Los autores desean agradecer a los donantes y sus familias por su generosa donación. Gracias al Sr. Xiao Xuan Li que grabaron el video y ayudó con la edición de vídeo; Apoyan a la Sra. Hannah Lewis y el Sr. Louis Szabo para proporcionar técnicas; y el profesor Jan Provis para revisar el video y entrada a contenido de vídeo.

todos los métodos aquí descritos han sido aprobados por el Comité de ética de investigación humana de la Universidad nacional australiana. Para crear el modelo ventricular utilizamos fibra Klingler disección técnica 12 , 14. La técnica de Klingler es un método de disección táctil que consiste en extirpar pequeñas porciones de la materia gris de la corteza y pelar de haces de fibras nerviosas, proporcionando así una guía paso a paso a través de las capas de tejido de la superficie de las estructuras profundas del cerebro.
 Nota: la muestra de cerebro utilizada para demostrar este protocolo en el vídeo adjunto y las imágenes se quitó cuidadosamente de un cadáver humano embalsamado en formol, obtenido desde el programa donantes de cuerpo de la Facultad de medicina, Australia Universidad Nacional. El donante no tenía antecedentes conocidos de enfermedad neuropathological. Después del retiro de la duramadre, el cerebro se almacenaba en solución de etanol del 10% a temperatura ambiente durante tres años.
 1. preparación <ol><li>obtener un total del cerebro de un cadáver humano embalsamado y retirar la duramadre y el cerebro en 10% de etanol a temperatura ambiente antes de la disección. 
	PRECAUCIÓN: Use equipo de protección personal en una habitación bien ventilada de acuerdo a las pautas locales al manejar. Asegurar que todos los participantes están familiarizados con los procedimientos institucionales para el manejo y eliminación de un bisturí y sharp los objetos antes de comenzar el protocolo de disección.</li>
	<li>Preparar los siguientes instrumentos: tijeras, pinzas, hojas de bisturí (Nº 15 y Nº 22), sonda de metal y el extremo romo de un mango de bisturí metálico ( figura 1). Utilice el extremo romo de la manija del escalpelo para minimizar el daño a las delicadas fibras del nervio y la conservación de los principales de la materia blanca fibra extensiones ( figura 2) 13.</li>
	<li>Colocar el cerebro de modo que su superficie ventral quede mirando hacia arriba.</li>
</ol> 2. Procedimiento de disección Nota: la disección toma aproximadamente 2 a 3 h para completar <ol><li>Retire el aracnoides y la vasculatura asociada de ambos hemisferios cerebrales con un par de pinzas atraumáticas (blunt).</li>
	<li>Suavemente Levante el cerebelo y localizar los colículos inferiores. Coloque la hoja de bisturí (Nº 15) unida a un mango de bisturí largo justo caudal a los colículos inferiores y corte axial a través del médula oblonga. Evitar que el disco tan cerca horizontal como sea posible para evitar daños en el cerebelo. Tenga cuidado de preservar El tectum del midbrain.</li>
	<li>Posición del cerebro para ver la fisura lateral derecho o izquierda. A partir de la convolución del cerebro supramarginal, utilice el extremo romo del mango del bisturí para eliminar suavemente las capas corticales superficiales. Suavemente avanzar primero por encima y por debajo del surco lateral para revelar los haces de fibras de asociación horizontal funcionando en el parietal, frontal y lóbulos temporales, respectivamente.</li>
	<li>Seguir la dirección de las fibras arqueadas alrededor del borde posterior de la ínsula conectando el fasciculi longitudinales superior e inferior para revelar el fascículo arqueado.</li>
