Bu çalışma Ulusal Sağlık Enstitüsü / NINDS Omurilik ve Indiana Eyalet Sağlık Bölümü Beyin Hasarı Araştırma fonu hibe 4R00 NS 057.940, ve hibe SCBI 200-12 tarafından desteklenmiştir.

1. Alttan ameliyatı için basit bir cihaz yapma <ol><li> Yarattığımız alttan cihazı (Şekil 1) üç bölümden oluşur: (1), yol gösterici bir tüp eki ve bir iğne sağlar ve ameliyat sırasında kranyal pencere karşısına geçiyor, paslanmaz çelik veya plastikten yapılmış bir destek plakası; (2) pozisyonda bir iğne tutan ve iğne dönme izin veren yol gösterici bir tüp; ve (3) ucundan ~ 3 mm 90 derece bükülmüş ve rotasyon ve ekleme hareketli bir iğne. </li><li> 1-1.5 mm kalınlığında, 7 ~ 10 x 30 mm dikdörtgen, paslanmaz çelik veya şeffaf plastik (1) destekleyen plaka yapmak için bir parça kesin. 1.5 inç 22-gauge (BD şirketi, # 305.156) ve 1.5 inç 25-gauge (BD şirketi, # 305.127) şırınga iğnesinin hazırlayın. Yol gösterici bir tüp yapmak için, plastik sonu ve böylece toplam uzunluğu 31-32 mm (2) yaklaşık 22-gauge şırınga iğne iğne ucu alt kesti. Kum metal uç düz ve pürüzsüz bir hale getirmek için. Bu kılavuz tüp nihai uzunluğu 25 gauge enjektör iğnesi 5-6 mm daha kısa olmalıdır. </li><li> Iğne metal kenarına dik olduğundan emin olun, destek plakası üzerine kılavuz tüpü düzeltmek için siyanoakrilat yapıştırıcı kullanın. </li><li> 25-gauge enjektör iğnesi kılavuz tüp içine yerleştirin ve ucu (Şekil 1) 2.5-3 mm iğne 90 derece bükün. </li><li> Iğne sonuna kadar (Şekil 1, iğne durdurma) üzerine yapıştırma küçük bir plastik tüp iğne dikey hareket aralığı ayarlayın. İğnenin son hareket aralığı ne kadar derin iğne korteks eklenebilir belirler ve, P21 sıçanlar için 1.6-1.8 mm ve P21 fareler için 1,3-1,5 mm olmalıdır. Bu uzunluk, farklı hayvan türleri ve yaşları için ayarlanması gerekiyor. </li><li> Kıvrık ucu ile aynı yönde (Şekil 1 iğne göstergesi) üst ucuna iğne küçük bir bakır tel veya küçük bir parça bant takın. Tel veya bant alttan ameliyat sırasında iğne dönme açısı (Şekil 1) gösterir. </li></ol> 2. Hayvan hazırlık <ol><li> Tüm cerrahi aletler, cam boncuk sterilizatör, veya% 70 etanol ile dezenfekte ile sterilize otoklava gerekir. </li><li> Postnatal yaş 20 - 22 gün (P20-22) ya da aynı yaşta CD1 fareler, Sprague / Dawley rat kullanır. , Fareler ya da sıçanlar farklı türlerin belirli projeler için kullanılıyor olabilir. </li><li> Fareyi bir ip ketamin enjeksiyonu ile anestezi 80 mg / kg ve ksilizin 8 mg / kg. Hayvan kuyruk tutam yanıt vermiyor sonra cerrahi prosedür başlar. Ameliyat sırasında, gerekirse, bir rapel doz üçte birinin orijinal doz anestezik