El trabajo fue financiado por una beca del Instituto de Medicina Traslacional y Terapéutica, y una beca de formación del Instituto de Medicina e Ingeniería (T32HL007954) de la Universidad de Pennsylvania y la Marlene L. Cohen y Jerome H. Fleisch Académico Grant en la Universidad de Connecticut Health Center (LHS) y NIH NS-029331 (FAW).

1. Preparación del equipo <ol><li> Limpie el marco estereotáxico y la bomba con el 75% de etanol para minimizar la contaminación bacteriana. </li><li> Esterilizar la jeringa y la aguja Hamilton silicio fundido. Nota: Si la esterilización química se utiliza, asegúrese de enjuagarse varias veces con agua estéril antes del uso. </li><li> Limpie la superficie de papel de cera de parafina con un 75% de etanol y dejar secar. </li></ol> 2. Preparación de ratón para la inyección Nota: Tenga ratones entregado a su instalación de animales al menos 7 días antes de la cirugía para permitir que se aclimate al nuevo ambiente y reducir el estrés. <ol><li> Pesar del ratón para la pre-operatoria de referencia. </li><li> Inducir la anestesia con un 30% de oxígeno, 70% de óxido nitroso, y el 4% isoflurano hasta que no responden a pellizcar la cola </li><li> Inyectar al ratón con buprenorfina 0.1mg/kg por vía intraperitoneal para la analgesia postoperatoria </li><li> Afeitarse el cuero cabelludo </li><li> Los ojos con agua estéril capa de vaselina </li><li> Preparar el cuero cabelludo con betadine x 3 paños, y luego permitir que se seque el cuero cabelludo </li><li> Haga una incisión sagital cm la línea media del cuero cabelludo con bisturí quirúrgico estéril Nota: Se hace una incisión generosa permitirá la exposición completa de los hitos del cráneo. </li><li> Afeitado 1 cm de la superficie ventral de la cola a partir del 1 cm de base y preparar con betadine x 3 toallitas </li><li> Coloque el ratón sobre el marco estereotáxico Nota: Es importante asegurarse de ratón está seguro en el marco de la superficie del cráneo es paralela a la base de la estructura, con una excelente exposición de bregma y por lo menos 3 mm a la derecha del bregma. </li></ol> 3. Cirugía de la hemorragia intracerebral Notas: Durante toda la cirugía del ratón se anestesia con un 30% de oxígeno, 70% de óxido nitroso, y el 1-3% de isoflurano, continuamente mantiene a 37 ± 0,5 ° C con una resistencia térmica controlada almohadilla térmica y control de un termómetro rectal. <ol><li> Coloque la aguja estéril 27 g en una jeringa de 1 cc en el marco. </li><li> Ajuste el brazo hasta que la aguja estereotáxica es exactamente más de bregma. </li><li> Ajuste el brazo para que la aguja está en 2,5 mm lateral a bregma e inferior a la superficie del cráneo. </li><li> Gire manualmente la jeringa para trepanación del cráneo en la superficie mientras se aplica el movimiento suave hacia abajo de la atención marco de la toma de no perforar el cráneo por completo. </li><li> Se retira la aguja y el agujero de trépano completo manual con jeringa / aguja Nota: Completar el agujero de trépano a mano permite el reconocimiento inmediato cuando haya perforado la tabla interna del cráneo y minimiza el riesgo de empujar inadvertidamente la aguja en el parénquima cerebral. </li><li> Realice una incisión transversal en la superficie ventral de la cola con hoja de bisturí estéril y dejar 2-3 gotas grandes de sangre arterial a caer en el papel de cera de parafina. Entonces, rápidamente, detener el sangrado con presión con una gasa estéril. </li><li> Retirar 17 l de sangre en la jeringa y una jeringa Hamilton lugar en la bomba. </li><li> Ajuste el brazo estereotáxica a punto 5 ° medialmente con relación al eje vertical. </li><li> Ajuste cuidadosamente el