Damos las gracias a Ye Wang para el debate y Pojoga Sorin para la formación del comportamiento. Apoyados por el Programa EUREKA NIH, el Instituto Nacional del Ojo, el Programa de Becas Pew, la Fundación James S. McDonnell (VD), y un NIH Visión Entrenamiento Grant (BJH).

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No hay conflictos de interés declarado.

De unidades múltiples grabaciones se han convertido en estándar para el análisis de cómo las redes neuronales en la corteza codificar la información de estímulo. Teniendo en cuenta los recientes avances en tecnología de electrodos, la aplicación de los electrodos laminar permite una caracterización sin precedentes de los locales de los circuitos corticales. A pesar de electrodos múltiples grabaciones ofrecen información útil sobre la dinámica de la población neuronal, múltiples electrodos laminares permit...

<table border="1"><tbody><tr><td> Nombre del Equipo </td><td> Empresa </td><td> Número de catálogo </td><td> Comentarios </td></tr><tr><td> Nan microdrive sistema </td><td> Nan los instrumentos </td><td> NAN-S4 </td><td> Figura 2. Abrazaderas personalizados son necesarios para utilizar el U-Probe. Todo lo mencionado con excepción de la U-Probe es proporcionado por los instrumentos NAN. </td></tr><tr><td> Microdrives tornillo </td><td> MIT taller mecánico </td><td></td><td> Todo lo que es capaz de asegurar un tubo de guía a la red NAN debe ser apropiado. </td></tr><tr><td> Tubos de acero inoxidable Guía </td><td> Partes pequeñas </td><td> B00137QHNS (1) o B00137QHO2 (5) </td><td> Estas son 60 de largo y cortado a la medida en el laboratorio usando un taladro Dremel </td></tr><tr><td> Plexon U-Probe </td><td> Plexon, Inc </td><td> PLX-UP-16-25ED-100-SE-360-25T-500 </td><td> Ver la sonda U-especificaciones disponibles en <a href="www.plexon.com">www.plexon.com</a> También vea la Figura 1. </td></tr></tbody></table> Tabla 1. Hardware. <table border="1"><tbody><tr><td> Nombre del Software </td><td> Empresa </td><td> Sitio web </td><td> Comentarios </td></tr><tr><td> NAN software </td><td> NAN </td><td> <a href="http://www.naninstruments.com/DesignConcept.htm">http://www.naninstruments.com/DesignConcept.htm</a> </td><td> Interfaz de la computadora requiere un puerto serial adicional para acomodar el sistema y el hardware Plexon NAN </td></tr><tr><td> Clasificador en línea, FPAlign, PlexUtil, programas de MATLAB </td><td> Plexon </td><td> <a href="http://www.plexon.com/downloads.html#Software">http://www.plexon.com/downloads.html # Software</a> </td><td> En &#39;paquetes de instalación &quot; </td></tr><tr><td> NeuroExplorer </td><td> NeuroExplorer </td><td> <a href="http://www.neuroexplorer.com/">http://www.neuroexplorer.com/</a> </td><td> Bajo &quot;Recursos&quot; </td></tr><tr><td> CSDplotter Versión 0.1.1 </td><td> Klas H. Petterson </td><td> <a href="http://arken.umb.no/~klaspe/user_guide.pdf">http://arken.umb.no/ ~ klaspe / user_guide.pdf</a> </td><td></td></tr></tbody></table> Tabla 2. Software.

examining local network processing using multi-contact laminar electrode recording