	<li>Anterior, quite con cuidado las restantes superficiales capas corticales de los lóbulos frontales temporales de media y las convoluciones del cerebro frontal inferior para revelar las fibras fasciculares ganchudas que conectan los temporales y</li>
	<li>Identificar las corto convoluciones del cerebro de la corteza insular y luego retire la ínsula. A continuación retire la cápsula extrema y claustrum para revelar la cápsula externa subyacente. Tenga en cuenta la protuberancia formada por el núcleo lentiforme profundo a la cápsula. Hacia la superficie dorsal de la corteza, revelan las fibras de la corona radiata ( figura 4).</li>
	<li>Quitar la corteza restante y la materia blanca subyacente en la superficie dorsal del cerebro para llegar a la circunvolución del cíngulo. Continuar con el extremo romo de la manija del escalpelo para quitar la corteza del cingulate para revelar el cíngulo, los tractos de materia blanca que conecta la sustancia perforada anterior con la convolución del cerebro parahippocampal.</li>
	<li>Utiliza la misma técnica para extraer el cíngulo posterior a anterior revelan el cuerpo calloso, compuesto de fibras commissural que conectan los dos hemisferios cerebrales. El dorso del cuerpo (tronco) del cuerpo calloso será visible ( figura 6).</li>
	<li>Repita los pasos 2.3 a 2.8 en el hemisferio cerebral contralateral.</li>
	<li>Palpate e identificar la extensión del ventrículo lateral en uno de los hemisferios. Utilizando una sonda, perfore la pared lateral del ventrículo en el sitio del trígono colateral. Usando una hoja de tamaño 24 (adjunta a un mango de bisturí no. 4) introduce a través del sitio de punción y corte inferiorly a abrir toda la longitud del cuerno inferior del ventrículo lateral.</li>
	<li>Ahora retomar el trígono colateral ventricular para ampliar el corte superior hacia el splenium del callosum de la recopilación (línea punteada en la figura 5).</li>
	<li>Repetir los pasos 2.10 y 2.11 en el otro hemisferio.</li>
	<li>Abrir el cuerpo del ventrículo lateral continuando la incisión desde el trígono rostral con un corte de aproximadamente 3 cm paralela al cuerpo calloso en ambos hemisferios (líneas punteadas en la figura 6).</li>
	<li>Únete las dos incisiones paralelas en cada hemisferio rostral a la altura de la rodilla y caudalmente al nivel del splenium del callosum de la recopilación. Con unas pinzas, sostuvo en la mano no dominante, levante suavemente el cuerpo calloso en el Esplenio. Con un fuerte par de tijeras, la mano dominante, separar el splenium del pellucidum del septo subyacente. Una vez haya llegado el extremo rostral del cuerpo, cortar el cuerpo calloso y quitar it</li>
	<li>Nestle la superficie ventral del cerebro a lo largo de la palma de su mano no dominante para estabilizar las áreas occipitales y temporales (parte posterior). Al mismo tiempo, utilizar su mano dominante firme pero suavemente el extremo anterior del cerebro mediante la colocación de los dedos opuestos y pulgar en los núcleos lentiformes de ambos lados del cerebro.</li>
	Movimientos y <li>con suave jalando separados físicamente las partes anteriores y posteriores del cerebro teniendo especial cuidado de mantener intacto el plexo coroideo. Se recomienda que un colega esté presente para guiar a la separación y sección suavemente cualquier resto conectar los tejidos durante el proceso con un bisturí.</li>
</ol>