brazo estereotáxica manera que la punta de la aguja sobre el agujero de trépano en el cráneo y luego baje la aguja 3,5 mm. </li><li> Espere 2 minutos y luego retirar la aguja de 0,5 mm (de modo que la punta es de 3 mm de profundidad) </li><li> Espere 5 minutos para permitir que el cerebro se expanda de nuevo alrededor de la aguja y minimizar el riesgo de reflujo de sangre por el camino de inserción de la aguja durante la inyección. </li><li> Inyectar sangre 1 l / min para 7,5 mL. </li><li> Esperar 5 minutos para permitir que la sangre inicial de coagulación y para los turnos de los tejidos que se produzcan para minimizar las elevaciones de la presión intracraneal. </li><li> Inyectar el restante 7,5 l de 1 l / minuto </li><li> Permita que la aguja permanezca en su lugar durante 25 minutos para permitir la coagulación de la sangre Nota: Si no espera por coagulación de la sangre se traducirá en la sangre de reflujo hasta el sitio de inserción de la aguja al retirar la aguja </li><li> Lentamente retire la aguja e inmediatamente enjuague con agua caliente para evitar que restos de sangre en la aguja de la coagulación y de garantizar la reutilización de agujas. </li><li> Quitar el ratón de la estructura y cerca de la cola y las incisiones del cuero cabelludo con pegamento quirúrgico veterinario. </li><li> Apague la anestesia. </li><li> Dejar el ratón para despertar mientras se calienta continuamente con libre acceso al alimento humedecido. </li><li> Volver a la jaula del ratón con la camada cuando esté completamente despierto. Lugar húmedo comida de pellets en la parte inferior de la jaula para ayudar a los animales en el acceso a los alimentos. </li></ol> 4. Los resultados representativos: <img src="/files/ftp_upload/2618/2618fig1.jpg" alt="Figura 1"/> Figura 1. Sección coronal de cerebro de ratón 15 minutos después de la cirugía de la ICH. Inmediatamente después del sacrificio del cerebro fue examinado por el éxito ICH basado en la inspección macroscópica de un corte coronal en el sitio de inserción de la aguja. Hemorragias que dio seguimiento a la base del cerebro, el trayecto de la aguja más allá del cuerpo calloso, o en los ventrículos se considerará fracasada y que el ratón fue eliminado de todos los análisis. En general succe ICHlas tasas de las SS fueron 75-85% en 50 ratones con 0% de mortalidad. <img src="/files/ftp_upload/2618/2618fig2.jpg" alt="Figura 2"/> Prueba de la Figura 2. Cilindro demuestra hemiparesia izquierda después de ICH derecho de los ganglios basales. (A) muestra parte trasera del ratón después de la cirugía ICH. Tenga en cuenta la colocación de la extremidad anterior derecha sólo en la pared del cilindro después de ICH izquierda los ganglios basales. (B) Gráfico de las pruebas de un cilindro de resultados de la cohorte de ratones después de la cirugía de la ICH (n = 5) en comparación con el tratamiento simulado (n = 4). Ratones Sham había todos los procedimientos, excepto la inyección de sangre (aguja se inserta en el cerebro). Cada ratón fue colocado en un cilindro de vidrio de 12 cm de diámetro interior y observaron durante 20 traseros. La colocación inicial de los miembros anteriores de la pared del cilindro fue anotado por detrás. Los movimientos posteriores (como la exploración lateral) no se obtuvo hasta que el ratón volvió a la tierra y anotó la parte trasera que viene. El índice se calcula como la lateralidad (# ubicaciones adecuadas extremidad anterior en el lado del cilindro - # izquierda colocaciones extremidad anterior) / (# + # derecha izquierda + # ambos), donde 0 indica ninguna preferencia patas delanteras y 1 indica que sólo la extremidad anterior derecha se utilizó .

