Este estudio fue apoyado por el Instituto Nacional sobre el Abuso de Drogas Subvenciones DA 006886, 032270 y DA.

<ol><li> Cablear los motores paso a paso bipolares enroscando los cables en los conectores que se suministran con la tarjeta de drivers. Colores de los cables de motores paso a paso bipolares están estandarizados (Figura 1). Nota: Los motores paso a paso se describen tienen una resolución de 1,8 ° / paso, orientado a 0,346 ° / paso. A stereotax estándar tiene 3 mm/360 ° de recorrido. La resolución final es de 2.88 m / paso. Los motores son también capaces de paso a paso fraccional. <ol><li> Conecte el cable verde a A +, conecte el cable negro a A-, conecte el cable rojo a B + y conectar el cable azul a B-. </li></ol></li><li> Deslice los enganches en los motores paso a paso, con cuidado para alinear los orificios de montaje y fije usando 12x Tornillos M3 de cabeza hueca (20 mm) (Figura 2). <ol><li> Asegúrese de que los acopladores estén firmemente sujetos a los motores. Nota: Los modelos 3D no incluyen roscas. Las partes se etiquetan con los diámetros de rosca, pero deben ser aprovechados después de que sefabricado. </li></ol></li><li> Quite los tornillos de fijación de los apoyos para el pulgar en los 3 ejes del instrumento estereotáxico utilizando una llave hexagonal pequeña. Los apoyos para el pulgar son roscados, para convertirlos a la izquierda para la extracción. Mantenga las arandelas de PTFE en su sitio en el brazo (Figura 3). </li><li> Atornillar el extremo roscado de los collares sobre las varillas roscadas de los brazos del instrumento estereotáxico (Figura 3). <ol><li> Asegúrese de que no hay diferencia entre los collares y PTFE juntas tóricas. Esto garantiza que las coordenadas se mantienen cuando el robot cambia de dirección. </li><li> Asegure los collares en la rosca de los brazos estereotáxicas utilizando 3x NF10-32 (1/4 de pulgada) tornillos de punta ahuecada. </li><li> Deslizar cada motor y el acoplador sobre los collarines y los brazos estereotáxica. Asegúrese de que los motores se sientan al ras con los brazos, y los agujeros de tornillo de fijación en los collares se alinean con la parte plana de los ejes del motor (Figura 4). </li><li> Asegure la couplers al stereotax utilizando los orificios de montaje y 6x NF10-32 (1/2 pulgada) tornillos de punta ahuecada (Figura 4). </li><li> Asegure los cuellos de los ejes del motor utilizando 3x NF10-32 (1/4 de pulgada) tornillos de fijación (Figura 4). </li></ol></li><li> Prepare el tablero de conductor del CNC mediante la creación de cada uno de los pines del controlador a medio paso a paso. Nota: Este controlador de motor paso a paso se presenta como una placa de circuito expuesto. Un caso puede ser construido, aunque no es necesario. También, se puede utilizar un número de diferentes controladores de motor paso a paso bipolar. Si es así, asegúrese de que las instrucciones de configuración se siguen para la junta que haya adquirido. <ol><li> Alinear los 6 alfileres por motor paso a paso de la misma manera. La mitad-paso a paso permite el doble de la resolución de paso en grados / paso (Figura 5). </li><li> Voltear el pin 1 a la posición de encendido, el pin 2 a la posición de apagado, el pin 3 para ponerlo en la posiciónción, el pin 4 a la posición de apagado, el pin 5 a la posición de encendido, y la patilla 6 a la posición de apagado (Figura 5). </li></ol></li><li> Enchufe los motores (X - Y - Z) en el controlador de motor paso a paso, junto con la fuente de alimentación de 12 V. La colocación correcta se marca en el controlador. También, conecte el conductor de pasos al puerto serie de un ordenador mediante un DB25cable (Figura 6). </li><li> Instalar el software de fresado CNC a un ordenador personal (este tendrá que ser situado en una zona quirúrgica) siguiendo las instrucciones predeterminadas. Una vez instalado, abra el software para comenzar la configuración. <ol><li> Configurar el software para comunicarse con los motores paso a paso. Nota: Las siguientes instrucciones son para uso exclusivo con el conductor del motor de pasos TB6560. </li><li> Haga clic a través de los menús del software de la siguiente manera. Abrir → Config → Puertos y pines→ Señales de salida. Rellene el símbolo para que coincida con la Figura 7 y pulse Aplicar. </li><li> Haga clic a través de los menús del software de la siguiente manera. Abrir → Config → Puertos y pines → Señales de entrada. Rellene el símbolo para que coincida con la Figura 8 y pulse Aplicar. </li><li> Haga clic a través de los menús del software de la siguiente manera. Abra → Config → Puertos y Pasadores → Salidas de motor. Rellene el símbolo para que coincida con la Figura 9 y pulse Aplicar. </li><li> Haga clic a través de los menús del software de la siguiente manera. Abrir → tuning Config → Motor. Rellene el símbolo para que coincida con la Figura 10 y haga clic en Guardar configuración del Eje. Repita el paso anterior para los 3 ejes con los mismos valores. </li></ol></li><li> Calibrar el stereotax a la escala de los programas de CNC. Nota: El software está diseñado para máquinas de fresado estándar, por lo que su unidad de medida no será proporcional al recorrido de un instrumento estereotáxico.