos ratones cayeron en ángulos significativamente más bajos (cuatro extremidades ángulo medio: CP 75.627 ± 24.48, n = 11, vs sham 96,57 ± 10.836, n = 9; p &lt;0,05, Figura 13). Estos datos muestran que hay un déficit significativo en la fuerza de agarre en ratones CP. <img alt="Figura 13" src="/files/ftp_upload/53569/53569fig13.jpg" /> Figura 13. Los ratones tienen CP Grip Más débil que Shams. Ratones Sham (barra de color negro) puede captar un ángulo invertido promedio de 96.57 ± 10.836 (n = 9). ratones CP (barra gris) sólo pueden llegar a un ángulo invertido de 75.627 ± 24,48 (n = 11). Los datos se expresan como media ± SEM; * Es p0; 0.05. <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/53569/53569fig13large.jpg" target="_blank">Haga clic aquí para ver una versión más grande de esta figura.</a> Los déficits aferramiento reflejas son evidentes en ratones CP Junto con los déficits de motricidad gruesa, motricidad fina también se vean afectados en pacientes con PC 25,26. El reflejo de agarre en el ser humano está presente al nacer y desaparece alrededor de 5 - 6 En meses. Sin embargo, los cambios en el reflejo de agarre, tales como la velocidad exagerada o la fuerza de agarre, la falta de agarrar, o la reaparición del reflejo de agarre después de 6 meses de edad, todos indican daño en el sistema nervioso. Para comparar captar en el modelo de CP, se determinaron los déficit de agarre reflexivos. A las 48 horas después de la lesión, los ratones CP demuestran una disminución de agarrar refleja (patas medias agarrarona las 48 horas: CP 2,429 ± 0,9376, n = 14, vs simulado 3.214 ± 0,8018, n = 14; p &lt;0,05, Figura 14A). Hubo un aumento ligero, pero no significativo en la preferencia de la pata derecha en las patas delanteras (datos no mostrados). Hubo una preferencia pata derecha significativa en las patas traseras (CP 75.0 ± 42.74, n = 14, vs simulacro 17.86 ± 54.09, n = 14; p &lt;0,005, Figura 14B). Una semana después de la lesión, los ratones CP muestran déficits de agarre (patas medias captados en la semana 1: CP 2,75 ± 1,035, n = 8, vs simulacro 3,80 ± 0,6325, n = 10; p &lt;0,05, Figura 14C), sin preferencia pata notable . <img alt="Figura 14" src="/files/ftp_upload/53569/53569fig14.jpg" /> Figura 14. Los ratones CP tienen déficit agarrar, en la patas traseras, contralateral a la región del cerebro lesionado. (A) 48 horas después de la lesión (PND 8), los ratones CP (barra gris) agarrar un palo con, en promedio, un menor número de patas que anima farsals (barra de color negro). ratones (B) CP (barra gris) muestran una preferencia para agarrar con la pata trasera derecha (contralateral a la lesión) en lugar de utilizar la pata trasera izquierda (ipsilateral a la lesión). ratones Sham (barra de color negro) no muestran esta preferencia pata derecha. Preferencia pata derecha se calcula como ([pata derecha - izquierda pata] / [pata derecha + pata izquierda + ambas patas] * 100) (C) Una semana después de la lesión, los ratones CP (barra gris) muestran todavía agarrando déficit en comparación con. Shams (barra de color negro). Los datos se expresan como media ± SEM; * Es p &lt;0,05, ** es p &lt;0,005. <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/53569/53569fig14large.jpg" target="_blank">Haga clic aquí para ver una versión más grande de esta figura.</a> Los ratones CP se alejan del borde del acantilado Durante Aversión La prueba se basa en la aversión al precipicio del miedo inherente de los ratones para convertir awaY desde un acantilado y se dirigen hacia la seguridad. Aunque algunos pacientes con PC tienen dificultades vestibulares, así como problemas de control del motor, los ratones CP no mostraron ningún déficit en esta prueba.