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Los autores declaran que no tienen ningún conflicto de interés.

El objetivo de este trabajo fue elaborar una guía de disección para su difusión a docentes y estudiantes que podrían utilizarse para mejorar la enseñanza y el aprendizaje de la profunda ventricular y periventricular de estructuras del cerebro humano. Hemos ideado a una guía paso a paso con imágenes, junto con un recurso de vídeo, que puede utilizarse para facilitar la comprensión de la morfología de los ventrículos y sus estructuras asociadas. La técnica de disección sí mismo no es nuevo. Disección de la f...

Muchos estudiantes luchan para entender los espacios negativos del sistema ventricular, situado en el cerebro humano1,2. Utilizados los recursos disponibles para los estudiantes para el estudio de los ventrículos proporcionan representaciones relativamente crudas de las intrincadas relaciones 3D de estas estructuras cerebrales profundas. Comprensión de la Anatomía 3D del sistema ventricular y las estructuras relacionadas es particularmente importante en neurocirugía ya que el acceso al sistema ventricular es una de las técnicas más utilizadas para medir la presión intracraneal, descomprimir el ventrículo sistema y administrar medicamentos3. Además, los avances rápidos en imágenes médicas han requerido el desarrollo de habilidades en la interpretación de la Anatomía 3D.
Dos dimensiones secciones (2D) del cerebro en diversos planos se suelen utilizar para visualizar las estructuras del cerebro profundo que forman los límites de los espacios ventriculares negativa4. Sin embargo, son insuficientes para permitir a los estudiantes a entender el alcance completo de la arquitectura 3D de los ventrículos y los detalles finos de la región como haces de fibras que conectan la corteza y estructuras subcorticales52D rebanadas del cerebro solamente. Por lo tanto, educadores que dependen de la capacidad de los estudiantes para calcular un concepto 3D comprensible de los ventrículos4. Los estudiantes que luchan con la conciencia espacial resulta extremadamente difícil crear esta imagen 3D. Mientras que los modelos de plástico y moldes ventriculares proporcionan una representación 3D del sistema ventricular, no llegan a demostrar la relación integral que forman los límites de los ventrículos. Los estudiantes a menudo sin pensar Saque las piezas del modelo plástico para acceder al sistema ventricular y comprender sus interconexiones. En este proceso, con frecuencia pasan por alto las posiciones relativas detalladas de cada estructura y perder la comprensión de sus relaciones (p. ej. formación del techo de los ventrículos laterales por el cuerpo calloso).
El desarrollo de nuevas herramientas de enseñanza informatizada ha abordado algunas de estas limitaciones. Sin embargo, muchos de estos modelos se limitan a imágenes y texto estático y no toman ventaja de la interactividad que ofrecen estas nuevas tecnologías7,8. Mientras que tecnologías interactivas que el usuario pueda rotar modelos 3D por computadora para estudiar múltiples puntos de vista, esto puede confundir a algunos usuarios, especialmente los principiantes que encuentran difícil orientarse estructuras6. Además, recursos informáticos interactivos han demostrado ser menos eficaz en la enseñanza de las estructuras anatómicas más complejas6. Por lo tanto, uno de los retos en la enseñanza de la neuroanatomía es proporcionar a los estudiantes con recursos que les permitan visualizar los ventrículos adecuadamente y apreciar su estructura 3D y relaciones anatómicas incluyendo la delicada asociativa, proyección, y haces de fibras commissural que forman relaciones complejas con las estructuras periventriculares del2.
La disección ha demostrado ser un excelente método educativo para el aprendizaje de anatomía7,8. Un reciente estudio proporciona evidencia de los beneficios de la disección de la estudiante en el aprendizaje de neuroanatomía. En 2016, Rae et al encontró mejor retención a corto plazo y a largo plazo de conocimiento de la neuroanatomía en los estudiantes que participan en las disecciones9. Mientras que los avances tecnológicos continúan mejorando la exactitud y la interactividad de modelos 3D por computadora, los conocimientos adquiridos a través de disección práctica no pueden repetirse digitalmente en el presente tiempo de10.
En este estudio, el objetivo fue producir una disección reproducible de un cerebro humano. Elegimos un método de disección de fibra ya que permite la preservación de los haces de fibras delicadas estructuras de la materia gris periventricular para mejor definen el espacio negativo de los ventrículos.
Aquí presentamos a una guía completa paso a paso para la creación de un modelo independiente de los ventrículos y las estructuras periventriculares junto con un acompañamiento vídeo de entrenamiento para usan en neuroanatomía de enseñanza y aprendizaje. Estos recursos se pueden utilizar para la enseñanza y el aprendizaje de la neuroanatomía del cerebro por educadores y estudiantes.

<table border="0" cellpadding="0" cellspacing="0" width="615">
	<tbody>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">Scalpel Blade No 15</td>
			<td style="width:113px;">Swann-Morton</td>
			<td style="width:119px;">0205</td>
			<td style="width:167px;">Scalpel blade</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Scalpel Blade No 11</td>
			<td style="width:113px;">Swann-Morton</td>
			<td style="width:119px;">0203</td>
			<td>Scalpel blade</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Scalpel Blade No 24</td>
			<td style="width:113px;">Swann-Morton</td>
			<td style="width:119px;">0211</td>
			<td>Scalpel blade</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Long Scalpel handle No3L</td>
			<td style="width:113px;">Swann-Morton</td>
			<td style="width:119px;">0913</td>
			<td>Scalpel handle</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Short Scalpel handle No4G</td>
			<td style="width:113px;">Swann-Morton</td>
			<td style="width:119px;">0934</td>
			<td>Scalpel handle</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Scissors</td>
			<td style="width:113px;"></td>
			<td style="width:119px;"></td>
			<td>Scissors</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Atraumatic Forceps</td>
			<td style="width:113px;"></td>
			<td style="width:119px;"></td>
			<td>Atraumatic forceps</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Toothed Forceps</td>
			<td style="width:113px;"></td>
			<td style="width:119px;"></td>
			<td>Toothed forceps</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Genelyn Arterial Enhanced</td>
			<td style="width:113px;">GMS Inovations</td>
			<td style="width:119px;">AE-475</td>
			<td>Arterial embalming media</td>
		</tr>
	</tbody>
</table>

exploring deep space - uncovering the anatomy of periventricular structures to reveal the lateral ventricles of the human brain

