Name: preparation of aplysia sensory-motor neuronal cell cultures
Uploaded: 2009-06-08T00:00:00
Description: Watch this Scientific Journal Video about Preparation of Aplysia Sensory-motor Neuronal Cell Cultures at JoVE.com

Le travail en laboratoire impliquant la mise en culture des neurones aplysie est financé par la liste des NIH et le NIH R01 R21MH077921.

Préparation (voir la section des solutions à la fin du protocole pour la composition des solutions) <ol><li> Préparer des boîtes de culture. Manteau en verre de plat ainsi Mattek la culture à fond de verre avec de la poly-L-lysine (made in borate de sodium). Ajouter suffisamment pour couvrir complètement le verre bien et laissez reposer&gt; 1h (peut être laissé toute la nuit). Enlever soigneusement la poly-L-lysine par un rinçage à l&#39;eau de mer artificielle (ASW) 4-5 fois. Après avoir retiré le dernier...

<ol>
	<li>Kandel, E. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+molecular+biology+of+memory+storage:+a+dialogue+between+genes+and+synapses.">The molecular biology of memory storage: a dialogue between genes and synapses.</a> Science. 294 (5544), 1030-1038 (2001).</li><li>Rayport, S. G., Schacher, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synaptic+plasticity+in+vitro:+cell+culture+of+identified+Aplysia+neurons+mediating+short-term+habituation+and+sensitization.">Synaptic plasticity in vitro: cell culture of identified Aplysia neurons mediating short-term habituation and sensitization.</a> J Neurosci. 6 (3), 759-763 (1986).</li><li>Goldberg, D. J., Schacher, S., Banker, G., Goslin, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Culturing Nerve Cells. , 213-236 (1998).</li><li>Glanzman, D. L., Kandel, E. R., Schacher, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Identified+target+motor+neuron+regulates+neurite+outgrowth+and+synapse+formation+of+aplysia+sensory+neurons+in+vitro.">Identified target motor neuron regulates neurite outgrowth and synapse formation of aplysia sensory neurons in vitro.</a> Neuron. 3 (4), 441-450 (1989).</li><li>Lyles, V., Zhao, Y., Martin, K. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synapse+formation+and+mRNA+localization+in+cultured+Aplysia+neurons.">Synapse formation and mRNA localization in cultured Aplysia neurons.</a> Neuron. 49 (3), 349-356 (2006).</li><li>Martin, K. C., Casadio, A., Zhu, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synapse-specific,+long-term+facilitation+of+aplysia+sensory+to+motor+synapses:+a+function+for+local+protein+synthesis+in+memory+storage.">Synapse-specific, long-term facilitation of aplysia sensory to motor synapses: a function for local protein synthesis in memory storage.</a> Cell. 91 (7), 927-938 (1997).</li><li>Guan, Z., Giustetto, M., Lomvardas, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Integration+of+long-term-memory-related+synaptic+plasticity+involves+bidirectional+regulation+of+gene+expression+and+chromatin+structure.">Integration of long-term-memory-related synaptic plasticity involves bidirectional regulation of gene expression and chromatin structure.</a> Cell. 111 (4), 483-493 (2002).</li></ol>

La bonne préparation des aplysie sensori-motrices des cultures a une courbe d&#39;apprentissage assez lent, car il implique le développement de la motricité fine associée à la microdissection et la manipulation des neurones individuels vue à travers un stéréomicroscope. Dans notre expérience, il faut environ 1-3 semaines de pratique pour obtenir saine isolées neurones sensoriels en culture et une semaine supplémentaire 1-3 pour apprendre à paire neurones sensoriels avec les neurones moteurs. Nous av...