<ol>
	<li>Hua, Y., Schallert, T., Keep, R. F., Wu, J., Hoff, J. T., Xi, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavioral+Tests+After+Intracerebral+Hemorrhage+in+the+Rat.">Behavioral Tests After Intracerebral Hemorrhage in the Rat.</a> Stroke. 33, 2478-2484 (2002).</li><li>James, M., Warner, D., Laskowitz, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preclinical+Models+of+Intracerebral+Hemorrhage:+A+Translational+Perspective.">Preclinical Models of Intracerebral Hemorrhage: A Translational Perspective.</a> Neurocritical Care. 9, 139-152 (2008).</li><li>Foerch, C., Arai, K., Jin, G., Park, K. -. P., Pallast, S., van Leyen, K., Lo, E. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+Model+of+Warfarin-Associated+Intracerebral+Hemorrhage.">Experimental Model of Warfarin-Associated Intracerebral Hemorrhage.</a> Stroke. 39, 3397-3404 (2008).</li><li>Nakamura, T., Xi, G., Hua, Y., Schallert, T., Hoff, J. T., Keep, R. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intracerebral+hemorrhage+in+mice:+model+characterization+and+application+for+genetically+modified+mice.">Intracerebral hemorrhage in mice: model characterization and application for genetically modified mice.</a> Journal of Cerebral Blood Flow and Metabolism. 24, 487-494 (2004).</li><li>Rynkowski, M. A., Kim, G. H., Komotar, R. J., Otten, M. L., Ducruet, A. F., Zacharia, B. E., Kellner, C. P., Hahn, D. K., Merkow, M. B., Garrett, M. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+mouse+model+of+intracerebral+hemorrhage+using+autologous+blood+infusion.">A mouse model of intracerebral hemorrhage using autologous blood infusion.</a> Nat. Protocols. 3, 122-128 (2008).</li><li>Wang, J., Fields, J., Dor&#233;, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+development+of+an+improved+preclinical+mouse+model+of+intracerebral+hemorrhage+using+double+infusion+of+autologous+whole+blood.">The development of an improved preclinical mouse model of intracerebral hemorrhage using double infusion of autologous whole blood.</a> Brain Research. 1222, 214-221 (2008).</li><li>Zhao, X., Sun, G., Zhang, J., Strong, R., Song, W., Gonzales, N., Grotta, J. C., Aronowski, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hematoma+resolution+as+a+target+for+intracerebral+hemorrhage+treatment:+Role+for+peroxisome+proliferator-activated+receptor+&#947;+in+microglia/macrophages.">Hematoma resolution as a target for intracerebral hemorrhage treatment: Role for peroxisome proliferator-activated receptor &#947; in microglia/macrophages.</a> Annals of Neurology. 61, 352-362 (2007).</li></ol>

No hay conflictos de interés declarado.

Este modelo quirúrgico de la hemorragia intracerebral en ratones utilizando autólogo cola resultados sanguíneo de la arteria en un modelo reproducible de la hemorragia espontánea ganglios basales. Un modelo en ratones ICH ofrece la ventaja de la disponibilidad de animales transgénicos para investigar la fisiopatología, sin embargo, su pequeño tamaño hace que los procedimientos neuroquirúrgicos técnicamente más difícil que en los animales más grandes. El modelo de la colagenasa y...

Marco estereotáxico para neurocirugía del ratón (Stoelting, 51.925) Microinfusión bomba y el procesador (UMP-3 y Micro4, Instrumentos del Mundo de precisión, Sarasota, FL) Ratón más caliente (Stoelting, 50300) La anestesia por inhalación del ratón (Braintree Scientific, EZ-AF9000) 25 l herméticas borosilicato Hamilton jeringa con el émbolo y la aguja no recubierto (Compañía de Hamilton, Reno, NV, una jeringa 1702RN: 765401, férula: 30949, separador: 30946) aguja de sílice fundida cortar a la longitud de 2 cm (Hamilton, 17739) * Nota Hamilton jeringa y la aguja de sílice fundida se pueden reutilizar para múltiples cirugías si esterilizar antes de cada cirugía. Estos materiales son cruciales para evitar la coagulación de la sangre. Guantes quirúrgicos estériles Bata, gorro y mascarilla Betadine 75% de etanol 27 g de agujas estériles (de un solo uso) estéril de 1 cc jeringa (de un solo uso) hoja de bisturí estéril Cidex agua estéril buprenorfina y el isoflurano gasa estéril cuadrados de papel de cera de parafina Pegamento quirúrgico veterinario (Vetbond, 3M, St. Paul, MN)