Los estudiantes de anatomía típicamente dos dimensiones secciones (2D) y de imágenes al estudiar anatomía ventricular cerebral y los estudiantes parece desafiante. Porque los ventrículos son espacios negativos profundamente dentro del cerebro, es la única manera de entender su anatomía por apreciar sus límites formados por estructuras relacionadas. Mirando una representación 2D de estos espacios, en cualquiera de los planos cardinales, no permitirá una visualización de todas las estructuras que forman los límites de los ventrículos. Así, usando secciones 2D solo requiere que los estudiantes a calcular su propia imagen mental de los espacios ventriculares 3D. El objetivo de este estudio fue desarrollar un método reproducible para la disección del cerebro humano para crear un recurso educativo para mejorar los estudiantes comprensión de las complejas relaciones entre los ventrículos y las estructuras periventriculares. Para lograr esto, hemos creado un recurso de video que ofrece a una guía paso a paso utilizando un método de disección de fibra para revelar los ventrículos lateral y terceros junto con las estructuras estrechamente relacionadas del sistema límbico y los ganglios basales. Una de las ventajas de este método es que permite la delineación de los tractos de materia blanca que son difíciles de distinguir de otras técnicas de disección. Este video va acompañado de un protocolo escrito que ofrece una descripción sistemática del proceso de ayuda en la reproducción de la disección del cerebro. Este paquete ofrece una valiosa anatomía enseñanza de recursos para educadores y estudiantes por igual. Siguiendo estas instrucciones educadores pueden crear recursos docentes y estudiantes pueden guiarse para producir su propia disección de cerebro como una actividad práctica práctica. Se recomienda que esta guía de video incorporados a neuroanatomía de enseñanza para mejorar el estudiante comprensión de la morfología y la relevancia clínica de los ventrículos.

Este método de disección expone el sistema ventricular mediante la separación del cerebro en una anterior y una parte posterior (figura 7 y figura 8). La parte posterior ofrece una visión interna al trígono colateral de que los cuernos posteriores e inferiores se pueden ver que se extiende hasta el occipital y lóbulos temporales, respectivamente (figura 8). En el cuerno inferi...

Watch this Scientific Journal Video about Exploring Deep Space - Uncovering the Anatomy of Periventricular Structures to Reveal the Lateral Ventricles of the Human Brain at JoVE.com

Exploring Deep Space - Uncovering the Anatomy of Periventricular Structures to Reveal the Lateral Ventricles of the Human Brain

Explorar el espacio profundo - descubrimiento de la anatomía de las estructuras periventriculares para revelar los ventrículos laterales del cerebro humano

Este trabajo demuestra el uso eficaz de un método de disección de fibra para revelar los tractos de sustancia blanca superficial y estructuras de periventricular del cerebro humano, en un espacio tridimensional, para facilitar la comprensión del estudiante de la morfología ventricular.

Anatomy students are typically provided with two-dimensional (2D) sections and images when studying cerebral ventricular anatomy and students find this ...

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Research

JoVE Journal

Neuroscience

16.5K vistas.  Australian National University. Este trabajo demuestra el uso eficaz de un método de disección de fibra para revelar los tractos de sustancia blanca superficial y estructuras de periventricular del cerebro humano, en un espacio tridimensional, para facilitar la comprensión del estudiante de la morfología ventricular.

Video: Exploring Deep Space - Uncovering the Anatomy of Periventricular Structures to Reveal the Lateral Ventricles of the Human Brain

Mouse Models of Periventricular Leukomalacia

We describe a protocol for establishing mouse models of periventricular leukomalacia (PVL). PVL is the predominant form of brain injury in premature infants and the most common antecedent of cerebral palsy. PVL is characterized by periventricular white matter damage with prominent oligodendroglial injury. Hypoxia/ischemia with or without systemic infection/inflammation are the primary causes of PVL. We use P6 mice to create models of neonatal brain injury by the induction of hypoxia/ischemia with or without systemic infection/inflammation with unilateral carotid ligation followed by exposure to hypoxia with or without injection of the endotoxin lipopolysaccharide (LPS). Immunohistochemistry of myelin basic protein (MBP) or O1 and electron microscopic examination show prominent myelin loss in cerebral white matter with additional damage to the hippocampus and thalamus. Establishment of mouse models of PVL will greatly facilitate the study of disease pathogenesis using available transgenic mouse strains, conduction of drug trials in a relatively high throughput manner to identify candidate therapeutic agents, and testing of stem cell transplantation using immunodeficiency mouse strains.

We established mouse models of periventricular leukomalacia (PVL), the predominant brain injury in premature infants characterized by periventricular white matter lesions. Hypoxia/ischemia with/without systemic infection are the primary causes of PVL. Unilateral carotid ligation and hypoxia exposure with/without lipopolysaccharide injection creates PVL-like lesions in P6 mice.