<ol><li> Ajuste la velocidad de los motores &#39;a 1 pulgada por minuto, y &quot;jog&quot; Z-eje del instrumento estereotáxico con RePág / AvPág al milímetro. Nota: La velocidad de avance lento recomendada máxima es de 3500 m / s, y la velocidad de corte máxima recomendada es de 500 m / seg. </li><li> Poner a cero el eje Z, y el Jog estereotáxica instrumento 1 mm. La distancia recorrida en el eje Z en Mach3 es la &quot;constante de escala&quot;. Coordenadas de la máquina se determinan multiplicando las coordenadas del cráneo (mm) por la &quot;constante de escala&quot;. </li><li> Realizar pruebas aleatorias de los 3 ejes programándolas viajar algunas distancias conocidas, y asegurar que los movimientos son precisos. Si el recorrido stereotax está demasiado lejos o definitiva, modificar la constante de escala en consecuencia. Nota: Una vez que la escala es completa, los scripts incluidos se pueden utilizar para generar código G para las cirugías. Sin embargo, se recomienda encarecidamente que los usuarios se familiaricen con el código G antes de intentar generar automáticamentecirugías. Esto es imprescindible para la solución de problemas y la modificación de las cirugías automatizados. </li></ol></li><li> Fije el taladro micro motor para el instrumento estereotáxico se utiliza el soporte extra grande sonda. Nota: La velocidad de la broca mínima recomendada es de 40.000 rpm. </li><li> Generación automática de código G para una craneotomía borde afilado con 3 orificios de los tornillos del cráneo. <ol><li> Coloque todos los scripts de la tabla de software en una sola carpeta en un PC. </li><li> Abra el script &quot;SharpEdgeCraniotomy.m&quot; y ejecutar el código. </li><li> Seleccionar Tanto al mensaje &quot;¿Qué tipo de cirugía usted estará presentando?&quot; (Figura 11). </li><li> Seleccione Personalizado para definir las esquinas de la ventana de cráneo. Rellene cada pregunta para que coincida con la figura 12. </li><li> Definir las posiciones X e Y de las esquinas de craneotomía. Cada coordenada se debe introducir en el orden correcto, según el ejemplo de la Figura 13 </strong&gt;. </li><li> Introduzca 3 en el símbolo para producir 3 agujeros del cráneo (Figura 14). </li><li> Seleccione Definir mediante coordenadas, e introduzca las coordenadas de cada agujero de la plantilla en la Figura 15. Nota: Si las coordenadas exactas no son importantes, hay una opción para apuntar y hacer clic en las posiciones de los agujeros &#39;en una imagen de un cráneo de la rata. Las posiciones serán generados automáticamente. </li><li> Defina los parámetros de perforación. Por primera cirugía de pruebas, acepte los valores por defecto. Nota: Estos valores dependen de los motores paso a paso, y el cráneo del animal. Cada lugar de la raza de la rata y el objetivo tiene un grosor del cráneo ligeramente diferente. Para las pocas cirugías iniciales usando este dispositivo, esté preparado para poner a prueba la profundidad de perforación y retire todos los trozos de cráneo restantes manualmente. Los valores se pueden modificar para futuras cirugías (Figura 16). </li><li> Nombra el código G, sino que se generará de forma automática y saved al directorio de trabajo. </li></ol></li><li> Cargue el código G en el software de fresado CNC y un cráneo de ensayo en los instrumentos estereotáxicas bares del oído. <ol><li> Refrescar manualmente la broca para Bregma con las teclas de flecha. Utilice una velocidad de trote lento (~ 5 pulgadas / m) para asegurar la exactitud. </li><li> Iniciar la broca que gira a mayor que 38.000 rpm. </li><li> Presione CycleStart, el stereotax realizará muchos pases del mismo corte, a diferentes profundidades. Entre cada paso, el stereotax hará una pausa, por lo que el cirujano puede continuar o cancelar el corte. Pulse Continuar Ciclo (Alt-R) para continuar cortando pases. </li></ol></li></ol>

