Ce travail a été soutenu par le NIH / NINDS NS octroi 4R00 057940, et accorder SCBI 200-12 de la moelle épinière et de financer des recherches sur le cerveau des blessures de l&#39;Indiana State Department de la Santé.

1. Faire un simple appareil pour la chirurgie de dégagement <ol><li> Le dispositif de dégagement nous avons créé est composé de trois parties (fig. 1): (1) une plaque de support en acier inoxydable ou en plastique qui permet la fixation d&#39;un tube de guidage et d&#39;une aiguille, et s&#39;assoit en face de la fenêtre lors de la chirurgie crânienne, (2) un tube de guidage qui détient une aiguille dans la position et permet la rotation d&#39;aiguilles, et (3) une aiguille qui est plié à 90 degrés à environ 3 mm de la pointe, et est mobile par rotation et d&#39;insertion. </li><li> Coupez un morceau de 1-1,5 mm d&#39;épaisseur, 7 ~ 10 x 30 mm en acier inoxydable ou rectangulaire en plastique transparent (1) pour faire la plaque de support. Préparer un pouce 1.5 de calibre 22 (BD société, # 305156) et un pouce 1.5 de calibre 25 (BD société, # 305127) aiguille de la seringue. Pour faire un tube de guidage, couper le bout de plastique et la pointe inférieure aiguille de l&#39;aiguille de la seringue de calibre 22 de sorte que sa longueur totale est d&#39;environ 31 à 32 mm (2). Sable fin en métal pour le rendre plat et lisse. La longueur finale de ce tube de guidage devrait être d&#39;environ 5-6 mm plus courte que l&#39;aiguille de la seringue de calibre 25. </li><li> Utilisez de la colle cyanoacrylate pour fixer le tube de guidage sur la plaque de support, en s&#39;assurant que l&#39;aiguille est perpendiculaire au bord du métal. </li><li> Insérez l&#39;aiguille de calibre 25 dans le tube de guidage, et pliez les 90 degrés l&#39;aiguille de 2,5-3 mm de la pointe (fig. 1). </li><li> Réglez la plage de déplacement vertical de l&#39;aiguille en collant un petit tube en plastique sur l&#39;extrémité de l&#39;aiguille (fig. 1, arrêt de l&#39;aiguille). La gamme de finale déplacement de l&#39;aiguille qui détermine la profondeur de l&#39;aiguille peut être insérée dans le cortex, et devrait être 1.6 à 1.8 mm pour P21 rats et 01.03 à 01.05 mm pour P21 souris. Cette longueur doit être ajusté pour les différentes espèces et l&#39;âge des animaux. </li><li> Attachez un fil de cuivre de petites ou un petit morceau de ruban adhésif sur l&#39;extrémité supérieure de l&#39;aiguille dans la même direction que le bout plié (Fig.1 indicateur à aiguille). Le fils ou des bandes indique l&#39;angle de braquage de l&#39;aiguille lors de la chirurgie inférieurs (Fig. 1). </li></ol> 2. La préparation des animaux <ol><li> Tous les instruments chirurgicaux doivent être autoclavés, stérilisés avec un stérilisateur perles de verre, ou désinfectée avec de l&#39;éthanol à 70%. </li><li> Nous utilisons des rats Sprague / Dawley à l&#39;âge post-natal de 20 à 22 jours (P20-22) ou de souris CD1 au même âge. Différentes souches de souris ou de rats peut être utilisé pour des projets spécifiques. </li><li> Anesthésier la souris avec une injection ip de la kétamine 80 mg / kg et de xylazine 8 mg / kg. La procédure chirurgicale commence après que l&#39;animal ne répond pas à pincer la queue. Pendant la chirurgie, si nécessaire, une dose de rappel d&#39;un tiers de la dose initiale du cocktail anesthésiant peut être donnée pour restaurer l&#39;état original anesthésique. </li><li> Couper les cheveux sur le cuir chevelu de l&#39;animal avec une coupe de cheveux électriques. Appliquez une petite quantité de pommade ophtalmique sur les yeux de l&#39;animal pour la protection de la sécheresse pendant l&#39;anesthésie. Désinfecter le cuir chevelu avec un 10% de povidone iodée solution, suivi par 70% d&#39;éthanol. </li><li> Mont de l&#39;animal sur un appareil stéréotaxique de garder la tête dans une position stable. Tout au long de la chirurgie, nous gardons l&#39;animal sur un coussin chauffant pour prévenir l&#39;hypothermie. </li><li> Faites une ligne médiane antéro-postérieure incision sur le cuir chevelu à l&#39;aide d&#39;un scalpel, s&#39;étendant de lambda pour entre les yeux. Utilisez hémostatiques de tirer la peau de côté et d&#39;exposer le crâne laissé suffisamment. </li><li> Faites une entaille rectangulaire sur le crâne de gauche, et utilisez le scalpel pour gratter le périoste. Cette étape permettra de réduire les saignements et faciliter le forage sur le crâne. </li></ol> 3. Faire des contre-dépouilles <ol><li> Ajoutez une petite quantité de solution saline stérile sur la zone du crâne exposé, et ensuite utiliser plusieurs Q-tips pour nettoyer le sang et sécher la zone. </li><li> Sous un microscope chirurgical, commencer le forage d&#39;une rainure rectangulaire (~ 5 x 7 mm chez le rat, 4 x 5 mm chez la souris) sur le centre du crâne à gauche (environ au-dessus du cortex sensorimoteur gauche). Appliquer une goutte de solution saline sur le crâne facilite le forage et la chaleur se dissipe. Après environ 2 / 3 la