autologous blood injection to model spontaneous intracerebral hemorrhage in mice

Investigation of the pathophysiology of injury after intracerebral
hemorrhage (ICH) requires a reproducible animal model. While ICH
accounts for 10-15% of all strokes, there remains no specific
effective therapy. The autologous blood injection model in mice
involves the stereotaxic injection of arterial blood into the basal
ganglia mimicking a spontaneous hypertensive hemorrhage in man. The
response to hemorrhage can then be studied in vivo and the
neurobehavioral deficits quantified, allowing for description of the
ensuing pathology and the testing of potential therapeutic agents. The
procedure described in this protocol uses the double injection
technique to minimize risk of blood reflux up the needle track, no
anticoagulants in the pumping system, and eliminates all dead space
and expandable tubing in the system.

Watch this Scientific Journal Video about Autologous Blood Injection to Model Spontaneous Intracerebral Hemorrhage in Mice at JoVE.com

Autologous Blood Injection to Model Spontaneous Intracerebral Hemorrhage in Mice

El modelo de inyección de sangre autóloga de la hemorragia intracerebral en ratones descritos en este protocolo se utiliza la técnica de doble inyección para minimizar el riesgo de reflujo de sangre hasta el trayecto de la aguja, no los anticoagulantes en el sistema de bombeo, y elimina todo el espacio muerto y el tubo extensible en el sistema.

Inyección de sangre autóloga al modelo hemorragia intracerebral espontánea en ratones

Investigation of the pathophysiology of injury after intracerebral
hemorrhage (ICH) requires a reproducible animal model.  While ICH
accounts for 10-15% ...

autologous-blood-injection-to-model-spontaneous-intracerebral

Research

JoVE Journal

Neuroscience

14.6K vistas.  University of Connecticut Health Center. El modelo de inyección de sangre autóloga de la hemorragia intracerebral en ratones descritos en este protocolo se utiliza la técnica de doble inyección para minimizar el riesgo de reflujo de sangre hasta el trayecto de la aguja, no los anticoagulantes en el sistema de bombeo, y elimina todo el espacio muerto y el tubo extensible en el sistema.

Video: Autologous Blood Injection to Model Spontaneous Intracerebral Hemorrhage in Mice

Methods to Characterize Spontaneous and Startle-induced Locomotion in a Rotenone-induced Parkinson's Disease Model of Drosophila

Parkinson&#8217;s disease is a neurodegenerative disorder that results from the degeneration of dopaminergic neurons in the central nervous system, primarily in the substantia nigra. The disease causes motor deficiencies, which present as rigidity, tremors and dementia in humans. Rotenone is an insecticide that causes oxidative damage by inhibiting the function of the electron transport chain in mitochondria. It is also used to model Parkinson&#8217;s disease in the Drosophila. Flies have an inherent negative geotactic response, which compels them to climb upwards upon being startled. It has been established that rotenone causes early mortality and locomotion defects that disrupt the flies&#8217; ability to climb after they have been tapped downwards. However, the effect of rotenone on spontaneous movement is not well documented. This study outlines two sensitive, reproducible, and high throughput assays to characterize rotenone-induced deficiencies in short-term startle-induced locomotion and long-term spontaneous locomotion in Drosophila. These assays can be conveniently adapted to characterize other Drosophila models of locomotion defects and efficacy of therapeutic agents.

Methods to Characterize Spontaneous and Startle-induced Locomotion in a Rotenone-induced Parkinson&#039;s Disease Model of Drosophila

Parkinson&#8217;s disease is a neurodegenerative disorder that results from the degeneration of dopaminergic neurons in the central nervous system, causing locomotion defects. Rotenone models Parkinson&#8217;s disease in Drosophila. This paper outlines two assays that characterize both spontaneous and startle-induced locomotion deficiencies caused by rotenone.