Solutions nécessaires pour la culture <ol><li> 0,35 M MgCl 2, stockées à température ambiante, utilisée pour l&#39;anesthésie. </li><li> Poly-L-lysine solution, Sigma: P-1524, PM&gt; 300000, faite en sodium borate 0,1 M pH8.2 à 0.5mg/ml solution. Vortex bien et stériliser par filtration à travers un filtre de 0,22 um, le stocker à ° 4C. Ne pas de gel-dégel. </li><li> Poudre de milieu L-15 (Leibovitz) (Sigma: L4386) complété par les sels comme ci-dessous pour faire 1 litre <table border="1" cellspacing="0" cellpadding="0"><tr><td> L-15 en poudre </td><td> 13,8 g </td></tr><tr><td> NaCl </td><td> 15,4 g </td></tr><tr><td> D-glucose </td><td> 6,24 g </td></tr><tr><td> MgSO 4 7H 2 0 </td><td> 6.45g </td></tr><tr><td> KCl </td><td> 350 mg </td></tr><tr><td> NaHCO 3 </td><td> 170 mg </td></tr><tr><td> MgCl 2 6H 2 O </td><td> 5,49 g </td></tr><tr><td> CaCl 2, 2H 2 O </td><td> 1.43g </td></tr><tr><td> HEPES </td><td> 3.53g </td></tr></table> Ajouter ddH20 à 1 litre. Le pH devrait être d&#39;environ 7.4 à 7.5, ajouter 10ml de 100X Pen / Strep solution, et le filtre de stériliser à travers un filtre 0,22 m. Entreposer à 4C0 pour les plus de 1 mois. </li><li> Solution de digestion de protéase: 1% de protéase IX (1unit/mg) est faite dans L15 (complété avec des sels comme ci-dessus) ou en ASW immédiatement avant usage, stérilisée par filtration à travers un Millipore 0,22 m. 5mls devrait être suffisant pour les ganglions de deux animaux (assurez-vous que les ganglions sont complètement immergés dans une solution de protéase). Sigma a déclaré qu&#39;ils vont cesser de vendre de protéase IX. Une protéase de substitution est: dispase II (Roche Applied Science catalogue # 04942078001). </li><li> Le milieu de culture. Immédiatement avant la préparation des cultures, décongeler un aliquot de 10 ml d&#39;hémolymphe et le mélanger avec 10 ml de L15 (complété avec des sels ci-dessus) pour faire un milieu de culture 20mls. Ajouter 200 pi de 200mm L-Glutamine, bien mélanger et à utiliser pour préparer des cultures. Ce milieu doit être préparé à chaque fois les cultures sont faites. </li><li> Eau de mer artificielle peut être faite de Instant Ocean (Aquarium Systems, Mentor, OH) ou comme suit: 450 mM NaCl, 10 mM de KCl, 30 mM MgCl 2 (6H 2 O), 20 mM MgSO 4, 10 mM de CaCl 2 (2H 2 O), 10 mM HEPES, avec un pH ajusté à 7,4. </li></ol>

preparation of aplysia sensory-motor neuronal cell cultures

Le système nerveux de l&#39;aplysie californica mollusque marin est relativement simple, composé d&#39;environ 20 000 neurones. Les neurones sont de grande taille (jusqu&#39;à 1 mm de diamètre) et identifiable, avec des tailles différentes, des formes, positions et les pigmentations et les corps cellulaires sont exposées à l&#39;extérieur en cinq ganglions jumelé distribués dans tout le corps de l&#39;animal. Ces propriétés ont permis aux enquêteurs de délimiter les circuits qui sous-tendent les comportements spécifiques chez l&#39;animal 1. La connexion monosynaptique entre les neurones sensoriels et moteurs est une composante centrale du réflexe de retrait des branchies chez l&#39;animal, un simple réflexe de défense dans lequel l&#39;animal se retire de ses branchies, en réponse à une stimulation tactile du siphon. Ce réflexe subit des formes d&#39;apprentissage non-associatif et associatif, y compris la sensibilisation, l&#39;accoutumance et le conditionnement classique. Un avantage particulier à l&#39;étude de la plasticité synaptique, la synapse sensori-moteur peut être reconstituée dans la culture, où bien caractérisés stimuli suscitent des formes de plasticité qui ont des corrélats directe dans le comportement de l&#39;animal 2,3. Plus précisément, l&#39;application de la sérotonine produit un renforcement synaptique qui, selon le protocole d&#39;application, dure quelques minutes (à court terme de facilitation), heures (à moyen terme de facilitation) ou jours (facilitation à long terme). En revanche, l&#39;application de la FMRFamide émetteur peptide produit un affaiblissement synaptique ou la dépression qui, selon le protocole d&#39;application, peut durer de quelques minutes à plusieurs jours (dépression à long terme). La grande taille des neurones permet d&#39;enregistrer l&#39;électrode répétées forte de la force synaptique au cours des périodes de jours ensemble avec microinjection de vecteurs d&#39;expression, et d&#39;autres composés siRNA pour cibler des cascades de signalisation et les molécules et ainsi d&#39;identifier les étapes moléculaires et cellulaires biologiques qui sous-tendent les changements de l&#39;efficacité synaptique. Un avantage supplémentaire du système de culture de l&#39;aplysie vient du fait que les neurones de démontrer la spécificité des synapses dans la culture 4,5. Ainsi, les neurones sensoriels ne forment des synapses avec eux-mêmes (autapses) ou avec d&#39;autres neurones sensoriels, et ils ne forment des synapses avec des non-cibles identifiés dans les neurones moteurs de la culture. Les varicosités, les sites de contact synaptiques entre les neurones sensoriels et moteurs, sont assez grandes (2-7 microns de diamètre) pour permettre la formation des synapses (ainsi que des changements dans la morphologie synaptique) avec les neurones moteurs cible à être étudiés au niveau du microscope optique. Dans cette vidéo, nous démontrons chaque étape de la préparation des cultures de neurones sensori-moteurs, y compris les adultes et les anesthésier aplysie juvénile, disséquant leurs ganglions, la digestion de la protéase du ganglion, l&#39;enlèvement du tissu conjonctif par microdissection, l&#39;identification des deux neurones sensoriels et moteurs et l&#39;enlèvement de chaque type cellulaire par microdissection, le placage du neurone moteur, plus du neurone sensoriel et la manipulation des neurites sensoriels pour former le contact avec la culture des neurones moteurs.