profondeur de l&#39;os est percé, enlever l&#39;excès salins et nettoyer la zone de forage avec un Q-tip. Lentement et sagement forer plus profond jusqu&#39;à ce que la pièce centrale de l&#39;os est mobile au toucher doux d&#39;une pince. </li><li> Retirez délicatement la pièce centrale de l&#39;os en insérant une pointe acérée d&#39;une pince sur le bord de l&#39;os et lentement la pince de levage pour exposer l&#39;hémisphère gauche. </li><li> Sous le microscope chirurgical, tenez l&#39;appareil contre-dépouilles, et d&#39;orienter l&#39;aiguille dans une direction parasagittale et la perpendiculaire à la plaque de support de la ligne médiane, inclinant l&#39;appareil légèrement caudalement afin de maintenir la visualisation de la pointe de l&#39;aiguille et la région cible corticale. Dirigez la pointe de l&#39;aiguille d&#39;une zone de 1-2 mm latéral à la suture sagittal supérieur, dans le milieu de la fenêtre crânienne, et d&#39;éviter la pénétration directe des grands navires. </li><li> Insérez l&#39;aiguille dans une direction horizontale à travers la dure-mère et sous la pie de manière à épargner les vaisseaux sanguins. Asseyez-vous en bas de l&#39;appareil inférieurssur les deux bords de la fenêtre crânienne afin que l&#39;aiguille est placée perpendiculairement à la surface corticale (Fig. 1). Reposant l&#39;appareil sur les bords de la fenêtre à la fois crânienne à la main permettra de réduire considérablement ou d&#39;éliminer la poignée de main pendant la procédure suivante. Soulevez l&#39;aiguille au-dessous du PIA, puis abaissez lentement pour créer une coupe transcorticale ce que l&#39;aiguille ne peut pas aller plus loin. Rotation ~ 135 ° loin de la ligne médiane afin de créer un sujet en demi-cercle blanc / couche profonde VI dépouille. Puis levez l&#39;aiguille de nouveau sous le pia. Inclinez l&#39;appareil vers l&#39;arrière et retirer l&#39;aiguille. </li><li> Optionnellement, on peut répéter la procédure ci-dessus (étape 2.5), mais sans tourner l&#39;aiguille de manière à créer une coupure supplémentaire transcorticale sur les bords latéraux de la fenêtre. Cela va créer un isolement plus complet corticale. </li><li> Placer un morceau de film plastique (6 x 6 mm) sur la fenêtre crânienne pour la protection et de suture au cuir chevelu. Placez l&#39;animal sur un tapis chauffé jusqu&#39;à ce que complètement récupéré de l&#39;anesthésie. </li></ol> 4. Les résultats représentatifs: Coronal tranches corticales peuvent être préparés pour confirmer le succès de la chirurgie dépouille. Dans les tranches préparées&gt; 2 semaines après la chirurgie, les coupures transcorticale et de saper sont perceptibles dans l&#39;objectif de faible puissance d&#39;un microscope (fig. 2). Le cortex partiellement isolé devient généralement un peu plus mince, et l&#39;activité épileptiforme peut être détecté dans la majorité des tranches de cortex avec enregistrement sur le terrain de potentiel (Fig. 3). En revanche, la formation de grands trous dans la matière blanche ou dans les couches corticales profondes, dramatique amincissement du cortex lésé, ou une coupe errants dessus ou en dessous de la substance blanche du cerveau fera inutilisable pour d&#39;autres expériences. <img src="/files/ftp_upload/2840/2840fig1.jpg" alt="Figure 1"/> Figure 1. Structure et application d&#39;un dispositif contre-dépouille. Un tube de guidage (2) est collé sur une plaque de support (1) qui est faite d&#39;acier inoxydable ou en plastique transparent. Une aiguille de la seringue (3) est inséré à travers le tube de guidage et plié à environ 3 mm à la pointe. Une aiguille d&#39;arrêt en tube de plastique est collée sur l&#39;extrémité supérieure de l&#39;aiguille de sorte que la gamme verticale mobile de l&#39;aiguille est limité à ~ 1,2 mm et ~ 1,5 mm pour une utilisation dans P21 souris et des rats, respectivement. Un segment de fil de cuivre est fixé petits dessous de la poignée comme aiguille de l&#39;indicateur pour l&#39;orientation de l&#39;aiguille tordue. Notez que l&#39;appareil est incliné et en contact avec les deux bords de la fenêtre crânienne afin qu&#39;une coupe peut être faite parallèlement à la surface piales. <img src="/files/ftp_upload/2840/2840fig2.jpg" alt="Figure 2"/> Figure 2. Une image représentative de la tranche dépouille. A. image fluorescente d&#39;une tranche qui a été préparé deux semaines après la lésion dépouille dans un rat P48. La blessure de coupe a été marqué par fluorescence DiI colorant. Les flèches blanches sur la droite indiquent coupé transcorticale, et les flèches en bas indiquent inférieurs qui sont passés si la frontière entre la couche IV et la matière blanche. <img src="/files/ftp_upload/2840/2840fig3.jpg" alt="Figure 3"/> Figure 3. Enregistrement sur ​​le terrain le potentiel d&#39;une tranche d&#39;enregistrement inférieurs potentiels. Champ d&#39;une tranche de cerveau présentaient une activité épileptiforme inférieurs, suggérant une hyperexcitabilité du tissu lésé corticale.