Métodos para caracterizar espontánea y de sobresalto-inducida Locomotion en un modelo de enfermedad de Parkinson inducida por rotenona-de<em&gt; Drosophila</em

Parkinson&#8217;s disease is a neurodegenerative disorder that results from the degeneration of dopaminergic neurons in the central nervous system, ...

Investigación

Neurociencias

Establishing Mouse Models for Zika Virus-induced Neurological Disorders Using Intracerebral Injection Strategies: Embryonic, Neonatal, and Adult

The Zika virus (ZIKV) is a flavivirus currently endemic in North, Central, and South America. It is now established that the ZIKV can cause microcephaly and additional brain abnormalities. However, the mechanism underlying the pathogenesis of ZIKV in the developing brain remains unclear. Intracerebral surgical methods are frequently used in neuroscience research to address questions about both normal and abnormal brain development and brain function. This protocol utilizes classical surgical techniques and describes methods that allow one to model ZIKV-associated human neurological disease in the mouse nervous system. While direct brain inoculation does not model the normal mode of virus transmission, the method allows investigators to ask targeted questions concerning the consequence after ZIKV infection of the developing brain. This protocol describes embryonic, neonatal, and adult stages of intraventricular inoculation of ZIKV. Once mastered, this method can become a straightforward and reproducible technique that only takes a few hours to perform.

Here we describe a method for establishing a model of Zika virus-induced microcephaly in mouse. This protocol includes methods for embryonic, neonatal, and adult-stage intracerebral inoculation of the Zika virus.

Establecimiento de modelos de ratón para trastornos neurológicos inducidos por el Virus Zika utilizando estrategias de inyección Intracerebral: embrionario, Neonatal y adulto

The Zika virus (ZIKV) is a flavivirus currently endemic in North, Central, and South America. It is now established that the ZIKV can cause microcephaly ...

Rat Model of Widespread Cerebral Cortical Demyelination Induced by an Intracerebral Injection of Pro-Inflammatory Cytokines

Multiple sclerosis (MS) is the most common immune-mediated disease of the central nervous system (CNS) and progressively leads to physical disability and death, caused by white matter lesions in the spinal cord and cerebellum, as well as by demyelination in grey matter. Whilst conventional models of experimental allergic encephalomyelitis are suitable for the investigation of the cell-mediated inflammation in the spinal and cerebellar white matter, they fail to address grey matter pathologies. Here, we present the experimental protocol for a novel rat model of cortical demyelination allowing the investigation of the pathological and molecular mechanisms leading to cortical lesions. The demyelination is induced by an immunization with low-dose myelin oligodendrocyte glycoprotein (MOG) in an incomplete Freund&#39;s adjuvant followed by a catheter-mediated intracerebral delivery of pro-inflammatory cytokines. The catheter, moreover, enables multiple rounds of demyelination without causing injection-induced trauma, as well as the intracerebral delivery of potential therapeutic drugs undergoing a preclinical investigation. The method is also ethically favorable as animal pain and distress or disability are controlled and relatively minimal. The expected timeframe for the implementation of the entire protocol is around 8 - 10 weeks.

The protocol presented here allows the reproduction of a widespread grey matter demyelination of both cortical hemispheres in adult male Dark Agouti rats. The method comprises of intracerebral implantation of a catheter, subclinical immunization against myelin oligodendrocyte glycoprotein, and intracerebral injection of a pro-inflammatory cytokine mixture through the implanted catheter.

Modelo de rata de desmielinización cortical cerebral generalizada inducida por una inyección intracerebral de citoquinas proinflamatorias

Multiple sclerosis (MS) is the most common immune-mediated disease of the central nervous system (CNS) and progressively leads to physical disability and ...