kokteyl orijinal anestezi devlet geri verilecektir. </li><li> Elektrikli saç kesme makineleri ile hayvan kafa derisi, saç kesin. Anestezi sırasında kuruluğa karşı koruma için bir hayvanın gözleri üzerine küçük bir miktar oftalmik merhem uygulayın. Kafa derisi,% 70 etanol,% 10 povidon-iyot çözeltisi ile dezenfekte edin. </li><li> Hayvan istikrarlı bir sabit pozisyonda kafa tutmak için stereotaksik bir aparat üzerine monte edin. Ameliyat boyunca, hipotermi önlemek için ısıtma pedi üzerinde hayvan tutun. </li><li> Lambda gelen gözler arasında uzanan bir neşter kullanılarak kafa derisi ön-arka orta hat kesi olun. Hemostat deri bir kenara çekin ve sol kafatası yeterince açığa çıkarmak için kullanın. </li><li> Sol kafatası üzerinde dikdörtgen bir kesim olun ve periost kazımak için neşter kullanabilirsiniz. Bu adım kanamayı azaltmak ve kafatası delme kolaylaştıracaktır. </li></ol> 3. Undercut&#39;lar yapma <ol><li> Maruz kafatası alan üzerine küçük bir miktar steril tuzlu su ekleyin ve sonra kan, temiz ve kuru alan birkaç Q-ipuçlarını kullanın. </li><li> Cerrahi bir mikroskop altında, sol kafatasının merkezinde dikdörtgen bir oluk (sıçanlarda ~ 5 x 7 mm, 4 farelerde x 5 mm) (yaklaşık, sol sensorimotor korteks üstte) sondaj başlar. Kafatası üzerinde bir damla serum fizyolojik uygulanması, sondaj ve dağıtır ısı kolaylaştırır. Kemik derinliği sonra yaklaşık 2 / 3 delinir, aşırı tuzlu su çıkarmak ve Q-tip sondaj alanını temiz. Kemik merkezi parçası üzerine hareketli bir forseps nazik bir dokunuş kadar yavaş ve dikkatli derin delme. </li><li> Merkezi parça kemik, kemik kenarına forseps keskin bir ucu ekleme ve sol hemisfer yavaş yavaş ortaya çıkarmak için forseps kaldırma dikkatlice çıkarın. </li><li> Cerrahi mikroskop altında, alttan cihazı ve yönlendirmek parasagital bir yönde iğne ve iğne ucu ve hedef kortikal bölgedeki görselleştirme korumak için biraz kaudal böylece orta hatta, devirme cihaz için destek plakası dik. Kraniyal penceresinin ortasında, superior sagittal sütür yanal bir alana 1-2 mm iğne ucu Amaç, ve büyük damarların doğrudan nüfuz önlemek. </li><li> Kan damarlarının Yedek şekilde dura aracılığıyla pia altında yatay yönde iğne takın. Alttan cihazın alt Sitböylece iki kraniyal pencere kenarları üzerine iğne kortikal yüzey (Şekil 1) normal konumlandırılmış. Elle kafatası pencere kenarlarını hem cihaz Dinlenme önemli ölçüde azaltmak veya aşağıdaki yordamı sırasında el sallayarak ortadan kaldıracaktır. Pia altına iğne yukarı kaldırın, sonra yavaş yavaş iğne derin gidemez kadar transkortikal bir kesim oluşturmak