Capas corticales son estructuras omnipresentes a lo largo de la neocorteza 1-4 que consisten en redes locales muy recurrente. En los últimos años, importantes avances se han realizado en nuestra comprensión de las diferencias en las propiedades de respuesta de las neuronas en diferentes capas corticales 5-8, pero aún queda mucho que aprender acerca de si y cómo la población neuronal codificar la información en un determinado laminar manera. Existentes de múltiples electrodos técnicas de matriz, aunque informativo para la medición de las respuestas a través de muchos milímetros de espacio cortical a lo largo de la superficie cortical, no son adecuadas para abordar el tema de los circuitos corticales laminar. Aquí, presentamos nuestro método para la creación y grabación de las neuronas individuales y los potenciales de campo locales (LFPs) a través de las capas corticales de la corteza visual primaria (V1) que utiliza electrodos de contacto multi-laminar (Figura 1; Plextrode U-Probe, Inc Plexon). Los métodos incluyen la construcción del dispositivo de grabación, la identificación de las capas corticales, y la identificación de los campos receptivos de las neuronas individuales. Para identificar las capas corticales, se mide el potencial de respuesta evocada (ERP) de la LFP de series de tiempo con todo el campo de estímulos brilló. A continuación, realice fuente de corriente densidad (CSD) un análisis para identificar la inversión de polaridad acompañado por la configuración del fregadero de código en la base de la capa 4 (el lavabo está dentro de la capa 4, posteriormente conocida como la capa granular 9-12). Fuente de corriente de densidad es útil porque proporciona un índice de la ubicación, la dirección y la densidad del flujo de corriente transmembrana, lo que nos permite colocar con precisión los electrodos de registro de todas las capas en una sola penetración 6, 11, 12.

Watch this Scientific Journal Video about Examining Local Network Processing using Multi-contact Laminar Electrode Recording at JoVE.com

Examining Local Network Processing using Multi-contact Laminar Electrode Recording

Una cuestión fundamental en nuestra comprensión de los circuitos corticales es cómo las redes en diferentes capas corticales codificar la información sensorial. Aquí se describe la utilización de técnicas electrofisiológicas de múltiples electrodos de contacto laminar para grabar unidades únicas y potenciales locales de campo y análisis de la actualidad para identificar a las capas corticales.

El examen de procesamiento de la red local usando multi-contacto laminar de grabación de electrodos

Cortical layers are ubiquitous structures throughout neocortex1-4 that consist of highly recurrent local networks. In recent years, significant progress ...

examining-local-network-processing-using-multi-contact-laminar

Research

JoVE Journal

Neuroscience

12.7K vistas.  University of Texas. Una cuestión fundamental en nuestra comprensión de los circuitos corticales es cómo las redes en diferentes capas corticales codificar la información sensorial. Aquí se describe la utilización de técnicas electrofisiológicas de múltiples electrodos de contacto laminar para grabar unidades únicas y potenciales locales de campo y análisis de la actualidad para identificar a las capas corticales.

Video: Examining Local Network Processing using Multi-contact Laminar Electrode Recording

Electrophysiology on Isolated Brainstem-spinal Cord Preparations from Newborn Rodents Allows Neural Respiratory Network Output Recording

While it is well known that the central respiratory drive is located in the brainstem, several aspects of its basic function, development, and response to stimuli remain to be fully understood. To overcome the difficulty of accessing the brainstem in the whole animal, isolation of the brainstem and part of the spinal cord is performed. This preparation is maintained in artificial cerebro-spinal fluid where gases, concentrations, and temperature are controlled and monitored. The output signal from the respiratory network is recorded by a suction electrode placed on the fourth ventral root. In this manner, stimuli can be directly applied onto the brainstem, and the effect can be recorded directly. The signal recorded is linked to the inspiratory signal sent to the diaphragm via the phrenic nerve, and can be described as bursts (around 8 bursts per minute). Analysis of these bursts (frequency, amplitude, length, and area under the curve) allows precise characterization of the stimulus effect on the respiratory network. The main limitation of this method is the viability of the preparation beyond the early post-natal stages. Thus, this method greatly focuses on the study of the whole network without the peripheral inputs in the newborn rat.

The central respiratory drive is located in the brainstem. Spontaneous respiratory motor output from an isolated brainstem-spinal cord is recorded by placing an electrode on the fourth ventral root. This experimental approach is valuable for pharmacological investigations or the assessment of respiratory challenges and genetic manipulations on rhythmic motor behavior.