Watch this Scientific Journal Video about A Battery of Motor Tests in a Neonatal Mouse Model of Cerebral Palsy at JoVE.com

A Battery of Motor Tests in a Neonatal Mouse Model of Cerebral Palsy

Presented is a concise battery of mouse neonatal motor tests. Using these tests, neonatal motor deficits can be demonstrated in a variety of neonatal motor disorders. By having a standardized set of tests, results from different studies can be compared, allowing for better and accurate reporting between groups.

Una batería de pruebas de motor en un modelo de ratón neonatal de la parálisis cerebral

As the sheer number of transgenic mice strains grow and rodent models of pediatric disease increase, there is an expanding need for a comprehensive, ...

a-battery-of-motor-tests-in-a-neonatal-mouse-model-of-cerebral-palsy

Universidad San Sebastian

Research

JoVE Journal

Medicine

24.0K vistas.  Lewiz Katz School of Medicine at Temple University. Presented is a concise battery of mouse neonatal motor tests. Using these tests, neonatal motor deficits can be demonstrated in a variety of neonatal motor disorders. By having a standardized set of tests, results from different studies can be compared, allowing for better and accurate reporting between groups.

Video: A Battery of Motor Tests in a Neonatal Mouse Model of Cerebral Palsy

Assessment of Sensorimotor Function in Mouse Models of Parkinson's Disease

Sensitive and reliable behavioral outcome measures are essential to the evaluation of potential therapeutic treatments in preclinical trials for many neurodegenerative diseases. In Parkinson's disease, sensorimotor tests sensitive to varying degrees of nigrostriatal dysfunction are fundamental for testing the efficacy of potential therapeutics. Reliable and quite elegant sensorimotor measures exist for rats, however many of these tests measure sensorimotor asymmetry within the rat and are not entirely suitable for the newer genetic mouse models of PD. We have put together a battery of sensorimotor tests inspired by the sensitive tests in rats and adapted for mice. The test battery highlighted in this study is chosen for a) its sensitivity in a wide variety of mouse models of PD, b) its ease in implementing into a study, and c) its low expense. These tests have proven useful in characterizing novel genetic mouse models of PD as well as in testing potential disease-modifying therapies.

In Parkinson's disease and movement disorders in general, sensitive and reliable behavioral assays are essential for testing novel potential therapeutics. Here, we describe a manageable battery of sensorimotor tests for mice that are sensitive to varying degrees of injury to the nigrostriatal system and useful for preclinical studies.

Evaluación de la función sensoriomotora en modelos de ratón de la enfermedad de Parkinson

Sensitive and  reliable behavioral outcome measures are essential to the evaluation of  potential therapeutic treatments in preclinical trials for many  ...

Investigación

Comportamiento

Paw-Dragging: a Novel, Sensitive Analysis of the Mouse Cylinder Test

The cylinder test is routinely used to predict focal ischemic damage to the forelimb motor cortex in rodents. When placed in the cylinder, rodents explore by rearing and touching the walls of the cylinder with their forelimb paws for postural support. Following ischemic injury to the forelimb sensorimotor cortex, rats rely more heavily on their unaffected forelimb paw for postural support resulting in fewer touches with their affected paw which is termed forelimb asymmetry. In contrast, focal ischemic damage in the mouse brain fails to result in comparable consistent deficits in forelimb asymmetry. While forelimb asymmetry deficits are infrequently observed, mice do demonstrate a novel behaviour post stroke termed &#8220;paw-dragging&#8221;. Paw-dragging is the tendency for a mouse to drag its affected paw along the cylinder wall rather than directly push off from the wall when dismounting from a rear to a four-legged stance. We have previously demonstrated that paw-dragging behaviour is highly sensitive to small cortical ischemic injuries to the forelimb motor cortex. Here we provide a detailed protocol for paw-dragging analysis. We define what a paw-drag is and demonstrate how to quantify paw-dragging behaviour. The cylinder test is a simple and inexpensive test to administer and does not require pre-training or food deprivation strategies. In using paw-dragging analysis with the cylinder test, it fills a niche for predicting cortical ischemic injuries such as photothrombosis and Endothelin-1 (ET-1)-induced ischemia &#8211; two models that are ever-increasing in popularity and produce smaller focal injuries than middle cerebral artery occlusion. Finally, measuring paw-dragging behaviour in the cylinder test will allow studies of functional recovery after cortical injury using a wide cohort of transgenic mouse strains where previous forelimb asymmetry analysis has failed to detect consistent deficits.