Los modelos de ratón de la Retinopatía

We describe a protocol for establishing mouse models of periventricular leukomalacia (PVL). PVL is the predominant form of brain injury in premature ...

Investigación

Neurociencias

Morphometric Analyses of Retinal Sections

Morphometric analyses of retinal sections have been used in examining retinal diseases. For examples, neuronal cells were significantly lost in the retinal ganglion cell layer (RGCL) in rat models with N-methyl-D-aspartate (NMDA)&#x2013;induced excitotoxicity1, retinal ischemia-reperfusion injury2 and glaucoma3. Reduction of INL and inner plexiform layer (IPL) thicknesses were reversed with citicoline treatment in rats' eyes subjected to kainic acid-mediated glutamate excitotoxicity4. Alteration of RGC density and soma sizes were observed with different drug treatments in eyes with elevated intraocular pressure3,5,6. Therefore, having objective methods of analyzing the retinal morphometries may be of great significance in evaluating retinal pathologies and the effectiveness of therapeutic strategies.
The retinal structure is multi-layers and several different kinds of neurons exist in the retina. The morphometric parameters of retina such as cell number, cell size and thickness of different layers are more complex than the cell culture system. Early on, these parameters can be detected using other commercial imaging software. The values are normally of relative value, and changing to the precise value may need further accurate calculation. Also, the tracing of the cell size and morphology may not be accurate and sensitive enough for statistic analysis, especially in the chronic glaucoma model. The measurements used in this protocol provided a more precise and easy way. And the absolute length of the line and size of the cell can be reported directly and easy to be copied to other files. For example, we traced the margin of the inner and outer most nuclei in the INL and formed a line then using the software to draw a 90 degree angle to measure the thickness. While without the help of the software, the line maybe oblique and the changing of retinal thickness may not be repeatable among individual observers. In addition, the number and density of RGCs can also be quantified. This protocol successfully decreases the variability in quantitating features of the retina, increases the sensitivity in detecting minimal changes.
 	This video will demonstrate three types of morphometric analyses of the retinal sections. They include measuring the INL thickness, quantifying the number of RGCs and measuring the sizes of RGCs in absolute value. These three analyses are carried out with Stereo Investigator (MBF Bioscience &mdash; MicroBrightField, Inc.). The technique can offer a simple but scientific platform for morphometric analyses.

This video demonstrates three types of morphometric analyses of the retina, which include measuring the inner nuclear layer thickness, quantifying the number of retinal ganglion cells (RGCs) and measuring the sizes of RGCs. The technique can offer a simple but scientific platform for morphometric analyses.

Los análisis morfométricos de las Secciones de retina

Morphometric analyses of retinal sections have been used in examining retinal diseases. For examples, neuronal cells were significantly lost in the ...

Manual Drainage of the Zebrafish Embryonic Brain Ventricles

Cerebrospinal fluid (CSF) is a protein rich fluid contained within the brain ventricles. It is present during early vertebrate embryonic development and persists throughout life. Adult CSF is thought to cushion the brain, remove waste, and carry secreted molecules1,2. In the adult and older embryo, the majority of CSF is made by the choroid plexus, a series of highly vascularized secretory regions located adjacent to the brain ventricles3-5. In zebrafish, the choroid plexus is fully formed at 144 hours post fertilization (hpf)6. Prior to this, in both zebrafish and other vertebrate embryos including mouse, a significant amount of embryonic CSF (eCSF) is present . These data and studies in chick suggest that the neuroepithelium is secretory early in development and may be the major source of eCSF prior to choroid plexus development7.
eCSF contains about three times more protein than adult CSF, suggesting that it may have an important role during development8,9. Studies in chick and mouse demonstrate that secreted factors in the eCSF, fluid pressure, or a combination of these, are important for neurogenesis, gene expression, cell proliferation, and cell survival in the neuroepithelium10-20. Proteomic analyses of human, rat, mouse, and chick eCSF have identified many proteins that may be necessary for CSF function. These include extracellular matrix components, apolipoproteins, osmotic pressure regulating proteins, and proteins involved in cell death and proliferation21-24. However, the complex functions of the eCSF are largely unknown.
We have developed a method for removing eCSF from zebrafish brain ventricles, thus allowing for identification of eCSF components and for analysis of the eCSF requirement during development. Although more eCSF can be collected from other vertebrate systems with larger embryos, eCSF can be collected from the earliest stages of zebrafish development, and under genetic or environmental conditions that lead to abnormal brain ventricle volume or morphology. Removal and collection of eCSF allows for mass spectrometric analysis, investigation of eCSF function, and reintroduction of select factors into the ventricles to assay their function. Thus the accessibility of the early zebrafish embryo allows for detailed analysis of eCSF function during development.