Electrofisiología en aislados Preparativos Cord-tronco cerebral espinal de roedores recién nacidos permite la grabación de salida de red neuronal Respiratorio

While it is well known that the central respiratory drive is located in the brainstem, several aspects of its basic function, development, and response to ...

Investigación

Neurociencias

Time-dependent Increase in the Network Response to the Stimulation of Neuronal Cell Cultures on Micro-electrode Arrays

Micro-electrode arrays (MEAs) can be used to investigate drug toxicity, design paradigms for next-generation personalized medicine, and study network dynamics in neuronal cultures. In contrast with more traditional methods, such as patch-clamping, which can only record activity from a single cell, MEAs can record simultaneously from multiple sites in a network, without requiring the arduous task of placing each electrode individually. Moreover, numerous control and stimulation configurations can be easily applied within the same experimental setup, allowing for a broad range of dynamics to be explored. One of the key dynamics of interest in these in vitro studies has been the extent to which cultured networks display properties indicative of learning. Mouse neuronal cells cultured on MEAs display an increase in response following training induced by electrical stimulation. This protocol demonstrates how to culture neuronal cells on MEAs; successfully record from over 95% of the plated dishes; establish a protocol to train the networks to respond to patterns of stimulation; and sort, plot, and interpret the results from such experiments. The use of a proprietary system for stimulating and recording neuronal cultures is demonstrated. Software packages are also used to sort neuronal units. A custom-designed graphical user interface is used to visualize post-stimulus time histograms, inter-burst intervals, and burst duration, as well as to compare the cellular response to stimulation before and after a training protocol. Finally, representative results and future directions of this research effort are discussed.

Mouse neuronal cells cultured on multi-electrode arrays display an increase in response following electrical stimulation. This protocol demonstrates how to culture neurons, how to record activity, and how to establish a protocol to train the networks to respond to patterns of stimulation.

Incremento dependiente del tiempo en la respuesta de la red a la estimulación de cultivos de células neuronales en matrices de microelectrodos

Micro-electrode arrays (MEAs) can be used to investigate drug toxicity, design paradigms for next-generation personalized medicine, and study network ...

Concurrent Recording of Co-localized Electroencephalography and Local Field Potential in Rodent

Although electroencephalography (EEG) is widely used as a non-invasive technique for recording neural activities of the brain, our understanding of the neurogenesis of EEG is still very limited. Local field potentials (LFPs) recorded via a multi-laminar microelectrode can provide a more detailed account of simultaneous neural activity across different cortical layers in the neocortex, but the technique is invasive. Combining EEG and LFP measurements in a pre-clinical model can greatly enhance understanding of the neural mechanisms involved in the generation of EEG signals, and facilitate the derivation of a more realistic and biologically accurate mathematical model of EEG. A simple procedure for acquiring concurrent and co-localized EEG and multi-laminar LFP signals in the anesthetized rodent is presented here. We also investigated whether EEG signals were significantly affected by a burr hole drilled in the skull for the insertion of a microelectrode. Our results suggest that the burr hole has a negligible impact on EEG recordings.

This protocol describes a simple method for concurrent recording of co-localized electroencephalography (EEG) and multi-laminar local field potential in an anesthetized rat. A burr hole drilled in the skull for the insertion of a microelectrode is shown to produce negligible distortion of the EEG signal.

Grabación concurrente de electroencefalografía Co localizada y potencial de campo Local en roedores

Although electroencephalography (EEG) is widely used as a non-invasive technique for recording neural activities of the brain, our understanding of the ...

Examining Monosynaptic Connections in Drosophila Using Tetrodotoxin Resistant Sodium Channels

Here, a new technique termed Tetrotoxin (TTX) Engineered Resistance for Probing Synapses (TERPS) is applied to test for monosynaptic connections between target neurons. The method relies on co-expression of a transgenic activator with the tetrodotoxin-resistant sodium channel, NaChBac, in a specific presynaptic neuron. Connections with putative post-synaptic partners are determined by whole-cell recordings in the presence of TTX, which blocks electrical activity in neurons that do not express NaChBac. This approach can be modified to work with any activator or calcium imaging as a reporter of connections. TERPS adds to the growing set of tools available for determining connectivity within networks. However, TERPS is unique in that it also reliably reports bulk or volume transmission and spillover transmission.