A Volumetric Method for Quantification of Cerebral Vasospasm in a Murine Model of Subarachnoid Hemorrhage

Subarachnoid hemorrhage (SAH) is a subtype of hemorrhagic stroke. Cerebral vasospasm that occurs in the aftermath of the bleeding is an important factor determining patient outcome and is therefore frequently taken as a study endpoint. However, in small animal studies on SAH, quantification of cerebral vasospasm is a major challenge. Here, an ex vivo method is presented that allows quantification of volumes of entire vessel segments, which can be used as an objective measure to quantify cerebral vasospasm. In a first step, endovascular casting of the cerebral vasculature is performed using a radiopaque casting agent. Then, cross-sectional imaging data are acquired by micro computed tomography. The final step involves 3-dimensional reconstruction of the virtual vascular tree, followed by an algorithm to calculate center lines and volumes of the selected vessel segments. The method resulted in a highly accurate virtual reconstruction of the cerebrovascular tree shown by a diameter-based comparison of anatomical samples with their virtual reconstructions. Compared with vessel diameters alone, the vessel volumes highlight the differences between vasospastic and non-vasospastic vessels shown in a series of SAH and sham-operated mice.

The goal of this article is to present a method that allows a 3-dimensional reconstruction of the cerebrovascular tree in mice after micro computed tomography and determination of volumes of entire vessel segments that can be used to quantify cerebral vasospasm in murine models of subarachnoid hemorrhage.

Un método volumétrico para la cuantificación de vasoespasmo Cerebral en un modelo murino de hemorragia subaracnoidea

Subarachnoid hemorrhage (SAH) is a subtype of hemorrhagic stroke. Cerebral vasospasm that occurs in the aftermath of the bleeding is an important factor ...

Minimally Invasive Endoscopic Intracerebral Hemorrhage Evacuation

Intracerebral hemorrhage (ICH) is a subtype of stroke with high mortality and poor functional outcomes, largely because there are no evidence-based treatment options for this devastating disease process. In the past decade, a number of minimally invasive surgeries have emerged to address this issue, one of which is endoscopic evacuation. Stereotactic ICH Underwater Blood Aspiration (SCUBA) is a novel endoscopic evacuation technique performed in a fluid-filled cavity using an aspiration system to provide an additional degree of freedom during the procedure. The SCUBA procedure utilizes a suction device, endoscope, and sheath and is divided into two phases. The first phase involves maximal aspiration and minimal irrigation to decrease clot burden. The second phase involves increasing irrigation for visibility, decreasing aspiration strength for targeted aspiration without disturbing the cavity wall, and cauterizing any bleeding vessels. Using the endoscope and aspiration wand, this technique aims to maximize hematoma evacuation while minimizing collateral damage to the surrounding brain. Advantages of the SCUBA technique include the use of a low-profile endoscopic sheath minimizing brain disruption and improved visualization with a fluid-filled cavity rather than an air-filled one.

This paper details the surgical protocol for minimally invasive endoscopic intracerebral hemorrhage evacuation using the SCUBA technique.

Evacuación endoscópica mínimamente invasiva de hemorragia intracerebral

Intracerebral hemorrhage (ICH) is a subtype of stroke with high mortality and poor functional outcomes, largely because there are no evidence-based ...

In Vivo Direct Reprogramming of Resident Glial Cells into Interneurons by Intracerebral Injection of Viral Vectors

Converting resident glia in the brain into functional and subtype-specific neurons in vivo provides a step forward towards the development of alternative cell replacement therapies while also creating tools to study cell fate in situ. To date, it has been possible to obtain neurons via in vivo reprogramming, but the precise phenotype of these neurons or how they mature has not been analyzed in detail. In this protocol, we describe a more efficient conversion and cell-specific identification of the in vivo reprogrammed neurons, using an AAV-based viral vector system. We also provide a protocol for functional assessment of the reprogrammed cells&#8217; neuronal maturation. By injecting flip-excision (FLEX) vectors, containing the reprogramming and synapsin-driven reporter genes&#160;to specific cell types in the brain that serve as the target for cell reprogramming. This technique allows for the easy identification of newly reprogrammed neurons. Results show that the obtained reprogrammed neurons functionally mature over time, receive synaptic contacts and show electrophysiological properties of different types of interneurons. Using the transcription factors Ascl1, Lmx1a and Nurr1, the majority of the reprogrammed cells have properties of fast-spiking, parvalbumin-containing interneurons.