We present a method to collect cerebrospinal fluid (CSF) and to create a system which lacks CSF within the embryonic zebrafish brain ventricular system. This allows for further examination of CSF composition and its requirement during embryonic brain development.

Drenaje Manual de los ventrículos del cerebro embrionario de pez cebra

Cerebrospinal fluid  (CSF) is a protein rich fluid contained within the brain ventricles. It is  present during early vertebrate embryonic development and ...

3D Modeling of the Lateral Ventricles and Histological Characterization of Periventricular Tissue in Humans and Mouse

The ventricular system carries and circulates cerebral spinal fluid (CSF) and facilitates clearance of solutes and toxins from the brain. The functional units of the ventricles are ciliated epithelial cells termed ependymal cells, which line the ventricles and through ciliary action are capable of generating laminar flow of CSF at the ventricle surface. This monolayer of ependymal cells also provides barrier and filtration functions that promote exchange between brain interstitial fluids (ISF) and circulating CSF. Biochemical changes in the brain are thereby reflected in the composition of the CSF and destruction of the ependyma can disrupt the delicate balance of CSF and ISF exchange. In humans there is a strong correlation between lateral ventricle expansion and aging. Age-associated ventriculomegaly can occur even in the absence of dementia or obstruction of CSF flow. The exact cause and progression of ventriculomegaly is often unknown; however, enlarged ventricles can show regional and, often, extensive loss of ependymal cell coverage with ventricle surface astrogliosis and associated periventricular edema replacing the functional ependymal cell monolayer. Using MRI scans together with postmortem human brain tissue, we describe how to prepare, image and compile 3D renderings of lateral ventricle volumes, calculate lateral ventricle volumes, and characterize periventricular tissue through immunohistochemical analysis of en face lateral ventricle wall tissue preparations. Corresponding analyses of mouse brain tissue are also presented supporting the use of mouse models as a means to evaluate changes to the lateral ventricles and periventricular tissue found in human aging and disease. Together, these protocols allow investigations into the cause and effect of ventriculomegaly and highlight techniques to study ventricular system health and its important barrier and filtration functions within the brain.

Using MRI scans (human), 3D imaging software, and immunohistological analysis, we document changes to the brain&#8217;s lateral ventricles. Longitudinal 3D mapping of lateral ventricle volume changes and characterization of periventricular cellular changes that occur in the human brain due to aging or disease are then modeled in mice.

Modelado 3D del lateral Ventrículos y histológico Caracterización de Tejidos periventricular en los seres humanos y de ratón

The ventricular system carries and circulates cerebral spinal fluid (CSF) and facilitates clearance of solutes and toxins from the brain. The functional ...

Analysis of Gene Expression Changes in the Rat Hippocampus After Deep Brain Stimulation of the Anterior Thalamic Nucleus

Deep brain stimulation (DBS) surgery, targeting various regions of the brain such as the basal ganglia, thalamus, and subthalamic regions, is an effective treatment for several movement disorders that have failed to respond to medication. Recent progress in the field of DBS surgery has begun to extend the application of this surgical technique to other conditions as diverse as morbid obesity, depression and obsessive compulsive disorder. Despite these expanding indications, little is known about the underlying physiological mechanisms that facilitate the beneficial effects of DBS surgery. One approach to this question is to perform gene expression analysis in neurons that receive the electrical stimulation. Previous studies have shown that neurogenesis in the rat dentate gyrus is elicited in DBS targeting of the anterior nucleus of the thalamus1. DBS surgery targeting the ATN is used widely for treatment refractory epilepsy. It is thus of much interest for us to explore the transcriptional changes induced by electrically stimulating the ATN. In this manuscript, we describe our methodologies for stereotactically-guided DBS surgery targeting the ATN in adult male Wistar rats. We also discuss the subsequent steps for tissue dissection, RNA isolation, cDNA preparation and quantitative RT-PCR for measuring gene expression changes. This method could be applied and modified for stimulating the basal ganglia and other regions of the brain commonly clinically targeted. The gene expression study described here assumes a candidate target gene approach for discovering molecular players that could be directing the mechanism for DBS.

The mechanism underlying the therapeutic effects of Deep Brain Stimulation (DBS) surgery needs investigation. The methods presented in this manuscript describe an experimental approach to examine the cellular events triggered by DBS by analyzing the gene expression profile of candidate genes that can facilitate neurogenesis post DBS surgery.