<ol>
	<li>Hoffman, S. N., Salin, P. A., Prince, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronic+neocortical+epileptogenesis+in+vitro.">Chronic neocortical epileptogenesis in vitro.</a> J Neurophysiol. 71, 1762-1773 (1994).</li><li>Topolnik, L., Steriade, M., Timofeev, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hyperexcitability+of+intact+neurons+underlies+acute+development+of+trauma-related+electrographic+seizures+in+cats+in+vivo.">Hyperexcitability of intact neurons underlies acute development of trauma-related electrographic seizures in cats in vivo.</a> Eur J Neurosci. 18, 486-496 (2003).</li><li>Graber, K., Prince, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Models of Seizures and Epilepsy. , 477-493 (2005).</li><li>Nita, D. A., Cisse, Y., Timofeev, I., Steriade, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increased+propensity+to+seizures+after+chronic+cortical+deafferentation+in+vivo.">Increased propensity to seizures after chronic cortical deafferentation in vivo.</a> J Neurophysiol. 95, 902-913 (2006).</li><li>Sharpless, S. K., Halpern, L. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+electrical+excitability+of+chronically+isolated+cortex+studied+by+means+of+permanently+implanted+electrodes.">The electrical excitability of chronically isolated cortex studied by means of permanently implanted electrodes.</a> Electroencephalogr Clin Neurophysiol. 14, 244-255 (1962).</li><li>Echlin, F. A., Battista, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epileptiform+Seizures+from+Chronic+Isolated+Cortex.">Epileptiform Seizures from Chronic Isolated Cortex.</a> Arch Neurol. 9, 154-170 (1963).</li><li>Prince, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epileptogenic+neurons+and+circuits.">Epileptogenic neurons and circuits.</a> Adv Neurol. 79, 665-684 (1999).</li><li>Marin-Padilla, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Developmental+neuropathology+and+impact+of+perinatal+brain+damage.+II:+white+matter+lesions+of+the+neocortex.">Developmental neuropathology and impact of perinatal brain damage. II: white matter lesions of the neocortex.</a> J Neuropathol Exp Neurol. 56, 219-235 (1997).</li><li>Jin, X., Prince, D. A., Huguenard, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enhanced+excitatory+synaptic+connectivity+in+layer+v+pyramidal+neurons+of+chronically+injured+epileptogenic+neocortex+in+rats.">Enhanced excitatory synaptic connectivity in layer v pyramidal neurons of chronically injured epileptogenic neocortex in rats.</a> J Neurosci. 26, 4891-4900 (2006).</li><li>Li, H., Prince, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synaptic+activity+in+chronically+injured,+epileptogenic+sensory-motor+neocortex.">Synaptic activity in chronically injured, epileptogenic sensory-motor neocortex.</a> J Neurophysiol. 88, 2-12 (2002).</li><li>Salin, P., Tseng, G. F., Hoffman, S., Parada, I., Prince, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Axonal+sprouting+in+layer+V+pyramidal+neurons+of+chronically+injured+cerebral+cortex.">Axonal sprouting in layer V pyramidal neurons of chronically injured cerebral cortex.</a> J Neurosci. 15, 8234-8245 (1995).</li><li>Avramescu, S., Nita, D. A., Timofeev, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neocortical+post-traumatic+epileptogenesis+is+associated+with+loss+of+GABAergic+neurons.">Neocortical post-traumatic epileptogenesis is associated with loss of GABAergic neurons.</a> J Neurotrauma. 26, 799-812 (2009).</li><li>Avramescu, S., Timofeev, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synaptic+strength+modulation+after+cortical+trauma:+a+role+in+epileptogenesis.">Synaptic strength modulation after cortical trauma: a role in epileptogenesis.</a> J Neurosci. 28, 6760-6772 (2008).</li></ol>

Aucun conflit d&#39;intérêt déclaré.