<ol>
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Los autores no tienen intereses financieros en competencia a revelar.

El uso de equipos de cirugía automatizada ayuda a eliminar algunos de los problemas más comunes en la investigación en neurociencias. En primer lugar, las trayectorias de la herramienta son 100% reproducible. Cada corte está garantizado a estar en la misma ubicación con respecto a Bregma. En segundo lugar, se debe reducir el error experimentador. Aunque muchos investigadores son cirujanos altamente calificados, se necesita una cantidad excepcional de la práctica a ser aún un cirujano competente. Este dispositivo ...

La cirugía estereotáctica roedor se utiliza en una amplia variedad de aplicaciones de neurociencia, incluyendo lesioning 1, iontoforesis 2, la implantación microhilo 3, 4 estimulación, y de formación de imágenes del cráneo delgada 5. Sin embargo, hay grandes obstáculos que enfrentan aquellos que desean aplicar estas técnicas, incluyendo la curva de aprendizaje para la realización de la cirugía estereotáctica exacta y la alta probabilidad de error humano. Los errores humanos incluyen la medición y fallas de cálculo, así como la poca precisión y replicabilidad de los movimientos humanos. En un esfuerzo para reducir estos errores de confusión, los cirujanos estereotácticas se beneficiarían de un sistema que asegura que todos los procedimientos quirúrgicos se realizan de forma idéntica en todos los sujetos. La reducción de los errores también es un método por el cual los investigadores pueden reducir al mínimo el uso de sujetos animales, un objetivo primordial de los Institutos Nacionales de la Salud para la experimentación animal 6. En un mundo ideal, todos los scirugías tereotactic serían perfectamente reproducible dentro de los experimentos y entre laboratorios. Para solucionar este problema, las empresas han desarrollado nuevos stereotaxics ultra-precisos, y pantallas digitales para la lectura de las mediciones. Para eliminar los errores de movimiento humano, manipuladores y stereotaxics micro motorizados fueron producidos comercialmente, pero su alto coste puede ser prohibitivo para un laboratorio con un presupuesto limitado. Además, su software es totalmente de propiedad, y no puede ser modificado por el investigador para dar cabida a un nuevo tipo de cirugía. Una solución asequible para el problema de error humano es construir una stereotax robótico del modelo existente de un laboratorio, utilizando un equipo estándar de la industria CNC. Debido a la creciente comunidad de aficionados del CNC, los materiales son mucho más baratos que los equipos científicos. Esto permite a uno construir un instrumento estereotáxico CNC precisa, que también es muy flexible y de bajo costo. Con un conocimiento básico de mecanizado CNC y G-Code, individualmenteals pueden programar cualquier cirugía estereotáctica que se imaginan, sin las limitaciones de software propietario. Y, a fin de acelerar la producción de g-código para cirugías simples, este protocolo incluye un software que permite al usuario diseñar cirugías (craneotomía afilada punta, ventanas cráneo delgado, taladrado y de implantes bajar) dentro de apuntar y hacer clic menús. Estos programas de salida de un código de g concluido que se pueden ejecutar directamente desde el software de CNC. En total, una actualización estereotáxica motorizado es ideal para aquellos que tienen un interés en el aumento de la precisión y la repetibilidad de las cirugías, al tiempo que conserva la flexibilidad y el bajo coste de una plataforma de código abierto.