Examining Monosynaptic Connections in Drosophila Using Tetrodotoxin Resistant Sodium Channels

This article features a method to test the monosynaptic connections between neurons by employing tetrodotoxin and the tetrodotoxin-resistant sodium channel, NaChBac.

Examinar las conexiones Monosynaptic en Drosophila utilizando canales de sodio Tetrodotoxin resistentes

Here, a new technique termed Tetrotoxin (TTX) Engineered Resistance for Probing Synapses (TERPS) is applied to test for monosynaptic connections between ...

Evaluation of Hemisphere Lateralization with Bilateral Local Field Potential Recording in Secondary Motor Cortex of Mice

This article demonstrates complete, detailed procedures for both in vivo bilateral recording and analysis of local field potential (LFP) in the cortical areas of mice, which are useful for evaluating possible laterality deficits, as well as for assessing brain connectivity and coupling of neural network activities in rodents. The pathological mechanisms underlying Alzheimer&#39;s disease (AD), a common neurodegenerative disease, remain largely unknown. Altered brain laterality has been demonstrated in aging people, but whether or not abnormal lateralization is one of the early signs of AD has not been determined. To investigate this, we recorded bilateral LFPs in 3-5-month-old AD model mice, APP/PS1, together with littermate wild type (WT) controls. The LFPs of the left and right secondary motor cortex (M2), specifically in the gamma band, were more synchronized in APP/PS1 mice than in WT controls, suggesting a declined hemispheric asymmetry of bilateral M2 in this AD mouse model. Notably, the recording and data analysis processes are flexible and easy to carry out, and can also be applied to other brain pathways when conducting experiments that focus on neuronal circuits.

We present in vivo electrophysiological recording of the local field potential (LFP) in bilateral secondary motor cortex (M2) of mice, which can be applied to evaluate hemisphere lateralization. The study revealed altered levels of synchronization between the left and right M2 in APP/PS1 mice compared to WT controls.

Evaluación de la lateralización del hemisferio con registro de potencial de campo local bilateral en corteza motora secundaria de ratones

This article demonstrates complete, detailed procedures for both in vivo bilateral recording and analysis of local field potential (LFP) in the cortical ...

Recording Network Activity in Spinal Nociceptive Circuits Using Microelectrode Arrays

The roles and connectivity of specific types of neurons within the spinal cord dorsal horn (DH) are being delineated at a rapid rate to provide an increasingly detailed view of the circuits underpinning spinal pain processing. However, the effects of these connections for broader network activity in the DH remain less well understood because most studies focus on the activity of single neurons and small microcircuits. Alternatively, the use of microelectrode arrays (MEAs), which can monitor electrical activity across many cells, provides high spatial and temporal resolution of neural activity. Here, the use of MEAs with mouse spinal cord slices to study DH activity induced by chemically stimulating DH circuits with 4-aminopyridine (4-AP) is described. The resulting rhythmic activity is restricted to the superficial DH, stable over time, blocked by tetrodotoxin, and can be investigated in different slice orientations. Together, this preparation provides a platform to investigate DH circuit activity in tissue from na&#239;ve animals, animal models of chronic pain, and mice with genetically altered nociceptive function. Furthermore, MEA recordings in 4-AP-stimulated spinal cord slices can be used as a rapid screening tool to assess the capacity of novel antinociceptive compounds to disrupt activity in the spinal cord DH.

The combined use of microelectrode array technology and 4-aminopyridine-induced chemical stimulation for investigating network-level nociceptive activity in the spinal cord dorsal horn is outlined.