Le modèle dégagement est un système très efficace pour étudier épileptogenèse post-traumatique. Une chirurgie typique prend seulement environ 20-30 minutes pour finir, et évoquée ou activité épileptiforme spontanée peut être enregistré dans les tranches de la plupart des animaux deux semaines après la chirurgie 1-2. Plus important encore, ce modèle imite les aspects des changements suite à une blessure traumatique du cerveau telles que saignements, inflammation, œdème, axotomie et la mort n...

<table border="1"><tbody><tr><td> Nom du produit </td><td> Tapez </td><td> Société </td><td> Numéro de catalogue </td></tr><tr><td> Foredom micromoteur kit </td><td> équipements </td><td> Foredom </td><td> K.1070 </td></tr><tr><td> 1,5 pouces aiguille de la seringue de calibre 22 </td><td> matériels </td><td> La société BD </td><td> 305156 </td></tr><tr><td> 1,5 pouces d&#39;aiguille de seringue de calibre 25 </td><td> matériels </td><td> La société BD </td><td> 305127 </td></tr><tr><td> Colle cyanoacrylate </td><td> matériels </td><td> Ted Pella </td><td> 14450 </td></tr></tbody></table>

preparing undercut model of posttraumatic epileptogenesis in rodents

Partiellement isolé du cortex («dégagement») est un modèle animal de l&#39;épileptogenèse post-traumatique. La procédure chirurgicale consiste à couper à travers le cortex sensori-moteur et le dessous de la substance blanche (dessous) afin qu&#39;une région spécifique du cortex cérébral est largement isolés du cortex voisins et régions sous-corticales 1-3. Après un temps de latence de deux semaines ou plus après la chirurgie, décharges épileptiformes peuvent être enregistrés dans des tranches de cerveau de rongeurs 1 et crises électriques ou de comportement peuvent être observés in vivo à partir d&#39;autres espèces comme chat et le singe 4-6. Ce modèle animal bien établi est efficace pour générer et imite plusieurs caractéristiques importantes de lésions cérébrales traumatiques. Cependant, il est techniquement difficile de tenter de faire précise des lésions corticales du cerveau petit rongeur avec une main libre. Basé sur la procédure établie initialement au laboratoire du Dr David Prince à l&#39;Université Stanford 1, ici nous présentons une technique améliorée pour effectuer une intervention chirurgicale pour la préparation de ce modèle chez la souris et les rats. Nous démontrons comment faire un simple dispositif chirurgical et l&#39;utiliser pour acquérir une meilleure maîtrise de la profondeur de coupe et l&#39;angle de générer des résultats plus précis et plus cohérents. L&#39;appareil est facile à faire, et la procédure est rapide à apprendre. La génération de ce modèle animal fournit un système efficace pour l&#39;étude sur les mécanismes de l&#39;épileptogenèse post-traumatique.

Watch this Scientific Journal Video about Preparing Undercut Model of Posttraumatic Epileptogenesis in Rodents at JoVE.com

Preparing Undercut Model of Posttraumatic Epileptogenesis in Rodents

Partiellement isolé du cortex («dégagement») est un modèle animal l&#39;efficacité de l&#39;épileptogenèse post-traumatique. Ici, nous montrer comment faire un nouveau dispositif chirurgical et l&#39;utiliser pour faire des lésions plus précise et plus cohérente pour générer ce modèle.

Préparer Modèle dépouille de épileptogenèse post-traumatique chez les rongeurs

Partially isolated cortex ("undercut") is an animal model of posttraumatic epileptogenesis. The surgical procedure involves cutting through the ...

preparing-undercut-model-of-posttraumatic-epileptogenesis-in-rodents

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Neuroscience

9.3K vues.  Indiana University School of Medicine. Partiellement isolé du cortex («dégagement») est un modèle animal l'efficacité de l'épileptogenèse post-traumatique. Ici, nous montrer comment faire un nouveau dispositif chirurgical et l'utiliser pour faire des lésions plus précise et plus cohérente pour générer ce modèle.