<table border="0" cellpadding="0" cellspacing="0" width="749">
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	<tbody>
		<tr height="44">
			<td height="44" style="height:44px;width:141px;">1x Standard U Frame Stereotax</td>
			<td style="width:137px;">Kopf</td>
			<td style="width:117px;"><a href="http://www.kopfinstruments.com/Stereotaxic/900.html">Kopf</a></td>
			<td style="width:355px;">This protocol should work with most existing stereotaxic devices.</td>
		</tr>
		<tr height="65">
			<td height="65" style="height:65px;width:141px;">3x 12 V, 1.6 A, 233 oz-inch Geared Bipolar Stepper Motor</td>
			<td style="width:137px;">Phidgets</td>
			<td style="width:117px;"><a href="http://www.robotshop.com/geared-bipolar-stepper-motor-3v-233-oz-in.html">Robot Shop</a></td>
			<td style="width:355px;">Any high torque geared stepper motor should do.&#160;</td>
		</tr>
		<tr height="73">
			<td height="73" style="height:73px;width:141px;">1x 3 Axis CNC Stepper Motor Driver Board Controller</td>
			<td style="width:137px;">Toshiba</td>
			<td style="width:117px;"><a href="http://www.ebay.com/itm/CNC-Router-3-Axis-TB6560-3-5A-Stepper-Motor-Driver-Board-For-Engraving-Machine-/300981888819">Ebay</a></td>
			<td style="width:355px;">Any 3 Axis CNC driver should do. Linked Item includes Mach3 CNC software.&#160;</td>
		</tr>
		<tr height="94">
			<td height="94" style="height:94px;width:141px;">2x Arm Couplers: medial-lateral (ML) &#38; dorsal-ventral (DV)</td>
			<td style="width:137px;">custom machined</td>
			<td style="width:117px;"><a href="http://eden.rutgers.edu/~mrcoffey/images/3D%20Models%20-%20CNC%20Stereotax%20Parts.zip">Part Drawings</a></td>
			<td style="width:355px;">These must be machined by your local machine shop. (costs will vary)</td>
		</tr>
		<tr height="76">
			<td height="76" style="height:76px;width:141px;">1x anterior-posterior (AP) Coupler</td>
			<td style="width:137px;">custom machined</td>
			<td style="width:117px;"><a href="http://eden.rutgers.edu/~mrcoffey/images/3D%20Models%20-%20CNC%20Stereotax%20Parts.zip">Part Drawings</a></td>
			<td style="width:355px;">These must be machined by your local machine shop. (costs will vary)</td>
		</tr>
		<tr height="127">
			<td height="172" rowspan="3" style="height:172px;width:141px;">3x Motor to Stereotax Collar</td>
			<td rowspan="3" style="width:137px;">custom machined</td>
			<td style="width:117px;"><a href="http://eden.rutgers.edu/~mrcoffey/images/3D%20Models%20-%20CNC%20Stereotax%20Parts.zip">Part Drawings</a></td>
			<td rowspan="3" style="width:355px;">These must be machined by your local machine shop. (costs will vary)</td>
		</tr>
		<tr height="23">
			<td height="23" style="height:23px;width:117px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:117px;"><a href="http://eden.rutgers.edu/~mrcoffey/images/Collar.html">View in Browser</a></td>
		</tr>
		<tr height="43">
			<td height="65" rowspan="2" style="height:65px;width:141px;">12x NF10-32 Cup Point Set Screws</td>
			<td rowspan="2" style="width:137px;">McMaster Carr</td>
			<td style="width:117px;"><a href="http://www.mcmaster.com/#92313A829">&#189;&#8221; Length</a></td>
			<td rowspan="2" style="width:355px;">You will need 6 of each.</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:117px;"><a href="http://www.mcmaster.com/#92313A825">&#188;&#8221; Length</a></td>
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		<tr height="46">
			<td height="46" style="height:46px;width:141px;">12x M3 Socket Head Screws (20 mm)</td>
			<td style="width:137px;">McMaster Carr</td>
			<td style="width:117px;"><a href="http://www.mcmaster.com/#92855A319">20mm Length</a></td>
			<td style="width:355px;">You will need 4 for each motor</td>
		</tr>
		<tr height="48">
			<td height="48" style="height:48px;width:141px;">1x Micro-Motor Drill&#160;</td>
			<td style="width:137px;">Buffalo Dental</td>
			<td style="width:117px;"><a href="http://www.buffalodental.com/pdfnew/x50.pdf">X50</a></td>
			<td style="width:355px;">Any Micromotor drill will work.&#160; At least 38,000 rpm recommended</td>
		</tr>
		<tr height="53">
			<td height="53" style="height:53px;width:141px;">1x 12 V DC Power Supply</td>
			<td style="width:137px;">12 Volt Adapters</td>
			<td style="width:117px;"><a href="http://www.12vadapters.com/adapter/power-supply/12v/7-amp-7a.html">12v DC &#8211; 7 Amp</a></td>
			<td style="width:355px;">Any 12 V DC PSU should work (ensure amperage rating is higher than the sum of the motors&#8217; amperage).</td>
		</tr>
		<tr height="44">
			<td height="44" style="height:44px;width:141px;">1x Extra Large Probe Holder</td>
			<td style="width:137px;">Stoelting</td>
			<td><a href="http://www.stoeltingco.com/neuroscience/accessories/stereotaxic-probe-holders/extra-large-probe-holder-551.html">Stoelting</a></td>
			<td style="width:355px;"></td>
		</tr>
		<tr height="44">
			<td height="44" style="height:44px;width:141px;">1x Grade B Rat Skull</td>
			<td style="width:137px;">Skulls Unlimited</td>
			<td><a href="http://www.skullsunlimited.com/record_variant.php?id=4009">Skulls Unlimited</a></td>
			<td style="width:355px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:141px;">Mach 3 Mill</td>
			<td style="width:137px;">ArtSoft USA</td>
			<td style="width:117px;"><a href="http://www.machsupport.com/ccount/click.php?id=1">Trial Download</a></td>
			<td style="width:355px;">Any Standard CNC controlling software should work.</td>
		</tr>
		<tr height="44">
			<td height="44" style="height:44px;width:141px;">Surgery Designer</td>
			<td style="width:137px;">Kevin Coffey &#38; David Barker</td>
			<td style="width:117px;"><a href="http://www.mathworks.com/matlabcentral/fileexchange/40282-robot-assisted-stereotactic-surgery">MATLAB File Exchange</a></td>
			<td style="width:355px;">These codes are available to modify. We accept no responsibility for your use or modification of code.</td>
		</tr>
	</tbody>
</table>

building an open-source robotic stereotaxic instrument

Este protocolo incluye los diseños y el software necesarios para actualizar un instrumento estereotáctico existente a un robot (CNC) instrumento estereotáxica por alrededor de $ 1.000 (sin incluir un taladro), utilizando motores de pasos estándar de la industria y el software de control CNC. Cada eje tiene control de velocidad variable y puede ser operado de manera simultánea o independientemente. La flexibilidad del robot y el sistema de codificación abierta (g-code) que sea capaz de realizar las tareas personalizadas que no son compatibles con los sistemas comerciales. Sus aplicaciones incluyen, pero no se limitan a, los agujeros de perforación, craneotomías borde afilado, adelgazamiento cráneo, y electrodos o cánula de descenso. Con el fin de agilizar la redacción del g-codificación para cirugías simples, hemos desarrollado scripts personalizados que permiten a los individuos para diseñar una cirugía sin ningún conocimiento de programación. Sin embargo, para que los usuarios obtener el máximo provecho de la stereotax motorizado, sería beneficioso tener conocimiento de programación matemática y G-Codificación de Mercancías (sencilla progembistiendo por CNC mecanizado). La velocidad recomendada de la fresa es mayor que 40.000 rpm. La resolución de motor paso a paso es de 1,8 ° / Paso, dirigido a 0.346 ° / Paso. A stereotax estándar tiene una resolución de 2,88 micras / paso. La velocidad de corte máxima recomendada es de 500 m / seg. La velocidad de avance lento recomendada máxima es de 3500 m / seg. El tamaño máximo recomendado broca es HP 2.