Watch this Scientific Journal Video about Preparation of Aplysia Sensory-motor Neuronal Cell Cultures at JoVE.com

Preparation of Aplysia Sensory-motor Neuronal Cell Cultures

Des cultures primaires de<em&gt; Aplysie</em&gt; Sensori-motrices des neurones fournir une préparation modèle pour étudier la formation des synapses et la plasticité synaptique<em&gt; In vitro</em&gt;. Cette vidéo montre l&#39;identification et la microdissection des neurones sensoriels et moteurs de<em&gt; Aplysie</emGanglions&gt; ainsi que les méthodes pour établir et maintenir sensori-motrices des neurones en culture.

Préparation des Aplysie Sensori-motrice cultures de cellules neuronales

The nervous system of the marine mollusk Aplysia californica is relatively simple, consisting of approximately 20,000 neurons.  The neurons are large (up ...

preparation-of-aplysia-sensory-motor-neuronal-cell-cultures

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Biology

Preparation of Aplysia Sensory-motor Neuronal Cell Cultures

ES Cell-derived Neuroepithelial Cell Cultures

ES cells have the potential to differentiate into cells from all germ layers, which makes them an attractive tool for the development of new therapies. In general, the differentiation of ES cells follows the concept to first generate immature progenitor cells, which then can be propagated and differentiated into mature cellular phenotypes. This also applies for ES cell-derived neurogenesis, in which the development of neural cells follows two major steps: First, the derivation and expansion of immature neuroepithelial precursors and second, their differentiation into mature neural cells. A common method to produce neural progenitors from ES cells is based on embryoid body (EB) formation, which reveals the differentiation of cells from all germ layers including neuroectoderm. An alternative and more efficient method to induce neuroepithelial cell development uses stromal cell-derived inducing activity (SDIA), which can be achieved by co-culturing ES cells with skull bone marrow-derived stromal cells (1). Both, EB formation and SDIA, reveal the development of rosette-like structures, which are thought to resemble neural tube- and/or neural crest-like progenitors. The neural precursors can be isolated, expanded and further differentiated into specific neurons and glia cells using defined culture conditions. Here, we describe the generation and isolation of such rosettes in co-culture experiments with the stromal cell line MS5 (2-5).

Derivation of neuroepithelial precursors from embryonic stem (ES) cells using stromal cell-derived inducing activity (SDIA).

ES Cell dérivé des cultures cellulaires neuroépithéliales

ES cells have the potential to differentiate into cells from all germ layers, which makes them an attractive tool for the development of new therapies. In ...