Classical forelimb asymmetry analysis of the cylinder test is routinely used to assess behavioural deficits in rats following brain injury or stroke; however, it fails to detect consistent deficits in mice. This study demonstrates that quantifying paw-dragging behaviour is a more sensitive analysis of brain injury in mice.

Paw-Arrastrar: una novela, Análisis sensible de la prueba Cilindro Ratón

The cylinder test is routinely used to predict focal ischemic damage to the forelimb motor cortex in rodents. When placed in the cylinder, rodents explore ...

Measuring Motor Coordination in Mice

Mice are increasingly being used in behavioral neuroscience, largely replacing rats as the behaviorist's animal of choice. Before aspects of behavior such as emotionality or cognition can be assessed, however, it is vital to determine whether the motor capabilities of e.g. a mutant or lesioned mouse allow such an assessment. Performance on a maze task requiring strength and coordination, such as the Morris water maze, might well be impaired in a mouse by motor, rather than cognitive, impairments, so it is essential to selectively dissect the latter from the former. For example, sensorimotor impairments caused by NMDA antagonists have been shown to impair water maze performance2. 
Motor coordination has traditionally been assessed in mice and rats by the rotarod test, in which the animal is placed on a horizontal rod that rotates about its long axis; the animal must walk forwards to remain upright and not fall off. Both set speed and accelerating versions of the rotarod are available.
The other three tests described in this article (horizontal bar, static rods and parallel bars) all measure coordination on static apparatus. The horizontal bar also requires strength for adequate performance, particularly of the forelimbs as the mouse initially grips the bar just with the front paws. 
Adult rats do not perform well on tests such as the static rods and parallel bars (personal observations); they appear less well coordinated than mice. I have only tested male rats, however, and male mice seem generally less well coordinated than females. Mice appear to have a higher strength:weight ratio than rats; the Latin name, Mus musculus, seems entirely appropriate. The rotarod, the variations of the foot fault test12 or the Catwalk (Noldus)15 apparatus are generally used to assess motor coordination in rats.

Protocols are presented for two established motor coordination tasks, the accelerating rotarod and horizontal bar, also two tests developed in Oxford recently, the static rods and parallel bars. These tests can detect motor impairments potentially of interest in their own right, as well as being possible variables in tests of other areas of behavior.

Medición de Coordinación motora en ratones

Mice are increasingly being used in behavioral neuroscience, largely replacing rats as the behaviorist's animal of choice. Before aspects of behavior such ...

Neurociencias

Neurodevelopmental Reflex Testing in Neonatal Rat Pups

Neurodevelopmental reflex testing is commonly used in clinical practice to assess the maturation of the nervous system. Neurodevelopmental reflexes are also referred to as primitive reflexes. They are sensitive and consistent with later outcomes. Abnormal reflexes are described as an absence, persistence, reappearance, or latency of reflexes, which are predictive indices of infants that are at high risk for neurodevelopmental disorders. Animal models of neurodevelopmental disabilities, such as cerebral palsy, often display aberrant developmental reflexes, as would be observed in human infants. The techniques described assess a variety of neurodevelopmental reflexes in neonatal rats. Neurodevelopmental reflex testing offers the investigator a testing method that is not otherwise available in such young animals. The methodology presented here aims to assist investigators in examining developmental milestones in neonatal rats as a method of detecting early-onset brain injury and/or determining the effectiveness of therapeutic interventions. The methodology presented here aims to provide a general guideline for investigators.