için daha düşük. ~ 135 ° uzakta VI alttan yarım daire beyaz madde / derin katmanı oluşturmak için orta hat döndürün. Sonra pia altına iğne tekrar yükseltin. Cihazı geriye doğru yatırın ve iğneyi çıkartın. </li><li> İsteğe bağlı olarak, yukarıdaki prosedürü (adım 2.5) tekrar ama pencerenin lateral kenarları ek bir transkortikal kesme oluşturacak kadar iğne açmadan. Bu, daha eksiksiz bir kortikal izolasyon yaratacaktır. </li><li> Plastik film (6 x 6 mm) bir parçası koruması için kafa pencere üzerine yerleştirin ve kafa derisi sütür. Anesteziden tamamen düzelene kadar ısıtılmış bir pad üzerinde hayvan yerleştirin. </li></ol> 4. Temsilcisi sonuçları: Koronal kortikal dilim alttan cerrahi başarı onaylamak için hazırlanmış olabilir. Ameliyattan 2 hafta sonra hazırlanan dilim, transkortikal ve alttan kesikler, mikroskop düşük güç hedefi (Şekil 2) altında görülebilir. Kısmen izole korteks genellikle biraz daha ince olur ve epileptiform aktivite alanında potansiyel kayıt (Şekil 3) ile kortikal dilim çoğunluğu tespit edilebilir. Kontrast, beyaz cevher veya derin kortikal tabakalar, büyük delik oluşumu Lezyonlu korteks, ya da altında veya üstünde beyaz cevher, hatalı kesim dramatik incelmesi, daha fazla deneyler için beyin kullanılamaz yapacak. <img src="/files/ftp_upload/2840/2840fig1.jpg" alt="Şekil 1"/> Şekil 1. Alttan bir cihazın yapısı ve uygulama yol gösterici tüp (2), paslanmaz çelik ya da şeffaf plastik bir plaka (1) üzerine yapıştırılır . Bir şırınga (3) iğne ucu ~ 3 mm kılavuz tüp ve bükülmüş sokulur. Bir iğne, plastik tüp yapılmış durdurmak iğne iğne dikey hareket aralığı ~ 1.2 mm ve kullanım için sırasıyla P21 fare ve sıçanlarda ~ 1.5 mm ile sınırlıdır, böylece üst ucuna yapıştırılır. A segmentinde küçük bakır tel yönelim bükük iğne iğne göstergesi olarak kolu altında bağlanır. Not cihaz hareket ettirildiğinde ve kraniyal pencerede iki kenarı ile temas eden bir kesim pial yüzeyine paralel böylece. <img src="/files/ftp_upload/2840/2840fig2.jpg" alt="Şekil 2"/> Şekil 2. Alttan dilim bir temsilcisi görüntü. P48 sıçan alttan lezyon iki hafta sonra hazırlanan bir dilim A. Floresan görüntü. Kesme yara, floresan boya DII etiketli oldu. Sağ tarafta beyaz oklar transkortikal kesim gösteren ve alt okları katmanlı IV ve beyaz madde arasındaki sınır rağmen geçti alttan göstermektedir. <img src="/files/ftp_upload/2840/2840fig3.jpg" alt="Şekil 3"/> Şekil 3. Alan potansiyeli alttan alttan bir beyin dilim dilim Alan potansiyel kayıt kayıt yaralı kortikal doku hipereksitabilite düşündüren, epileptiform aktivite sergiledi.