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Biologie

Primary Neuronal Cultures from the Brains of Late Stage Drosophila Pupae

In this video, we demonstrate the preparation of primary neuronal cultures from the brains of late stage Drosophila pupae. The procedure begins with the removal of brains from animals at 70-78 hrs after puparium formation. The isolated brains are shown after brief incubation in papain followed by several washes in serum-free growth medium. The process of mechanical dissociation of each brain in a 5 ul drop of media on a coverslip is illustrated. The axons and dendrites of the post-mitotic neurons are sheered off near the soma during dissociation but the neurons begin to regenerate processes within a few hours of plating. Images show live cultures at 2 days. Neurons continue to elaborate processes during the first week in culture. Specific neuronal populations can be identified in culture using GAL4 lines to drive tissue specific expression of fluorescent markers such as  GFP or RFP. Whole cell recordings have demonstrated the cultured neurons form functional, spontaneously active cholinergic and GABAergic synapses. A short video segment illustrates calcium dynamics in the cultured neurons using Fura-2 as a calcium indicator dye to monitor spontaneous calcium transients and nicotine evoked calcium responses in a dish of cultured neurons. These pupal brain cultures are a useful model system in which genetic and pharmacological tools can be used to identify intrinsic and extrinsic factors that influence formation and function of central synapses.

This video demonstrates the preparation of primary neuronal cultures from the brains of late stage Drosophila pupae. Views of live cultures show neurite outgrowth and imaging of calcium levels using Fura-2.

Cultures neuronales primaires du cerveau des drosophiles stade tardif pupes

In this video, we demonstrate the preparation of primary neuronal cultures from the brains of late stage Drosophila pupae. The procedure begins with the ...

Preparation of Neuronal Cultures from Midgastrula Stage Drosophila Embryos

This video illustrates the procedure for making primary neuronal cultures from midgastrula stage Drosophila embryos. The methods for collecting embryos and their dechorionation using bleach are demonstrated. Using a glass pipet attached to a mouth suction tube, we illustrate the removal of all cells from single embryos. The method for dispersing cells from each embyro into a small (5 l) drop of medium on an uncoated glass coverslip is demonstrated. A view through the microscope at 1 hour after plating illustrates the preferred cell density. Most of the cells that survive when grown in defined medium are neuroblasts that divide one or more times in culture before extending neuritic processes by 12-24 hours. A view through the microscope illustrates the level of neurite outgrowth and branching expected in a healthy culture at 2 days in vitro. The cultures are grown in a simple bicarbonate based defined medium, in a 5% CO2 incubator at 22-24&deg;C. Neuritic processes continue to elaborate over the first week in culture and when they make contact with neurites from neighboring cells they often form functional synaptic connections. Neurons in these cultures express voltage-gated sodium, calcium, and potassium channels and are electrically excitable. This culture system is useful for studying molecular genetic and environmental factors that regulate neuronal differentiation, excitability, and synapse formation/function.

This video demonstrates the preparation of primary neuronal cultures from midgastrula stage Drosophila embryos. Views of live cultures show cells 1 hour after plating and differentiated neurons after 2 days of growth in a bicarbonate-based defined medium. The neurons are electrically excitable and form synaptic connections.

Préparation des cultures de neurones d&#39;embryons de drosophile Midgastrula scène

This video illustrates the procedure for making primary neuronal cultures from midgastrula stage Drosophila embryos. The methods for collecting embryos ...

Preparation of Dissociated Mouse Cortical Neuron Cultures

This video will guide you through the process for generating cortical neuronal cultures from late embryo and early postnatal mouse brain. These cultures can be used for a variety of applications including immunocytochemistry, biochemistry, electrophysiology, calcium and sodium imaging, protein and/or RNA isolation. These cultures also provide a platform to study the neuronal development of transgenic animals that carry a late embryonic or postnatal lethal gene mutation. The procedure is relatively straight forward, requires some experience in tissue culture technique and should not take longer than two to three hours if you are properly prepared. Careful separation of the cortical rind from the thalamo-cortical fiber tract will reduce the number of unwanted non-neuronal cells. To increase yields of neuronal cells triturate the pieces of the cortical tissue gently after the enzyme incubation step. This is imperative as it prevents unnecessary injury to cells and premature neuronal cell death. Since these cultures are maintained in the absence of glia feeder cells, they also offer an added advantage of growing cultures enriched in neurons.