This protocol aims at generating directly reprogrammed interneurons in vivo, using an AAV-based viral system in the brain and a FLEX synapsin-driven GFP reporter, which allows for cell identification and further analysis in vivo.

En Vivo Reprogramación Directa de Células Gliales Residentes en Interneuronas por Inyección Intracerebral de Vectores Virales

Converting resident glia in the brain into functional and subtype-specific neurons in vivo provides a step forward towards the development of alternative ...

Double Direct Injection of Blood into the Cisterna Magna as a Model of Subarachnoid Hemorrhage

Among strokes, subarachnoid hemorrhage (SAH) consecutive to the rupture of a cerebral arterial aneurysm represents 5-9% but is responsible for about 30% of the total stroke-related mortality with an important morbidity in terms of neurological outcome. A delayed cerebral vasospasm (CVS) may occur most often in association with a delayed cerebral ischemia. Different animal models of SAH are now being used including endovascular perforation and direct injection of blood into the cisterna magna or even the prechiasmatic cistern, each exhibiting distinct advantages and disadvantages. In this article, a standardized mouse model of SAH by double direct injection of determined volumes of autologous whole blood into the cisterna magna is presented. Briefly, mice were weighed and then anesthetized by isoflurane inhalation. Then, the animal was placed in a reclining position on a heated blanket maintaining a rectal temperature of 37 &#176;C and positioned in a stereotactic frame with a cervical bend of about 30&#176;. Once in place, the tip of an elongated glass micropipette filled with the homologous arterial blood taken from carotid artery of another mouse of the same age and gender (C57Bl/6J) was positioned at a right angle in contact with the atlanto-occipital membrane by means of a micromanipulator. Then 60 &#181;L of blood was injected in the cisterna magna followed by a 30&#176; downward tilt of the animal for 2 minutes. The second infusion of 30 &#181;L of blood into the cisterna magna was performed 24 h after the first one. The individual follow-up of each animal is carried out daily (careful evaluation of weight and well-being). This procedure allows a predictable and highly reproducible distribution of blood, likely accompanied by intracranial pressure elevation that can be mimicked by an equivalent injection of an artificial cerebral spinal fluid (CSF), and represents an acute to mild-model of SAH inducing low mortality.

We described in this protocol a standardized subarachnoid hemorrhage (SAH) mouse model by a double injection of autologous whole-blood into the cisterna magna. The high degree of standardization of the double-injection procedure represents a middle-to-acute model of SAH with relative safety regarding mortality.

Doble inyección directa de sangre en la Cisterna Magna como modelo de hemorragia subaracnoidea

Among strokes, subarachnoid hemorrhage (SAH) consecutive to the rupture of a cerebral arterial aneurysm represents 5-9% but is responsible for about 30% ...

Massive Pontine Hemorrhage by Dual Injection of Autologous Blood

We provide a protocol to establish a massive pontine hemorrhage model in a rat. Rats weighing about 250 grams were used in this study. One hundred microliters of autologous blood was taken from the tail vein and stereotaxically injected into the pons. The injection process was divided into 2 steps: First, 10 &#181;L of blood was injected into a specific location, anteroposterior position (AP) -9.0 mm; lateral (Lat) 0 mm; vertical (Vert) -9.2 mm, followed by a second injection of the residual blood located at AP -9.0 mm; Lat 0 mm; Vert -9.0 mm with a 20-minute interval. The balance beam test, limb placement test, and the modified Voestch neuroscore were used to evaluate neurological function. Magnetic Resonance Imaging (MRI) was used to assess the volume of hemorrhage in vivo. The symptoms of this model were in line with patients with massive pontine hemorrhage.