Análisis de los cambios de expresión génica en la Rata Hippocampus Después de la estimulación cerebral profunda del tálamo anterior Núcleo

Deep brain stimulation (DBS) surgery, targeting various regions of the brain such as the basal ganglia, thalamus, and subthalamic regions, is an effective ...

Investigating the Function of Deep Cortical and Subcortical Structures Using Stereotactic Electroencephalography: Lessons from the Anterior Cingulate Cortex

Stereotactic Electroencephalography (SEEG) is a technique used to localize seizure foci in patients with medically intractable epilepsy. This procedure involves the chronic placement of multiple depth electrodes into regions of the brain typically inaccessible via subdural grid electrode placement. SEEG thus provides a unique opportunity to investigate brain function. In this paper we demonstrate how SEEG can be used to investigate the role of the dorsal anterior cingulate cortex (dACC) in cognitive control. We include a description of the SEEG procedure, demonstrating the surgical placement of the electrodes. We describe the components and process required to record local field potential (LFP) data from consenting subjects while they are engaged in a behavioral task. In the example provided, subjects play a cognitive interference task, and we demonstrate how signals are recorded and analyzed from electrodes in the dorsal anterior cingulate cortex, an area intimately involved in decision-making. We conclude with further suggestions of ways in which this method can be used for investigating human cognitive processes.

Stereotactic Electroencephalography (SEEG) is an operative technique used in epilepsy surgery to help localize seizure foci. It also affords a unique opportunity to investigate brain function. Here we describe how SEEG can be used to investigate cognitive processes in human subjects.

La investigación de la función de profunda cortical y estructuras subcorticales Usando estereotáctica Electroencefalografía: Lecciones de la corteza cingulada anterior

Stereotactic Electroencephalography (SEEG) is a technique used to localize seizure foci in patients with medically intractable epilepsy. This procedure ...

Live Imaging of the Ependymal Cilia in the Lateral Ventricles of the Mouse Brain

Multiciliated ependymal cells line the ventricles in the adult brain. Abnormal function or structure of ependymal cilia is associated with various neurological deficits. The current ex vivo live imaging of motile ependymal cilia technique allows for a detailed study of ciliary dynamics following several steps. These steps include: mice euthanasia with carbon dioxide according to protocols of The University of Toledo&#8217;s Institutional Animal Care and Use Committee (IACUC); craniectomy followed by brain removal and sagittal brain dissection with a vibratome or sharp blade to obtain very thin sections through the brain lateral ventricles, where the ependymal cilia can be visualized. Incubation of the brain&#8217;s slices in a customized glass-bottom plate containing Dulbecco&#8217;s Modified Eagle&#8217;s Medium (DMEM)/High-Glucose at 37 &#176;C in the presence of 95%/5% O2/CO2 mixture is essential to keep the tissue alive during the experiment. A video of the cilia beating is then recorded using a high-resolution differential interference contrast microscope. The video is then analyzed frame by frame to calculate the ciliary beating frequency. This allows distinct classification of the ependymal cells into three categories or types based on their ciliary beating frequency and angle. Furthermore, this technique allows the use of high-speed fluorescence imaging analysis to characterize the unique intracellular calcium oscillation properties of ependymal cells as well as the effect of pharmacological agents on the calcium oscillations and the ciliary beating frequency. In addition, this technique is suitable for immunofluorescence imaging for ciliary structure and ciliary protein localization studies. This is particularly important in disease diagnosis and phenotype studies. The main limitation of the technique is attributed to the decrease in live motile cilia movement as the brain tissue starts to die.

Using high-resolution differential interference contrast (DIC) microscopy, an ex vivo observation of the beating of motile ependymal cilia located within the mouse brain ventricles is demonstrated by live-imaging. The technique allows a recording of the unique ciliary beating frequency and beating angle as well as their intracellular calcium oscillation pacing properties.

Imágenes en vivo de la ependimarios cilios en los ventrículos laterales del cerebro del ratón

Multiciliated ependymal cells line the ventricles in the adult brain. Abnormal function or structure of ependymal cilia is associated with various ...