El resultado final de la cirugía diseñada en los métodos será un cráneo de rata con una craneotomía borde afilado, y 3 agujeros del cráneo (Figura 17). Tenga en cuenta que el cráneo utilizado para demostrar la cirugía fue mucho más amplio que el cráneo de rata prototípico. La craneotomía borde afilado se puede usar para insertar una matriz de microhilo en el cerebro, para las grabaciones de alta densidad neuronales. El stereotax CNC también se puede usar para reducir la matriz con gran pre...

Watch this Scientific Journal Video about Building An Open-source Robotic Stereotaxic Instrument at JoVE.com

Building An Open-source Robotic Stereotaxic Instrument

Este protocolo incluye los diseños y el software necesario para actualizar un instrumento estereotáxico existente a un robot (numérico controlado por ordenador; CNC) instrumento estereotáxico para alrededor de 1.000 dólares (sin incluir un taladro).

La construcción de un código abierto robótica estereotáxica Instrumento

This protocol includes the designs and software necessary to upgrade an existing stereotaxic instrument to a robotic (CNC) stereotaxic instrument for ...

building-an-open-source-robotic-stereotaxic-instrument

Research

JoVE Journal

Neuroscience

14.7K vistas.  Rutgers, The State University of New Jersey.  Este protocolo incluye los diseños y el software necesario para actualizar un instrumento estereotáxico existente a un robot (numérico controlado por ordenador; CNC) instrumento estereotáxico para alrededor de 1.000 dólares (sin incluir un taladro). 

Video: Building An Open-source Robotic Stereotaxic Instrument

Rodent Stereotaxic Surgery and Animal Welfare Outcome Improvements for Behavioral Neuroscience

Stereotaxic surgery for the implantation of cannulae into specific brain regions has for many decades been a very successful experimental technique to investigate the effects of locally manipulated neurotransmitter and signaling pathways in awake, behaving animals. Moreover, the stereotaxic implantation of electrodes for electrophysiological stimulation and recording studies has been instrumental to our current understanding of neuroplasticity and brain networks in behaving animals. Ever-increasing knowledge about optimizing surgical techniques in rodents1-4, public awareness concerning animal welfare issues and stringent legislation (e.g., the 2010 European Union Directive on the use of laboratory animals5) prompted us to refine these surgical procedures, particularly with respect to implementing new procedures for oxygen supplementation and the continuous monitoring of blood oxygenation and heart rate levels during the surgery as well as introducing a standardized protocol for post-surgical care. Our observations indicate that these modifications resulted in an increased survival rate and an improvement in the general condition of the animals after surgery (e.g. less weight loss and a more active animal). This video presentation will show the general procedures involved in this type of stereotaxic surgery with special attention to our several modifications. We will illustrate these surgical procedures in rats, but it is also possible to perform this type of surgery in mice or other small laboratory animals by using special adaptors for the stereotaxic apparatus6.

Stereotaxic surgery on rodents allows for targeted administration of drugs or electrical stimulation and recordings in awake, behaving animals. In this video presentation we will demonstrate recent procedural refinements to this long-standing procedure that successfully improved survival rate and reduced post-surgical weight loss.

Cirugía estereotáctica de roedores y mejoras del bienestar de los animales de resultados para la revista Behavioral Neuroscience

Stereotaxic surgery for the implantation of cannulae into specific brain regions has for many decades been a very successful experimental technique to ...

Investigación

Neurociencias

Stereotaxic Surgery for Excitotoxic Lesion of Specific Brain Areas in the Adult Rat

Many behavioral functions in mammals, including rodents and humans, are mediated principally by discrete brain regions. A common method for discerning the function of various brain regions for behavior or other experimental outcomes is to implement a localized ablation of function. In humans, patient populations with localized brain lesions are often studied for deficits, in hopes of revealing the underlying function of the damaged area. In rodents, one can experimentally induce lesions of specific brain regions.
Lesion can be accomplished in several ways. Electrolytic lesions can cause localized damage but will damage a variety of cell types as well as traversing fibers from other brain regions that happen to be near the lesion site. Inducible genetic techniques using cell-type specific promoters may also enable site-specific targeting. These techniques are complex and not always practical depending on the target brain area. Excitotoxic lesion using stereotaxic surgery, by contrast, is one of the most reliable and practical methods of lesioning excitatory neurons without damaging local glial cells or traversing fibers.
Here, we present a protocol for stereotaxic infusion of the excitotoxin, N-methyl-D-aspartate (NMDA), into the basolateral amygdala complex. Using anatomical indications, we apply stereotaxic coordinates to determine the location of our target brain region and lower an injection needle in place just above the target. We then infuse our excitotoxin into the brain, resulting in excitotoxic death of nearby neurons. While our experimental subject of choice is a rat, the same methods can be applied to other mammals, with the appropriate adjustments in equipment and coordinates.
This method can be used on a variety of brain regions, including the basolateral amygdala1-6, other amygdala nuclei6, 7, hippocampus8, entorhinal cortex9 and prefrontal cortex10. It can also be used to infuse biological compounds such as viral vectors1, 11. The basic stereotaxic technique could also be adapted for implantation of more permanent osmotic pumps, allowing more prolonged exposure to a compound of interest.