Vidéo: Preparing Undercut Model of Posttraumatic Epileptogenesis in Rodents

Habituation and Prepulse Inhibition of Acoustic Startle in Rodents

The acoustic startle response is a protective response, elicited by a sudden and intense acoustic stimulus. Facial and skeletal muscles are activated within a few milliseconds, leading to a whole body flinch in rodents1. Although startle responses are reflexive responses that can be reliably elicited, they are not stereotypic. They can be modulated by emotions such as fear (fear potentiated startle) and joy (joy attenuated startle), by non-associative learning processes such as habituation and sensitization, and by other sensory stimuli through sensory gating processes (prepulse inhibition), turning startle responses into an excellent tool for assessing emotions, learning, and sensory gating, for review see 2, 3. The primary pathway mediating startle responses is very short and well described, qualifying startle also as an excellent model for studying the underlying mechanisms for behavioural plasticity on a cellular/molecular level3.
We here describe a method for assessing short-term habituation, long-term habituation and prepulse inhibition of acoustic startle responses in rodents. Habituation describes the decrease of the startle response magnitude upon repeated presentation of the same stimulus. Habituation within a testing session is called short-term habituation (STH) and is reversible upon a period of several minutes without stimulation. Habituation between testing sessions is called long-term habituation (LTH)4. Habituation is stimulus specific5. Prepulse inhibition is the attenuation of a startle response by a preceding non-startling sensory stimulus6. The interval between prepulse and startle stimulus can vary from 6 to up to 2000 ms. The prepulse can be any modality, however, acoustic prepulses are the most commonly used.
Habituation is a form of non-associative learning. It can also be viewed as a form of sensory filtering, since it reduces the organisms' response to a non-threatening stimulus. Prepulse inhibition (PPI) was originally developed in human neuropsychiatric research as an operational measure for sensory gating7. PPI deficits may represent the interface of "psychosis and cognition" as they seem to predict cognitive impairment8-10. Both habituation and PPI are disrupted in patients suffering from schizophrenia11, and PPI disruptions have shown to be, at least in some cases, amenable to treatment with mostly atypical antipsychotics12, 13. However, other mental and neurodegenerative diseases are also accompanied by disruption in habituation and/or PPI, such as autism spectrum disorders (slower habituation), obsessive compulsive disorder, Tourette's syndrome, Huntington's disease, Parkinson's disease, and Alzheimer's Disease (PPI)11, 14, 15 Dopamine induced PPI deficits are a commonly used animal model for the screening of antipsychotic drugs16, but PPI deficits can also be induced by many other psychomimetic drugs, environmental modifications and surgical procedures.

Habituation and prepulse inhibition of startle are operational measures of sensory gating. Sensory gating is disrupted in schizophrenia, and some other mental disorders and neurodegenerative diseases. We here describe a standard protocol to assess short-term and long-term habituation as well as prepulse inhibition of acoustic startle responses in rats and mice.

Inhibition préimpulsion habituation et du réflexe de sursaut acoustique dans Rongeurs

The acoustic startle response is a protective response, elicited by a sudden and intense acoustic stimulus. Facial and skeletal muscles are activated ...

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Neurosciences

Large-scale Recording of Neurons by Movable Silicon Probes in Behaving Rodents

A major challenge in neuroscience is linking behavior to the collective activity of neural assemblies. Understanding of input-output relationships of neurons and circuits requires methods with the spatial selectivity and temporal resolution appropriate for mechanistic analysis of neural ensembles in the behaving animal, i.e. recording of representatively large samples of isolated single neurons. Ensemble monitoring of neuronal activity has progressed remarkably in the past decade in both small and large-brained animals, including human subjects1-11. Multiple-site recording with silicon-based devices are particularly effective because of their scalability, small volume and geometric design.
Here, we describe methods for recording multiple single neurons and local field potential in behaving rodents, using commercially available micro-machined silicon probes with custom-made accessory components. There are two basic options for interfacing silicon probes to preamplifiers: printed circuit boards and flexible cables. Probe supplying companies (<a href="http://www.neuronexustech.com/" >http://www.neuronexustech.com/</a>; <a href="http://www.sbmicrosystems.com/">http://www.sbmicrosystems.com/</a>; <a href="http://www.acreo.se/">http://www.acreo.se/</a>) usually provide the bonding service and deliver probes bonded to printed circuit boards or flexible cables. Here, we describe the implantation of a 4-shank, 32-site probe attached to flexible polyimide cable, and mounted on a movable microdrive. Each step of the probe preparation, microdrive construction and surgery is illustrated so that the end user can easily replicate the process.

We describe methods for large-scale recording of multiple single units and local field potential in behaving rodents with silicon probes. Drive fabrication, probe attachment to the drive and probe implantation processes are illustrated in sufficient details for easy replication.

À grande échelle d&#39;enregistrement des neurones par des sondes de silicium mobiles à se comporter Rongeurs

A major challenge in neuroscience is linking behavior to the collective activity of neural assemblies. Understanding of input-output relationships of ...