<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>

Çıkar çatışması ilan etti.

Alttan model posttravmatik epileptogenezisin eğitimi için son derece etkili bir sistem. Tipik bir cerrahi bitirmek için sadece yaklaşık 20-30 dakika sürer ve uyarılmış veya cerrahi 1-2 iki hafta sonra spontan epileptiform aktivite çoğu hayvanlardan dilimleri kaydedilebilir. Daha da önemlisi, bu model, travmatik beyin hasarı, kanama, inflamasyon, ödem, axotomy ve nöronal ölüm 7 gibi aşağıdaki değişikliklerden yönlerini taklit eder. Kemirgenler ve diğer hayvanlardan sadece epi...

<table border="1"><tbody><tr><td> Malzeme Adı </td><td> Tip </td><td> Şirket </td><td> Katalog Numarası </td></tr><tr><td> Foredom mikromotor kiti </td><td> ekipman </td><td> Foredom </td><td> K.1070 </td></tr><tr><td> 1.5 inç 22-gauge enjektör iğnesi </td><td> malzeme </td><td> BD şirket </td><td> 305156 </td></tr><tr><td> 1.5 inç 25-gauge şırınga iğnesinin </td><td> malzeme </td><td> BD şirket </td><td> 305127 </td></tr><tr><td> Siyanoakrilat yapıştırıcı </td><td> malzeme </td><td> Ted Pella </td><td> 14450 </td></tr></tbody></table>

preparing undercut model of posttraumatic epileptogenesis in rodents

Kısmen izole korteks (&quot;alttan&quot;), travma sonrası epileptogenezisin bir hayvan modeli. Cerrahi işlem sensorimotor korteks ve serebral korteksin belirli bir bölgeye komşu korteks ve subkortikal bölgelerde 1-3 büyük ölçüde izole edilmiş, böylece beyaz madde (alttan) altında. Üzerinden kesme içerir. Epileptiform deşarjlar, ameliyatı takip eden iki hafta veya daha fazla bir gecikme sonra kemirgenler beyin dilimleri 1; ve elektrik veya davranış nöbet gibi kedi ve maymun 4-6 gibi diğer türler in vivo olarak görülebilir. Bu iyi kurulmuş bir hayvan modeli oluşturmak için etkili ve travmatik beyin hasarı birçok önemli özelliklerini taklit ederek. Ancak, teknik olarak ücretsiz bir el ile küçük kemirgen beyin hassas kortikal lezyonlar haline getirmeye çalışıyor zordur. Başlangıçta, Stanford Üniversitesi&#39;nden 1 Dr David Prince laboratuarda kurulan prosedürü dayanarak, burada fare ve sıçanlarda bu modelin hazırlanması için bir ameliyat gerçekleştirmek için geliştirilmiş bir teknik mevcut . Biz basit bir cerrahi cihaz ve daha kesin ve tutarlı sonuçlar üretmek için derinlik ve açı kesme daha iyi bir kontrol elde etmek için kullanmak için nasıl göstermektedir. Cihaz yapmak kolaydır ve prosedürü öğrenmek için hızlı. Bu hayvan modeli nesil posttravmatik epileptogenezisin mekanizmaları için verimli bir çalışma sistemi sağlar.

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

Kısmen izole korteks (&quot;alttan&quot;), travma sonrası epileptogenezisin verimli bir hayvan modeli. Burada nasıl bir roman cerrahi cihaz yapmak ve bu model oluşturmak için daha kesin ve tutarlı lezyonlar yapmak için kullanabilirsiniz göstermektedir.

Kemirgenler Travma Sonrası epileptogenezisin Alt kesilme Model hazırlanması

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

Research

JoVE Journal

Neuroscience

9.3K Görüntüleme.  Indiana University School of Medicine. Kısmen izole korteks ("alttan"), travma sonrası epileptogenezisin verimli bir hayvan modeli. Burada nasıl bir roman cerrahi cihaz yapmak ve bu model oluşturmak için daha kesin ve tutarlı lezyonlar yapmak için kullanabilirsiniz göstermektedir.

Video: 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.

Kemirgenler Akustik irkilme alışkanlık ve prepulse inhibisyonu

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

Nörobilim

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.

Kemirgenler davranmak yılında Hareketli Silikon Problar tarafından Nöronlar Büyük ölçekli Kaydı

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.

Kemirgenler Davranış Odyometri için Düşük Maliyetli Kurulum

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.

Kemirgenler Faaliyet teker Running kayıt ve Sirkadiyen Rhythms Analizi

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.

Kemirgenler Eşzamanlı fMRI ile derin beyin stimülasyonu

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.

Kemirgenler Akrilik Reçine Kalıp Esaslı Başkanı Fixation Tekniği

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.

Kemirgenler dorsal kök gangliyon Omurilik ve İzolasyon hidrolik eldesi

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.

Transkraniyal elektrik beyin stimülasyonu uyarı Rodents

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.

Kanül implantasyon Sisterna Magna kemirgenler içine

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.

Kafa içi farmakoterapi ve ağrı deneyleri Rodents

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