Targeted ablation of specific brain region(s) by infusion of an excitotoxin using stereotaxic coordinates is described. This technique could also be adapted for infusion of other chemicals into the rat brain.

La cirugía estereotáxica para la lesión excitotóxica de áreas específicas del cerebro en la rata adulta

Many behavioral functions in mammals, including rodents and humans, are mediated principally by discrete brain regions. A common method for discerning the ...

Stereotaxic Infusion of Oligomeric Amyloid-beta into the Mouse Hippocampus

Alzheimer&#8217;s disease is a neurodegenerative disease affecting the aging population. A key neuropathological feature of the disease is the over-production of amyloid-beta and the deposition of amyloid-beta plaques in brain regions of the afflicted individuals. Throughout the years scientists have generated numerous Alzheimer&#8217;s disease mouse models that attempt to replicate the amyloid-beta pathology. Unfortunately, the mouse models only selectively mimic the disease features. Neuronal death, a prominent effect in the brains of Alzheimer&#8217;s disease patients, is noticeably lacking in these mice. Hence, we and others have employed a method of directly infusing soluble oligomeric species of amyloid-beta - forms of amyloid-beta that have been proven to be most toxic to neurons - stereotaxically into the brain. In this report we utilize male C57BL/6J mice to document this surgical technique of increasing amyloid-beta levels in a select brain region. The infusion target is the dentate gyrus of the hippocampus because this brain structure, along with the basal forebrain that is connected by the cholinergic circuit, represents one of the areas of degeneration in the disease. The results of elevating amyloid-beta in the dentate gyrus via stereotaxic infusion reveal increases in neuron loss in the dentate gyrus within 1 week, while there is a concomitant increase in cell death and cholinergic neuron loss in the vertical limb of the diagonal band of Broca of the basal forebrain. These effects are observed up to 2 weeks. Our data suggests that the current amyloid-beta infusion model provides an alternative mouse model to address region specific neuron death in a short-term basis. The advantage of this model is that amyloid-beta can be elevated in a spatial and temporal manner.

Here, we present a protocol for direct stereotaxic brain infusion of amyloid-beta. This methodology provides an alternative in vivo mouse model to address the short-term effects of amyloid-beta on brain neurons.

Infusión estereotáxica de Oligomeric beta-amiloide en el hipocampo del ratón

Alzheimer&#8217;s disease is a neurodegenerative disease affecting the aging population. A key neuropathological feature of the disease is the ...

Non-restraining EEG Radiotelemetry: Epidural and Deep Intracerebral Stereotaxic EEG Electrode Placement

Implantable EEG radiotelemetry is of central relevance in the neurological characterization of transgenic mouse models of neuropsychiatric and neurodegenerative diseases as well as epilepsies. This powerful technique does not only provide valuable insights into the underlying pathophysiological mechanisms, i.e., the etiopathogenesis of CNS related diseases, it also facilitates the development of new translational, i.e., therapeutic approaches. Whereas competing techniques that make use of recorder systems used in jackets or tethered systems suffer from their unphysiological restraining to semi-restraining character, radiotelemetric EEG recordings overcome these disadvantages. Technically, implantable EEG radiotelemetry allows for precise and highly sensitive measurement of epidural and deep, intracerebral EEGs under various physiological and pathophysiological conditions. First, we present a detailed protocol of a straight forward, successful, quick and efficient technique for epidural (surface) EEG recordings resulting in high-quality electrocorticograms. Second, we demonstrate how to implant deep, intracerebral EEG electrodes, e.g., in the hippocampus (electrohippocampogram). For both approaches, a computerized 3D stereotaxic electrode implantation system is used. The radiofrequency transmitter itself is implanted into a subcutaneous pouch in both mice and rats. Special attention also has to be paid to pre-, peri- and postoperative treatment of the experimental animals. Preoperative preparation of mice and rats, suitable anesthesia as well as postoperative treatment and pain management are described in detail.

Non-restraining EEG radiotelemetry is a valuable methodological approach to record in vivo long-term electroencephalograms from freely moving rodents. This detailed protocol describes stereotaxic epidural and deep intracerebral electrode placement in different brain regions in order to obtain reliable recordings of CNS rhythmicity and CNS related behavioral stages.

No restricción EEG radiotelemetría: epidural y Deep intracerebral estereotáxica colocación de electrodos EEG

Implantable EEG radiotelemetry is of central relevance in the neurological characterization of transgenic mouse models of neuropsychiatric and ...