A Low Cost Setup for Behavioral Audiometry in Rodents

In auditory animal research it is crucial to have precise information about basic hearing parameters of the animal subjects that are involved in the experiments. Such parameters may be physiological response characteristics of the auditory pathway, e.g. via brainstem audiometry (BERA). But these methods allow only indirect and uncertain extrapolations about the auditory percept that corresponds to these physiological parameters. To assess the perceptual level of hearing, behavioral methods have to be used. A potential problem with the use of behavioral methods for the description of perception in animal models is the fact that most of these methods involve some kind of learning paradigm before the subjects can be behaviorally tested, e.g. animals may have to learn to press a lever in response to a sound. As these learning paradigms change perception itself 1,2 they consequently will influence any result about perception obtained with these methods and therefore have to be interpreted with caution. Exceptions are paradigms that make use of reflex responses, because here no learning paradigms have to be carried out prior to perceptual testing. One such reflex response is the acoustic startle response (ASR) that can highly reproducibly be elicited with unexpected loud sounds in na&iuml;ve animals. This ASR in turn can be influenced by preceding sounds depending on the perceptibility of this preceding stimulus: Sounds well above hearing threshold will completely inhibit the amplitude of the ASR; sounds close to threshold will only slightly inhibit the ASR. This phenomenon is called pre-pulse inhibition (PPI) 3,4, and the amount of PPI on the ASR gradually depends on the perceptibility of the pre-pulse. PPI of the ASR is therefore well suited to determine behavioral audiograms in na&iuml;ve, non-trained animals, to determine hearing impairments or even to detect possible subjective tinnitus percepts in these animals. In this paper we demonstrate the use of this method in a rodent model (cf. also ref. 5), the Mongolian gerbil (Meriones unguiculatus), which is a well know model species for startle response research within the normal human hearing range (e.g. 6).

A fast and inexpensive method for the behavioral determination of hearing parameters like hearing thresholds, hearing impairments or phantom perceptions (subjective tinnitus) is described. It uses pre-pulse inhibition of the acoustic startle response and can be easily implemented in a personal computer using a programmable AD/DA-converter and a piezo sensor.

Une configuration à faible coût pour l&#39;audiométrie comportementale chez les rongeurs

In auditory animal research it is crucial to have precise information about basic hearing parameters of the animal subjects that are involved in the ...

Recording and Analysis of Circadian Rhythms in Running-wheel Activity in Rodents

When rodents have free access to a running wheel in their home cage, voluntary use of this wheel will depend on the time of day1-5. Nocturnal rodents, including rats, hamsters, and mice, are active during the night and relatively inactive during the day. Many other behavioral and physiological measures also exhibit daily rhythms, but in rodents, running-wheel activity serves as a particularly reliable and convenient measure of the output of the master circadian clock, the suprachiasmatic nucleus (SCN) of the hypothalamus. In general, through a process called entrainment, the daily pattern of running-wheel activity will naturally align with the environmental light-dark cycle (LD cycle; e.g. 12 hr-light:12 hr-dark). However circadian rhythms are endogenously generated patterns in behavior that exhibit a ~24 hr period, and persist in constant darkness. Thus, in the absence of an LD cycle, the recording and analysis of running-wheel activity can be used to determine the subjective time-of-day. Because these rhythms are directed by the circadian clock the subjective time-of-day is referred to as the circadian time (CT). In contrast, when an LD cycle is present, the time-of-day that is determined by the environmental LD cycle is called the zeitgeber time (ZT).
Although circadian rhythms in running-wheel activity are typically linked to the SCN clock6-8, circadian oscillators in many other regions of the brain and body9-14 could also be involved in the regulation of daily activity rhythms. For instance, daily rhythms in food-anticipatory activity do not require the SCN15,16 and instead, are correlated with changes in the activity of extra-SCN oscillators17-20. Thus, running-wheel activity recordings can provide important behavioral information not only about the output of the master SCN clock, but also on the activity of extra-SCN oscillators. Below we describe the equipment and methods used to record, analyze and display circadian locomotor activity rhythms in laboratory rodents.

Circadian rhythms in voluntary wheel-running activity in mammals are tightly coupled to the molecular oscillations of a master clock in the brain. As such, these daily rhythms in behavior can be used to study the influence of genetic, pharmacological, and environmental factors on the functioning of this circadian clock.

Enregistrement et analyse des rythmes circadiens dans la course roue-activité chez les rongeurs

When rodents have free access to a running wheel in their home cage, voluntary use of this wheel will depend on the time of day1-5. Nocturnal rodents, ...