Behavioral testing is the gold standard for determining outcomes following brain injury, and can identify the presence of developmental disabilities in infants and children. Neurodevelopmental reflexes are an early indicator of these abnormalities. A host of easily accomplished developmental reflex tests in the neonatal rodent were developed and described here.

Neurodesarrollo pruebas de reflejos neonatales en crías de rata

Neurodevelopmental reflex testing is commonly used in clinical practice to assess the maturation of the nervous system. Neurodevelopmental reflexes are ...

Battery of Behavioral Tests Assessing General Locomotion, Muscular Strength, and Coordination in Mice

Behavioral testing is used in pre-clinical trials to assess the phenotypic effects and outcomes that a particular disease or treatment has on the animal&#39;s wellbeing and health. There are numerous behavioral tests that may be applied. We selected a test for general locomotion, the open field test (OFT); a test for muscular strength, the mesh test (MT); and a test for coordination, the rotarod test (RR). Testing can be accomplished on a weekly or monthly basis. As a test for general locomotion, the OFT works by objectively monitoring movement parameters while the mouse is in an open field apparatus. The field is generally a 2&#39; x 2&#39; box, and the movements are recorded through laser sensing or through video capture. The mouse is placed in the center of the open field and allowed to move freely for the test. The MT uses the latency for a mouse to fall off an inverted screen as a measure of muscular strength. A mouse is placed on a screen, which is inverted over a clear box, and is timed for their latency to fall. Three trials are performed, with the best of the three trials scored for that day. A score of 60 s is the maximum time a mouse is left inverted. Mice are given a 5-min rest period between mesh test trials. Lastly, an accelerated protocol on the RR assesses motor coordination and endurance. During a trial, a mouse walks on a rotating rod as it increases in speed from 4 rpm to 40 rpm over 5 min. The trial ends when the mouse touches the magnetized pressure sensor upon falling. Each mouse undergoes three trials, and the best trial is scored for that day. This combined behavioral data allows for the global assessment of mobility, coordination, strength, and movement of the test animals. At least two out of the three behavioral testing measures must show improvement for an animal to qualify as having overall improved motor function.

Behavioral testing is used in pre-clinical trials to assess the phenotypic effects of a disease or treatment on an animal's wellbeing. In order to globally assess motor functioning, we selected tests for general locomotion, muscular strength, and coordination: the open field test, the mesh test, and the rotarod test, respectively.

Batería de comportamiento pruebas evaluación General locomoción, fuerza Muscular y la coordinación en ratones

Behavioral testing is used in pre-clinical trials to assess the phenotypic effects and outcomes that a particular disease or treatment has on the ...

A Neonatal Mouse Spinal Cord Compression Injury Model

Spinal cord injury (SCI) typically causes devastating neurological deficits, particularly through damage to fibers descending from the brain to the spinal cord. A major current area of research is focused on the mechanisms of adaptive plasticity that underlie spontaneous or induced functional recovery following SCI. Spontaneous functional recovery is reported to be greater early in life, raising interesting questions about how adaptive plasticity changes as the spinal cord develops. To facilitate investigation of this dynamic, we have developed a SCI model in the neonatal mouse. The model has relevance for pediatric SCI, which is too little studied. Because neural plasticity in the adult involves some of the same mechanisms as neural plasticity in early life1, this model may potentially have some relevance also for adult SCI. Here we describe the entire procedure for generating a reproducible spinal cord compression (SCC) injury in the neonatal mouse as early as postnatal (P) day 1. SCC is achieved by performing a laminectomy at a given spinal level (here described at thoracic levels 9-11) and then using a modified Yasargil aneurysm mini-clip to rapidly compress and decompress the spinal cord. As previously described, the injured neonatal mice can be tested for behavioral deficits or sacrificed for ex vivo physiological analysis of synaptic connectivity using electrophysiological and high-throughput optical recording techniques1. Earlier and ongoing studies using behavioral and physiological assessment have demonstrated a dramatic, acute impairment of hindlimb motility followed by a complete functional recovery within 2 weeks, and the first evidence of changes in functional circuitry at the level of identified descending synaptic connections1.