High-density Electroencephalographic Acquisition in a Rodent Model Using Low-cost and Open-source Resources

Advanced electroencephalographic analysis techniques requiring high spatial resolution, including electrical source imaging and measures of network connectivity, are applicable to an expanding variety of questions in neuroscience. Performing these kinds of analyses in a rodent model requires higher electrode density than traditional screw electrodes can accomplish. While higher-density electroencephalographic montages for rodents exist, they are of limited availability to most researchers, are not robust enough for repeated experiments over an extended period of time, or are limited to use in anesthetized rodents.1-3 A proposed low-cost method for constructing a durable, high-count, transcranial electrode array, consisting of bilaterally implantable headpieces is investigated as a means to perform advanced electroencephalogram analyses in mice or rats.

Procedures for headpiece fabrication and surgical implantation necessary to produce high signal to noise, low-impedance electroencephalographic and electromyographic signals are presented. While the methodology is useful in both rats and mice, this manuscript focuses on the more challenging implementation for the smaller mouse skull. Freely moving mice are only tethered to cables via a common adapter during recording. One version of this electrode system that includes 26 electroencephalographic channels and 4 electromyographic channels is described below.

Instructions for the low-cost construction and surgical implantation of a chronic transcranial high-density electroencephalographic montage into mice are provided. Signal recording, extraction, and processing techniques are also described.

De alta densidad Electroencefalográfica Adquisición en un modelo de roedor Uso de recursos de bajo costo y de código abierto

Advanced electroencephalographic analysis techniques requiring high spatial resolution, including electrical source imaging and measures of network ...

An Alternative Approach to Study Primary Events in Neurodegeneration Using Ex Vivo Rat Brain Slices

Despite&#160;numerous studies that attempt to develop reliable animal models&#160;which reflecting the primary processes underlying neurodegeneration, very few have been widely accepted. Here, we propose&#160;a new procedure&#160;adapted from the well-known ex vivo brain slice technique, which offers a closer in vivo-like scenario than in vitro preparations, for investigating the early events triggering cell degeneration, as observed in Alzheimer&#39;s disease (AD). This variation consists of simple and easily reproducible steps, which enable preservation of the anatomical cytoarchitecture of the selected brain region and its local functionality in a physiological milieu. Different anatomical areas can be obtained from the same brain, providing the opportunity to perform multiple experiments with the treatments in question in a site-, dose-, and time-dependent manner. Potential limitations&#160;which could affect the outcomes related to this methodology are related to the conservation of the tissue, i.e., the maintenance of its anatomical integrity during the slicing and incubation steps and the section thickness, which can influence the biochemical and immunohistochemical analysis. This approach can be employed for different purposes, such as exploring molecular mechanisms involved in physiological or pathological conditions, drug screening, or dose-response assays. Finally, this protocol could also reduce the number of animals employed in behavioral studies. The application reported here has been recently described and tested for the first time on ex vivo rat brain slices containing the basal forebrain (BF), which is one of the cerebral regions primarily affected in AD. Specifically, it has been demonstrated that the administration of a toxic peptide derived from the C-terminus of acetylcholinesterase (AChE) could prompt an AD-like profile, triggering, along the antero-posterior axis of the BF, a differential expression of proteins altered in AD, such as the alpha7 nicotinic receptor (&#945;7-nAChR), phosphorylated Tau (p-Tau), and amyloid beta (A&#946;).

An Alternative Approach to Study Primary Events in Neurodegeneration Using Ex Vivo Rat Brain Slices

We present a method which can&#160;provide further insights into the early events underlying neurodegeneration and based on the established ex-vivo brain technique, combining the advantages of in vivo and in vitro experiments. Moreover, it represents a unique opportunity for direct comparison of treated and untreated group in the same anatomical plane.

Un enfoque alternativo para estudiar acontecimientos primarios en neurodegeneración con rodajas de cerebro de rata Ex Vivo

Despite&#160;numerous studies that attempt to develop reliable animal models&#160;which reflecting the primary processes underlying neurodegeneration, ...

Stereotaxic Surgery for Implantation of Microelectrode Arrays in the Common Marmoset (Callithrix jacchus)

Marmosets (Callithrix jacchus) are small non-human primates that are gaining popularity in biomedical and preclinical research, including the neurosciences. Phylogenetically, these animals are much closer to humans than rodents. They also display complex behaviors, including a wide range of vocalizations and social interactions. Here, an effective stereotaxic neurosurgical procedure for implantation of recording electrode arrays in the common marmoset is described. This protocol also details the pre- and postoperative steps of animal care that are required to successfully perform such a surgery. Finally, this protocol shows an example of local field potential and spike activity recordings in a freely behaving marmoset 1 week after the surgical procedure. Overall, this method provides an opportunity to study the brain function in awake and freely behaving marmosets. The same protocol can be readily used by researchers working with other small primates. In addition, it can be easily modified to allow other studies requiring implants, such as stimulating electrodes, microinjections, implantation of optrodes or guide cannulas, or ablation of discrete tissue regions.