We present a protocol to establish a massive pontine hemorrhage model in a rat via dual injection of autologous blood.

Hemorragia pontina masiva por inyección dual de sangre autóloga

We provide a protocol to establish a massive pontine hemorrhage model in a rat. Rats weighing about 250 grams were used in this study. One hundred ...

Pre-Chiasmatic, Single Injection of Autologous Blood to Induce Experimental Subarachnoid Hemorrhage in a Rat Model

Despite advances in treatment over the last decades, subarachnoid hemorrhage (SAH) continues to carry a high burden of morbidity and mortality, largely afflicting a fairly young population. Several animal models of SAH have been developed to investigate the pathophysiological mechanisms behind SAH and to test pharmacological interventions. The pre-chiasmatic, single injection model in the rat presented in this article is an experimental model of SAH with a predetermined blood volume. Briefly, the animal is anesthetized, intubated, and kept under mechanical ventilation. Temperature is regulated with a heating pad. A catheter is placed in the tail artery, enabling continuous blood pressure measurement as well as blood sampling. The atlantooccipital membrane is incised and a catheter for pressure recording is placed in the cisterna magna to enable intracerebral pressure measurement. This catheter can also be used for intrathecal therapeutic interventions. The rat is placed in a stereotaxic frame, a burr hole is drilled anteriorly to the bregma, and a catheter is inserted through the burr hole and placed just anterior to the optic chiasm. Autologous blood (0.3 mL) is withdrawn from the tail catheter and manually injected. This results in a rise of intracerebral pressure and a decrease of cerebral blood flow. The animal is kept sedated for 30 min and given subcutaneous saline and analgesics. The animal is extubated and returned to its cage. The pre-chiasmatic model has a high reproducibility rate and limited variation between animals due to the pre-determined blood volume. It mimics SAH in humans making it a relevant model for SAH research.

Subarachnoid hemorrhage continues to carry a high burden of mortality and morbidity in man. To facilitate further research into the condition and its pathophysiology, a pre-chiasmatic, single injection model is presented.

Inyección única prequiasmática de sangre autóloga para inducir hemorragia subaracnoidea experimental en un modelo de rata

Despite advances in treatment over the last decades, subarachnoid hemorrhage (SAH) continues to carry a high burden of morbidity and mortality, largely ...

Intracranial Pressure Monitoring In Nontraumatic Intraventricular Hemorrhage Rodent Model

Survivors of intraventricular hemorrhage are often left with significant long-term memory impairment; thus, research utilizing intraventricular hemorrhage animal models is essential. In this study, we sought out ways to measure intracranial pressure, mean arterial pressure, and cerebral perfusion pressure during nontraumatic intraventricular hemorrhage in rats. The experimental design included three Sprague Dawley groups: sham, standard 200 &#181;l intraventricular hemorrhage, and vehicle control groups. By introducing an intraparenchymal fiberoptic pressure sensor, precise intracranial pressure measurements were obtained in all groups. Cerebral perfusion pressures were calculated with the knowledge of intracranial pressure and mean arterial pressure values. As expected, the intraventricular hemorrhage and vehicle control groups both experienced a rise in the intracranial pressure and subsequent decline in cerebral perfusion pressure during intraventricular injection of autologous blood and artificial cerebrospinal fluid, respectively. The addition of an intraparenchymal fiberoptic pressure sensor is beneficial in monitoring precise intracranial pressure changes.

Monitoring intracranial pressure in rodent&#160;models of nontraumatic intraventricular hemorrhage is not common in the current literature. Herein, we demonstrate a technique for measuring intracranial pressure, mean arterial pressure, and cerebral perfusion pressure during intraventricular hemorrhage in a rat&#160;animal model.

Monitorización de la presión intracraneal en un modelo de roedor con hemorragia intraventricular no traumática

Survivors of intraventricular hemorrhage are often left with significant long-term memory impairment; thus, research utilizing intraventricular hemorrhage ...