This article describes a method for generating a reproducible spinal cord compression injury (SCI) in the neonatal mouse. The model provides an advantageous platform for studying mechanisms of adaptive plasticity that underlie spontaneous functional recovery.

Un ratón neonatal lesión de médula espinal de compresión Modelo

Spinal cord injury (SCI) typically causes devastating neurological deficits, particularly through damage to fibers descending from the brain to the spinal ...

Medicina

Neurobehavioral Assessments in a Mouse Model of Neonatal Hypoxic-ischemic Brain Injury

We performed unilateral carotid artery occlusion on CD-1 mice to create a neonatal hypoxic-ischemic (HI) model and investigated the effects of neonatal HI brain injury by studying neurobehavioral functions in these mice compared to non-operated (i.e., normal) mice. During the study, Rice-Vannucci&#39;s method was used to induce neonatal HI brain damage in postnatal day 7-10 (P7-10) mice. The HI operation was performed on the pups by unilateral carotid artery ligation and exposure to hypoxia (8% O2 and 92% N2 for 90 min). One week after the operation, the damaged brains were evaluated with the naked eye through the semi-transparent skull and were categorized into subgroups based on the absence (&#34;no cortical injury&#34; group) or presence (&#34;cortical injury&#34; group) of cortical injury, such as a lesion in the right hemisphere. On week 6, the following neurobehavioral tests were performed to evaluate the cognitive and motor functions: passive avoidance task (PAT), ladder walking test, and grip strength test. These behavioral tests are helpful in determining the effects of neonatal HI brain injury and are used in other mouse models of neurodegenerative diseases. In this study, neonatal HI brain injury mice showed motor deficits that corresponded to right hemisphere damage. The behavioral test results are relevant to the deficits observed in human neonatal HI patients, such as cerebral palsy or neonatal stroke patients. In this study, a mouse model of neonatal HI brain injury was established and showed different degrees of motor deficits and cognitive impairment compared to non-operated mice. This work provides basic information on the HI mouse model. MRI images demonstrate the different phenotypes, separated according to the severity of brain damage by motor and cognitive tests.

We performed unilateral carotid artery occlusion on postnatal day 7-10 CD-1 mouse pups to create a neonatal hypoxic-ischemic (HI) model and investigated the effects of HI brain injury. We studied neurobehavioral functions in these mice compared to non-operated normal mice.

Evaluación neuroconductual en un modelo murino de lesión de cerebro hipóxico-isquémica Neonatal

We performed unilateral carotid artery occlusion on CD-1 mice to create a neonatal hypoxic-ischemic (HI) model and investigated the effects of neonatal HI ...

A Controlled Mouse Model for Neonatal Polymicrobial Sepsis

Neonatal sepsis remains a global burden. A preclinical model to screen effective prophylactic or therapeutic interventions is needed. Neonatal mouse polymicrobial sepsis can be induced by injecting cecal slurry intraperitoneally into day of life 7 mice and monitoring them for the following week. Presented here are the detailed steps necessary for the implementation of this neonatal sepsis model. This includes making a homogeneous cecal slurry stock, diluting it to a weight- and litter-adjusted dose, an outline of the monitoring schedule, and a definition of observed health categories used to define humane endpoints. The generation of a homogeneous cecal slurry stock from pooled donors allows for the administration into many litters over time, reducing the variation between donors, and preventing the use of potentially toxic glycerol. The monitoring strategy used allows for the anticipation of survival outcome and the identification of mice that would later progress to death, allowing for an earlier identification of the humane endpoint. Two main behavioral features are used to define the health scores, namely, the ability of the neonatal mice to right themselves when placed on their back and their level of mobility. These criteria could potentially be applied to address humane endpoints in other studies of neonatal disease in mice, as long as a pilot study is performed to confirm accuracy. In conclusion, this approach provides a standardized method to model newborn sepsis in mice, while providing resources to assess animal welfare used to define early humane endpoints for challenged animals.