Stereotaxic Surgery for Implantation of Microelectrode Arrays in the Common Marmoset Callithrix jacchus

This work presents a protocol to perform a stereotaxic, neurosurgical implantation of&#160;microelectrode arrays in the common marmoset. This method specifically enables electrophysiological recordings in freely behaving animals but can be easily adapted to any other similar neurosurgical intervention in this species (e.g., cannula for drug administration or electrodes for brain stimulation).

Cirugía estereotaxia para la implantación de matrices de microelectrodos en el Marmoset común (Jacchus Callithrix)

Marmosets (Callithrix jacchus) are small non-human primates that are gaining popularity in biomedical and preclinical research, including the ...

An Open-Source Virtual Reality System for the Measurement of Spatial Learning in Head-Restrained Mice

Head-restrained behavioral experiments in mice allow neuroscientists to observe neural circuit activity with high-resolution electrophysiological and optical imaging tools while delivering precise sensory stimuli to a behaving animal. Recently, human and rodent studies using virtual reality (VR) environments have shown VR to be an important tool for uncovering the neural mechanisms underlying spatial learning in the hippocampus and cortex, due to the extremely precise control over parameters such as spatial and contextual cues. Setting up virtual environments for rodent spatial behaviors can, however, be costly and require an extensive background in engineering and computer programming. Here, we present a simple yet powerful system based upon inexpensive, modular, open-source hardware and software that enables researchers to study spatial learning in head-restrained mice using a VR environment. This system uses coupled microcontrollers to measure locomotion and deliver behavioral stimuli while head-restrained mice run on a wheel in concert with a virtual linear track environment rendered by a graphical software package running on a single-board computer. The emphasis on distributed processing allows researchers to design flexible, modular systems to elicit and measure complex spatial behaviors in mice in order to determine the connection between neural circuit activity and spatial learning in the mammalian brain.

Here, we present a simplified open-source hardware and software setup for investigating mouse spatial learning using virtual reality (VR). This system displays a virtual linear track to a head-restrained mouse running on a wheel by utilizing a network of microcontrollers and a single-board computer running an easy-to-use Python graphical software package.

Un sistema de realidad virtual de código abierto para la medición del aprendizaje espacial en ratones con cabeza restringida

Head-restrained behavioral experiments in mice allow neuroscientists to observe neural circuit activity with high-resolution electrophysiological and ...

Open-Source Real-Time Closed-Loop Electrical Threshold Tracking for Translational Pain Research

Nociceptors are a class of primary afferent neurons that signal potentially harmful noxious stimuli. An increase in nociceptor excitability occurs in acute and chronic pain conditions. This produces abnormal ongoing activity or reduced activation thresholds to noxious stimuli. Identifying the cause of this increased excitability is required for the development and validation of mechanism-based treatments. Single-neuron electrical threshold tracking can quantify nociceptor excitability. Therefore, we have developed an application to allow such measurements and demonstrate its use in humans and rodents. APTrack provides real-time data visualization and action potential identification using a temporal raster plot. Algorithms detect action potentials by threshold crossing and monitor their latency after electrical stimulation. The plugin then modulates the electrical stimulation amplitude using an up-down method to estimate the electrical threshold of the nociceptors. The software was built upon the Open Ephys system (V0.54) and coded in C++ using the JUCE framework. It runs on Windows, Linux, and Mac operating systems. The open-source code is available (<a data-saferedirecturl="https://www.google.com/url?q=https://github.com/Microneurography/APTrack&amp;source=gmail&amp;ust=1680789829274000&amp;usg=AOvVaw0nMk9J_EI2jUn8SlmC5JJm" href="https://github.com/Microneurography/APTrack" target="_blank">https://github.com/Microneurography/APTrack</a>). The electrophysiological recordings were taken from nociceptors in both a mouse skin-nerve preparation using the teased fiber method in the saphenous nerve and in healthy human volunteers using microneurography in the superficial peroneal nerve. Nociceptors were classified by their response to thermal and mechanical stimuli, as well as by monitoring the activity-dependent slowing of the conduction velocity. The software facilitated the experiment by simplifying the action potential identification through the temporal raster plot. We demonstrate real-time closed-loop electrical threshold tracking of single-neuron action potentials during&#160;in vivo&#160;human microneurography, for the first time, and during&#160;ex vivo&#160;mouse electrophysiological recordings of C-fibers and A&#948;-fibers. We establish proof of principle by showing that the electrical threshold of a human heat-sensitive C-fiber nociceptor is reduced by heating the receptive field. This plugin enables the electrical threshold tracking of single-neuron action potentials and allows the quantification of changes in nociceptor excitability.

APTrack is a software plugin developed for the Open Ephys platform that enables real-time data visualization and the closed-loop electrical threshold tracking of neuronal action potentials. We have successfully used this in microneurography for human C-fiber nociceptors and mouse C-fiber and A&#948;-fiber nociceptors.

Seguimiento de umbral eléctrico de circuito cerrado en tiempo real de código abierto para la investigación traslacional del dolor

Nociceptors are a class of primary afferent neurons that signal potentially harmful noxious stimuli. An increase in nociceptor excitability occurs in ...