This video shows a procedure for generating neuronal cultures from late embryo and early postnatal mouse cortex. These cultures can be used for immunocytochemistry, biochemistry, electrophysiology, calcium and sodium imaging and provide a platform to study the neuronal development of transgenic animals that carry a postnatal lethal gene mutation.

Préparation de cultures souris Dissocié neurones corticaux

This video will guide you through the process for generating cortical neuronal cultures from late embryo and early postnatal mouse brain. These cultures ...

Neuronal Cell Cultures from Aplysia for High-Resolution Imaging of Growth Cones

Neuronal growth cones are the highly motile structures at the tip of axons that can detect guidance cues in the environment and transduce this information into directional movement towards the appropriate target cell. To fully understand how guidance information is transmitted from the cell surface to the underlying dynamic cytoskeletal networks, one needs a model system suitable for live cell imaging of protein dynamics at high temporal and spatial resolution. Typical vertebrate growth cones are too small to quantitatively analyze F-actin and microtubule dynamics. Neurons from the sea hare Aplysia californica are 5-10 times larger than vertebrate neurons, can easily be kept at room temperature and are very robust cells for micromanipulation and biophysical measurements. Their growth cones have very defined cytoplasmic regions and a well-described cytoskeletal system. The neuronal cell bodies can be microinjected with a variety of probes for studying growth cone motility and guidance. In the present protocol we demonstrate a procedure for dissection of the abdominal ganglion, culture of bag cell neurons and setting up an imaging chamber for live cell imaging of growth cones.

Aplysia californica neurons develop large growth cones in culture that are excellent for high-resolution imaging of growth cone motility and guidance. Here, we present a protocol for dissection and plating of Aplysia bag cell neurons as well as for setting up a chamber for live cell imaging.

Cultures de cellules neuronales de l&#39;aplysie pour l&#39;imagerie haute résolution des cônes de croissance

Neuronal growth cones are the highly motile structures at the tip of axons that can detect guidance cues in the environment and transduce this information ...

Mouse Epidermal Neural Crest Stem Cell (EPI-NCSC) Cultures

EPI-NCSC are remnants of the embryonic neural crest in an adult location, the bulge of hair follicles. They are multipotent stem cells that have the physiological property to generate a wide array of differentiated cell types, including neurons, nerve supporting cells, smooth muscle cells, bone/cartilage cells and melanocytes. EPI-NCSC are easily accessible in the hairy skin and can be isolated as a highly pure population of stem cells. This video provides a detailed protocol for preparing mouse EPI-NCSC cultures from whisker follicles. The whisker pad of an adult mouse is removed, and whisker follicles dissected. The follicles are then cut longitudinally and subsequently transversely above and below the bulge region. The bulge is removed from the collagen capsule and placed in a culture plate. EPI-NCSC start to emigrate from the bulge explants 3 to 4 days later.

Mouse Epidermal Neural Crest Stem Cell EPI-NCSC Cultures

Here we show our method to isolate mouse epidermal neural crest stem cells (EPI-NCSC). Technique involves micro-dissecting whisker follicles, isolating the bulge and placeing it into tissue culture. EPI-NCSC start to emigrate from bulge explants onto the substratum within 3 - 4 days.

Souris épidermique Neural Stem Cell Crest (EPI-NCSC) cultures

EPI-NCSC are remnants of the embryonic neural crest in an adult location, the bulge of hair follicles. They are multipotent stem cells that have the ...

Preparation of 2-dGuo-Treated Thymus Organ Cultures

In the thymus, interactions between developing T-cell precursors and stromal cells that include cortical and medullary epithelial cells are known to play a key role in the development of a functionally competent T-cell pool. However, the complexity of T-cell development in the thymus in vivo can limit analysis of individual cellular components and particular stages of development. In vitro culture systems provide a readily accessible means to study multiple complex cellular processes. Thymus organ culture systems represent a widely used approach to study intrathymic development of T-cells under defined conditions in vitro. Here we describe a system in which mouse embryonic thymus lobes can be depleted of endogenous haemopoeitic elements by prior organ culture in 2-deoxyguanosine, a compound that is selectively toxic to haemopoeitic cells. As well as providing a readily accessible source of thymic stromal cells to investigate the role of thymic microenvironments in the development and selection of T-cells, this technique also underpins further experimental approaches that include the reconstitution of alymphoid thymus lobes in vitro with defined haemopoietic elements, the transplantation of alymphoid thymuses into recipient mice, and the formation of reaggregate thymus organ cultures. (This article is based on work first reported Methods in Molecular Biology 2007, Vol. 380 pages 185-196).