Deep Brain Stimulation with Simultaneous fMRI in Rodents

In order to visualize the global and downstream neuronal responses to deep brain stimulation (DBS) at various targets, we have developed a protocol for using blood oxygen level dependent (BOLD) functional magnetic resonance imaging (fMRI) to image rodents with simultaneous DBS. DBS fMRI presents a number of technical challenges, including accuracy of electrode implantation, MR artifacts created by the electrode, choice of anesthesia and paralytic to minimize any neuronal effects while simultaneously eliminating animal motion, and maintenance of physiological parameters, deviation from which can confound the BOLD signal.&#160;Our laboratory has developed a set of procedures that are capable of overcoming most of these possible issues. For electrical stimulation, a homemade tungsten bipolar microelectrode is used, inserted stereotactically at the stimulation site in the anesthetized subject. In preparation for imaging, rodents are fixed on a plastic headpiece and transferred to the magnet bore. For sedation and paralysis during scanning, a cocktail of dexmedetomidine and pancuronium is continuously infused, along with a minimal dose of isoflurane; this preparation minimizes the BOLD ceiling effect of volatile anesthetics. In this example experiment, stimulation of the subthalamic nucleus (STN) produces BOLD responses which are observed primarily in ipsilateral cortical regions, centered in motor cortex. Simultaneous DBS and fMRI allows the unambiguous modulation of neural circuits dependent on stimulation location and stimulation parameters, and permits observation of neuronal modulations free of regional bias. This technique may be used to explore the downstream effects of modulating neural circuitry at nearly any brain region, with implications for both experimental and clinical DBS.

This protocol describes a standard method for simultaneous functional magnetic resonance imaging and deep brain stimulation in the rodent. The combined use of these experimental tools allows for the exploration of global downstream activity in response to electrical stimulation at virtually any brain target.

La stimulation cérébrale profonde avec simultanée IRMf chez les rongeurs

In order to visualize the global and downstream neuronal responses to deep brain stimulation (DBS) at various targets, we have developed a protocol for ...

Acrylic Resin Molding Based Head Fixation Technique in Rodents

Head fixation is a technique of immobilizing animal's head by attaching a head-post on the skull for rigid clamping. Traditional head fixation requires surgical attachment of metallic frames on the skull. The attached frames are then clamped to a stationary platform resulting in immobilization of the head. However, metallic frames for head fixation have been technically difficult to design and implement in general laboratory environment. In this study, we provide a novel head fixation method. Using a custom-made head fixation bar, head mounter is constructed during implantation surgery. After the application of acrylic resin for affixing implants such as electrodes and cannula on the skull, additional resins applied on top of that to build a mold matching to the port of the fixation bar. The molded head mounter serves as a guide rails, investigators conveniently fixate the animal&#8217;s head by inserting the head mounter into the port of the fixation bar. This method could be easily applicable if implantation surgery using dental acrylics is necessary and might be useful for laboratories that cannot easily fabricate CNC machined metal head-posts.

Traditional head fixation techniques have used metallic frames attached on the skull as an interface for head fixation apparatus. This protocol demonstrates a frameless, acrylic resin molding based head fixation technique for behavioral tasks in rodents.

Acrylique mouler à base Chef Fixation Technique chez les rongeurs

Head fixation is a technique of immobilizing animal's head by attaching a head-post on the skull for rigid clamping. Traditional head fixation requires ...

Hydraulic Extrusion of the Spinal Cord and Isolation of Dorsal Root Ganglia in Rodents

Traditionally, the spinal cord is isolated by laminectomy, i.e. by breaking open the spinal vertebrae one at a time. This is both time consuming and may result in damage to the spinal cord caused by the dissection process. Here, we show how the spinal cord can be extruded using hydraulic pressure. Handling time is significantly reduced to only a few minutes, likely decreasing protein damage. The low risk of damage to the spinal cord tissue improves subsequent immunohistochemical analysis. By performing hydraulic spinal cord extrusion instead of traditional laminectomy, the rodents can further be used for DRG isolation, thereby lowering the number of animals and allowing analysis across tissues from the same rodent. We demonstrate a consistent method to identify and isolate the DRGs according to their localization relative to the costae. It is, however, important to adjust this method to the particular animal used, as the number of spinal cord segments, both thoracic and lumbar, may vary according to animal type and strain. In addition, we illustrate further processing examples of the isolated tissues.

Here, we present a protocol for hydraulic extrusion of the spinal cord as well as identification and isolation of specific dorsal root ganglia (DRGs) in the same rodent. Compared to standard spinal cord isolation methods, this method is significantly faster and reduces the risk of tissue damage.

Extrusion hydraulique de la moelle épinière et isolement de ganglions rachidiens en Rongeurs

Traditionally, the spinal cord is isolated by laminectomy, i.e. by breaking open the spinal vertebrae one at a time. This is both time consuming and may ...