This protocol provides the necessary steps to establish and evaluate neonatal sepsis in 7-day-old mice.

Un modelo de ratón controlado para la Sepsis Neonatal polimicrobiana

Neonatal sepsis remains a global burden. A preclinical model to screen effective prophylactic or therapeutic interventions is needed. Neonatal mouse ...

Inmunología e Infección

A Mouse Model of Single and Repetitive Mild Traumatic Brain Injury

Mild Traumatic Brain Injury (mTBI) can result in the acute loss of brain function, including a period of confusion, a loss of consciousness (LOC), focal neurological deficits and even amnesia. Athletes participating in contact sports are at high risk of exposure to large number of mTBIs. In terms of the level of injury in a sporting athlete, a mTBI is defined as a mild injury that does not cause gross pathological changes, but does cause short-term neurological deficits that are spontaneously resolved. Despite previous attempts to model mTBI in mice and rats, many have reported gross adverse effects including skull fractures, intracerebral bleeding, axonal injury and neuronal cell death. Herein, we describe our highly reproducible animal model of mTBI that reproduces clinically relevant symptoms. This model uses a custom made pneumatic impactor device to deliver a closed-head trauma. This impact is made under precise velocity and deformation parameters, creating a reliable and reproducible model to examine the mechanisms that contribute to effects of single or repetitive concussive mTBI.

Athletes absorb several hundred mild traumatic brain injuries (mTBI)/concussions every year; however, the consequence of these on the brain is poorly understood. Therefore, an animal model of single and repetitive mTBI that consistently replicates clinically relevant symptoms provides the means to advance the study of mTBI and concussion.

Un Modelo de Ratón de Único y Repetitivo Lesión Cerebral Traumática Leve

Mild Traumatic Brain Injury (mTBI) can result in the acute loss of brain function, including a period of confusion, a loss of consciousness (LOC), focal ...

Cuantificación de la disfunción locomotora en ratones con lesión medular utilizando MouseWalker

Using the MouseWalker to Quantify Locomotor Dysfunction in a Mouse Model of Spinal Cord Injury

The execution of complex and highly coordinated motor programs, such as walking and running, is dependent on the rhythmic activation of spinal and supra-spinal circuits. After a thoracic spinal cord injury, communication with upstream circuits is impaired. This, in turn, leads to a loss of coordination, with limited recovery potential. Hence, to better evaluate the degree of recovery after the administration of drugs or therapies, there is a necessity for new, more detailed, and accurate tools to quantify gait, limb coordination, and other fine aspects of locomotor behavior in animal models of spinal cord injury. Several assays have been developed over the years to quantitatively assess free-walking behavior in rodents; however, they usually lack direct measurements related to stepping gait strategies, footprint patterns, and coordination. To address these shortcomings, an updated version of the MouseWalker, which combines a frustrated total internal reflection (fTIR) walkway with tracking and quantification software, is provided. This open-source system has been adapted to extract several graphical outputs and kinematic parameters, and a set of post-quantification tools can be to analyze the output data provided. This manuscript also demonstrates how this method, allied with already established behavioral tests, quantitatively describes locomotor deficits following spinal cord injury.

Autor destacado: Uso del MouseWalker para cuantificar la disfunción locomotora en un modelo de ratón de lesión de la médula espinal  

An experimental pipeline to quantitatively describe the locomotor pattern of freely walking mice using the MouseWalker (MW) toolbox is provided, ranging from initial video recordings and tracking to post-quantification analysis. A spinal cord contusion injury model in mice is employed to demonstrate the usefulness of the MW system.