Registro de la actividad de la red en circuitos nociceptivos espinales utilizando matrices de microelectrodos

The roles and connectivity of specific types of neurons within the spinal cord dorsal horn (DH) are being delineated at a rapid rate to provide an ...

Visualización de redes de microtúbulos en uniones neuromusculares y células musculares de Drosophila

Using Drosophila Larval Neuromuscular Junction and Muscle Cells to Visualize Microtubule Network

The microtubule network is an essential component of the nervous system. Mutations in many microtubules regulatory proteins are associated with neurodevelopmental disorders and neurological diseases, such as microtubule-associated protein Tau to neurodegenerative diseases, microtubule severing protein Spastin and Katanin 60 cause hereditary spastic paraplegia and neurodevelopmental abnormalities, respectively. Detection of microtubule networks in neurons is advantageous for elucidating the pathogenesis of neurological disorders. However, the small size of neurons and the dense arrangement of axonal microtubule bundles make visualizing the microtubule networks challenging. In this study, we describe a method for dissection of the larval neuromuscular junction and muscle cells, as well as immunostaining of &#945;-tubulin and microtubule-associated protein Futsch to visualize microtubule networks in Drosophila melanogaster. The neuromuscular junction permits us to observe both pre-and post-synaptic microtubules, and the large size of muscle cells in Drosophila larva allows for clear visualization of the microtubule network. Here, by mutating and overexpressing Katanin 60 in Drosophila melanogaster,&#160;and then examining the microtubule networks in the neuromuscular junction and muscle cells, we&#160;accurately reveal the regulatory role of Katanin 60 in neurodevelopment. Therefore, combined with the powerful genetic tools of Drosophila&#160;melanogaster, this protocol greatly facilitates genetic screening and microtubule dynamics analysis for the role of microtubule network regulatory proteins in the nervous system.

Autor destacado: Comprensión de la red de microtúbulos en las uniones neuromusculares de Drosophila 

Here, we present a detailed protocol to visualize the microtubule networks in neuromuscular junctions and muscle cells. Combined with the powerful genetic tools of Drosophila melanogaster, this protocol greatly facilitates genetic screening and microtubule-dynamics analysis for the role of microtubule network regulatory proteins in the nervous system.

Uso de la unión neuromuscular de las larvas de Drosophila y células musculares para visualizar la red de microtúbulos

The microtubule network is an essential component of the nervous system. Mutations in many microtubules regulatory proteins are associated with ...

Micro-Drive System Assembly and Electrode Array Implantation for Multichannel Recording in Mice

Begin by assembling the Microdrive system. Connect two computer-designed boards using two stulls and a screw that holds the movable Microdrive, and attach the connector to one board. Ensure the Microdrive can carry two sets of eight guide tubes for each side of the motor cortex or MC region.Cut the guide tubes to the same length. Cut 16 nichrome wires, each measuring approximately five centimeters long and 35 micrometers in diameter. Load the wires into the guide tubes and apply glue to fix them.Strip the wire insulation and twine each exposed wire to each pin from the connector following the channel map and the reference and ground electrodes, then slowly coat conducting paint onto each pin. Cover the pins using epoxy resin, then perform gold plating via an impedance tester to reduce the impedance of the electrode tips to approximately 350 kiloohms. For electrode array implantation, fix the anesthetized mouse in a stereotaxic apparatus and use a temperature controller to maintain its rectal temperature at 37 degrees Celsius.Then make a small midline incision to expose the skull. Remove the residual tissue using scissors and clean the skull with sterile saline-soaked cotton buds. Using a glass microelectrode filled with ink, mark the desired locations of the bilateral MC for implantation.Next, using a skull drill, carefully drill two small holes on both the left and right sides of the coordinated skull in the MC regions. Gently remove the dura mater from the holes with fine forceps. Then insert the Microdrive system into the center of the holes, using a micromanipulator at 10 micrometers per second.After finishing the insertion, fill the petroleum jelly into the dental cement walls and join the bottom plate of the Microdrive system and the dental cement walls with the mixed dental cement.