This video demonstrates the dissection and removal of the fetal thymus as well the preparation of ex vivo cultures of 2-dGuo-treated thymus.

Préparation de la 2-dGuo cultures traitées orgue Thymus

In the thymus, interactions between developing T-cell precursors and stromal cells that include cortical and medullary epithelial cells are known to play ...

Preparation and Maintenance of Dorsal Root Ganglia Neurons in Compartmented Cultures

Neurons extend axonal processes that are far removed from the cell body to innervate target tissues, where target-derived growth factors are required for neuronal survival and function. Neurotrophins are specifically required to maintain the survival and differentiation of innervating sensory neurons but the question of how these target-derived neurotrophins communicate to the cell body of innervating neurons has been an area of active research for over 30 years. The most commonly accepted model of how neurotrophin signals reach the cell body proposes that signaling endosomes carry this signal retrogradely along the axon. In order to study retrograde transport, a culture system was originally devised by Robert Campenot, in which cell bodies are isolated from their axons. The technique of preparing these compartmented chambers for culturing sensory neurons recapitulates the selective stimulation of neuron terminals that occurs in vivo following release of target-derived neurotrophins. Retrograde signaling events that require long-range microtubule dependent retrograde transport have important implications for the treatment of neurodegenerative disorders.

Here we describe the technique of preparing and maintaining compartmented chambers for culturing sensory neurons of the dorsal root ganglia.

Préparation et entretien des neurones ganglions de la racine dorsale dans les cultures à compartiments

Neurons extend axonal processes that are far removed from the cell body to innervate target tissues, where target-derived growth factors are required for ...

The Preparation of Primary Hematopoietic Cell Cultures From Murine Bone Marrow for Electroporation

It is becoming increasingly apparent that electroporation is the most effective way to introduce plasmid DNA or siRNA into primary cells. The Gene Pulser MXcell  electroporation system and Gene Pulser  electroporation buffer were specifically developed to transfect nucleic acids into mammalian cells and difficult-to-transfect cells, such as primary and stem cells.This video demonstrates how to establish primary hematopoietic cell cultures from murine bone marrow, and then prepare them for electroporation in the MXcell system. We begin by isolating femur and tibia. Bone marrow from both femur and tibia are then harvested and cultures are established. Cultured bone marrow cells are then transfected and analyzed.

This procedure describes how to establish primary hematopoietic cell cultures from murine bone marrow and is followed by transfection using the Gene Pulser MXCell electroporation system.

La préparation des cultures primaires de cellules hématopoïétiques de la moelle osseuse murine pour l&#39;électroporation

It is becoming increasingly apparent that electroporation is the most effective way to introduce plasmid DNA or siRNA into primary cells. The Gene Pulser ...

Preparation of Rat Brain Aggregate Cultures for Neuron and Glia Development Studies

An in vitro system that recapitulates the development and differentiation of progenitors into mature neurons and glia in the central nervous system (CNS) would provide a powerful platform for neuroscientists to investigate axo-glial interactions, properties and differentiation of multipotent progenitors, and progression of oligodendroglial lineage cells at the cellular and molecular level.  We describe here a CNS aggregate culture system from embryonic rat forebrains, which can be maintained in a serum-free medium up to 3-4 weeks and is used in our laboratory as a model to study neuron-glia interaction and CNS myelination.  This video clip will demonstrate how to isolate and grow these CNS aggregate cultures from E16 rat brain.  Furthermore, from the same brain dissection, highly enriched regular dissociated neuronal cultures can be readily obtained and used for various studies on CNS neurons or used for co-cultures with other cells. 

A protocols for an embryonic rat brain aggregate culture system is described. Multipotent progenitors in the aggregates can develop and differentiate into neurons, astrocytes and oligodendrocytes.

Préparation des cultures de cerveau de rat d&#39;agrégation pour Neuron et études sur le développement Glia

An in vitro system that recapitulates the development and differentiation of progenitors into mature neurons and glia in the central nervous system (CNS) ...