A Simple One-step Dissection Protocol for Whole-mount Preparation of Adult Drosophila Brains

There is an increasing interest in using Drosophila to model human brain degenerative diseases, map neuronal circuitries in adult brains, and study the molecular and cellular basis of higher brain functions. A whole-mount preparation of adult brains with well-preserved morphology is critical for such whole brain-based studies, but can be technically challenging and time-consuming. This protocol describes an easy-to-learn, one-step dissection approach of an adult fly head in less than 10 s, while keeping the intact brain attached to the rest of the body to facilitate subsequent processing steps. The procedure helps remove most of the eye and tracheal tissues normally associated with the brain that can interfere with the later imaging step, and also places less demand on the quality of the dissecting forceps. Additionally, we describe a simple method that allows convenient flipping of the mounted brain samples on a coverslip, which is important for imaging both sides of the brains with similar signal intensity and quality. As an example of the protocol, we present an analysis of dopaminergic (DA) neurons in adult brains of WT (w1118) flies. The high efficacy of the dissection method makes it particularly useful for large-scale adult brain-based studies in Drosophila.

A Simple One-step Dissection Protocol for Whole-mount Preparation of Adult Drosophila Brains

The adult Drosophila brain is a valuable system for studying neuronal circuitry, higher brain functions, and complex disorders. An efficient method to dissect whole brain tissue from the small fly head will facilitate brain-based studies. Here we describe a simple, one-step dissection protocol of adult brains with well-preserved morphology.

Un simple de un solo paso para la disección Protocolo Todo el montaje Preparación de adulto<em&gt; Drosophila</emLos cerebros&gt;

There is an increasing interest in using Drosophila to model human brain degenerative diseases, map neuronal circuitries in adult brains, and study the ...

Sublimation of DAN Matrix for the Detection and Visualization of Gangliosides in Rat Brain Tissue for MALDI Imaging Mass Spectrometry

Sample preparation is key for optimal detection and visualization of analytes in Matrix-assisted Laser Desorption/Ionization (MALDI) Imaging Mass Spectrometry (IMS) experiments. Determining the appropriate protocol to follow throughout the sample preparation process can be difficult as each step must be optimized to comply with the unique characteristics of the analytes of interest. This process involves not only finding a compatible matrix that can desorb and ionize the molecules of interest efficiently, but also selecting the appropriate matrix deposition technique. For example, a wet matrix deposition technique, which entails dissolving a matrix in solvent, is superior for desorption of most proteins and peptides, whereas dry matrix deposition techniques are particularly effective for ionization of lipids. Sublimation has been reported as a highly efficient method of dry matrix deposition for the detection of lipids in tissue by MALDI IMS due to the homogeneity of matrix crystal deposition and minimal analyte delocalization as compared to many wet deposition methods 1,2. Broadly, it involves placing a sample and powdered matrix in a vacuum-sealed chamber with the samples pressed against a cold surface. The apparatus is then lowered into a heated bath (sand or oil), resulting in sublimation of the powdered matrix onto the cooled tissue sample surface. Here we describe a sublimation protocol using 1,5-diaminonaphthalene (DAN) matrix for the detection and visualization of gangliosides in the rat brain using MALDI IMS.

A protocol for the sublimation of DAN matrix onto rat brain tissue for the detection of gangliosides using MALDI Imaging Mass Spectrometry is presented.

Sublimación de DAN Matriz para la detección y visualización de gangliósidos en el tejido cerebral de la rata por MALDI Imaging Espectrometría de Masas

Sample preparation is key for optimal detection and visualization of analytes in Matrix-assisted Laser Desorption/Ionization (MALDI) Imaging Mass ...

Recording Synaptic Plasticity in Acute Hippocampal Slices Maintained in a Small-volume Recycling-, Perfusion-, and Submersion-type Chamber System

Even though experiments on brain slices have been in use since 1951, problems remain that reduce the probability of achieving a stable and successful analysis of synaptic transmission modulation when performing field potential or intracellular recordings. This manuscript describes methodological aspects that might be helpful in improving experimental conditions for the maintenance of acute brain slices and for recording field excitatory postsynaptic potentials in a commercially available submersion chamber with an outflow-carbogenation unit. The outflow-carbogenation helps to stabilize the oxygen level in experiments that rely on the recycling of a small buffer reservoir to enhance the cost-efficiency of drug experiments. In addition, the manuscript presents representative experiments that examine the effects of different carbogenation modes and stimulation paradigms on the activity-dependent synaptic plasticity of synaptic transmission.

This protocol describes the stabilization of the oxygen level in a small volume of recycled buffer and methodological aspects of recording activity-dependent synaptic plasticity in submerged acute hippocampal slices.

Grabación de la plasticidad sináptica en rebanadas Hippocampal agudas mantenidas en un pequeño volumen reciclado - perfusión- y sistema de cámara de tipo inmersión

Even though experiments on brain slices have been in use since 1951, problems remain that reduce the probability of achieving a stable and successful ...