Transcranial Electrical Brain Stimulation in Alert Rodents

Transcranial electrical brain stimulation can modulate cortical excitability and plasticity in humans and rodents. The most common form of stimulation in humans is transcranial direct current stimulation (tDCS). Less frequently, transcranial alternating current stimulation (tACS) or transcranial random noise stimulation (tRNS), a specific form of tACS using an electrical current applied randomly within a pre-defined frequency range, is used. The increase of noninvasive electrical brain stimulation research in humans, both for experimental and clinical purposes, has yielded an increased need for basic, mechanistic, safety studies in animals. This article describes a model for transcranial electrical brain stimulation (tES) through the intact skull targeting the motor system in alert rodents. The protocol provides step-by-step instructions for the surgical set-up of a permanent epicranial electrode socket combined with an implanted counter electrode on the chest. By placing a stimulation electrode into the epicranial socket, different electrical stimulation types, comparable to tDCS, tACS, and tRNS in humans, can be delivered. Moreover, the practical steps for tES in alert rodents are introduced. The applied current density, stimulation duration, and stimulation type may be chosen depending on the experimental needs. The caveats, advantages, and disadvantages of this set-up are discussed, as well as safety and tolerability aspects.

This protocol describes a surgical set-up for a permanent epicranial electrode socket and an implanted chest electrode in rodents. By placing a second electrode into the socket, different types of transcranial electrical brain stimulation can be delivered to the motor system in alert animals through the intact skull.

Stimulation transcrânienne électrique cérébrale chez les rongeurs alertes

Transcranial electrical brain stimulation can modulate cortical excitability and plasticity in humans and rodents. The most common form of stimulation in ...

Cannula Implantation into the Cisterna Magna of Rodents

Cisterna magna cannulation (CMc) is a straightforward procedure that enables direct access to the cerebrospinal fluid (CSF) without operative damage to the skull or the brain parenchyma. In anesthetized rodents, the exposure of the dura mater by blunt dissection of the neck muscles allows the insertion of a cannula into the cisterna magna (CM). The cannula, composed either by a fine beveled needle or borosilicate capillary, is attached via a polyethylene (PE) tube to a syringe. Using a syringe pump, molecules can then be injected at controlled rates directly into the CM, which is continuous with the subarachnoid space. From the subarachnoid space, we can trace CSF fluxes by convective flow into the perivascular space around penetrating arterioles, where solute exchange with the interstitial fluid (ISF) occurs. CMc can be performed for acute injections immediately following the surgery, or for chronic implantation, with later injection in anesthetized or awake, freely moving rodents. Quantitation of tracer distribution in the brain parenchyma can be performed by epifluorescence, 2-photon microscopy, and magnetic resonance imaging (MRI), depending on the physico-chemical properties of the injected molecules. Thus, CMc in conjunction with various imaging techniques offers a powerful tool for assessment of the glymphatic system and CSF dynamics and function. Furthermore, CMc can be utilized as a conduit for fast, brain-wide delivery of signaling molecules and metabolic substrates that could not otherwise cross the blood brain barrier (BBB).

Here we describe a protocol to perform cisterna magna cannulation (CMc), a minimally invasive way to deliver tracers, substrates and signaling molecules into the cerebrospinal fluid (CSF). Combined with different imaging modalities, CMc enables glymphatic system and CSF dynamics assessment, as well as brain-wide delivery of various compounds.

Implantation de la canule dans la Magna de Cisterna de rongeurs

Cisterna magna cannulation (CMc) is a straightforward procedure that enables direct access to the cerebrospinal fluid (CSF) without operative damage to ...

Intracranial Pharmacotherapy and Pain Assays in Rodents

Pain is a salient sensory experience with affective and cognitive dimensions. However, central mechanisms for pain remain poorly understood, hindering the development of effective therapeutics. Intracranial pharmacology presents an important tool for understanding the molecular and cellular mechanisms of pain in the brain, as well as for novel treatments. Here we present a protocol that integrates intracranial pharmacology with pain behavior testing. Specifically, we show how to infuse analgesic drugs into a select brain region, which may be responsible for pain modulation. Furthermore, to determine the effect of the candidate drug in the central nerve system, pain assays are performed after intracranial treatment. Our results demonstrate that intracranial administration of analgesic drugs in a targeted region can provide relief of pain in rodents. Thus, our protocol successfully demonstrates that intracranial pharmacology, combined with pain behavior testing, can be a powerful tool for the study of pain mechanisms in the brain.

Here we present a protocol to perform intracranial pharmacological experiments followed by pain behavior assays in rodents. This protocol allows researchers to deliver molecular and cellular targets in the brain, for pharmacologic agents in the treatment of pain.

Pharmacothérapie intracrânienne et dosages de la douleur chez les rongeurs

Pain is a salient sensory experience with affective and cognitive dimensions. However, central mechanisms for pain remain poorly understood, hindering the ...