This research was supported by Basic Science Research Program through the NRF funded by the Ministry of Education, Science and Technology (2010-0022362) and (2012R1A2A2A02046812).

Etik Beyanı: Bu deney Kyungpook Ulusal Üniversitesi Kurumsal Hayvan Bakım ve Kullanım Kurulu tarafından onaylandı ve Bakımı ve Kullanımı Laboratuvar Hayvanları (Ulusal Sağlık Enstitüsü, 1996) Kılavuzu göre yapıldı. Deneyden sonra, karbon dioksit ötenazi altında kurban edilmiştir. Sıçanlar için başlıklar tespit Bar 1. Tasarım NOT: Bu adım, bir tespit çubuğu tasarımı için genel bir kılavuzdur. Boyut ve malzeme ile ilgili herhangi bir değişiklik hayvanlar veya implantlar bağlı mümkündür. CAD yazılımı kullanarak bir kafa sabitleme çubuğunu Tasarımı önerilir, ancak gerekli değildir. <ol><li> Bir tespit çubuğu tasarlamak için CAD yazılımı kullanın. <ol><li> Bir dikdörtgen şeklinde (123 × 35 × 6 mm) (Şekil 1A) bir tespit çubuğu tasarlayın. Tespit çubuğunun ortasında, dikdörtgen bir çentik (port 16 × 20 mm) emin olun. Yamaçlarında ya da girintileri ekle yüklenebileceğinih (Şekil 1B -arrows Şekil 1A-) liman sınırları üzerinde kılavuz rayı olarak hizmet vermektedir. NOT: limanın alan kraniotomi yoluyla beyin bölgelerinin çoğunda bir stereotaksik manipülatör kullanarak dikey erişimlerini sağlamak için beynin üzerine sıçan kafatası tahmini dikdörtgen kapsar emin olun. </li><li> Sabitleme çubuğunun her iki tarafta iki uzatılmış sabitleme delikleri (50 x 20 x 6 mm) plakaları takın. Bu delikler 6 mm çaplı (Şekil 1A-d) vidalarını geçmesi emin olun. Bir kilitleme kancası (Şekil 1A-B) bağlantı noktasının önünde çapı 3 mm bir delik açın. </li><li> Bir kutu bir zemin (135 x 305 mm) ve yan duvarları (305 x 76 mm) sahip olun. Şekil 1A (ek) &#39;de gösterildiği gibi, iki 6 mm çaplı delik delin. Sabitleme bar (Şekil 1A-e) destek duvarları ekleyin. </li></ol></li><li> Tasarlanmış bir bar imalatı ve kutu gibi siyanoakrilat kullanılarak her biri bir arayaŞekil 1A (ek) gösterilmiştir. 6 mm çapında vidalar kullanılarak kutusunu çubuğu birleştirin. Ergonomik kafa sabitleme (Şekil 1A-ek) için çubuğun vidalı pozisyonunu ayarlayarak eğimi uygulayın. NOT: Fare için, sabitleme çubuğunun boyutu (2 mm kalınlığında akrilik paneller iki katmandan oluşan), 60 x 20 x 4 mm &#39;dir. Port 8 mm genişliğinde (üst) ve Şekil 1C &#39;de gösterildiği gibi 9 x 10 mm (alt) kare olduğunu. Ankraj plakaları fareler &#39;aynı boyuttadır. Kutu plaka ve yan duvarlar sırasıyla 100 × 72 mm ve 100 × 50 mm, vardır. </li></ol> İmplantasyon Cerrahisi sırasında 2. Başkanı Mounter İnşaat DİKKAT: Diş akrilik reçine ile uğraşırken dikkatli olun. Sıvı cilt tahrişine neden olabilir. Bir maske koyun ve eldiven giyin. Iyi havalandırılan bir alanda çalışın. <ol><li> Ameliyattan önce hazırlık <ol><li> Sıkıca tespit çubuğu kullanarak ap limanı sınır sarınbara uygulanan akrilik reçine yapışmasını önlemek için araffin film (Şekil 1B ve Şekil 2A &#39;ya bakınız). Kat akrilik reçine ince bir tabaka ile parafin film kaplı portun yatılı. </li><li> Sterilize cerrahi aletler kullanın. İyice stereotaksik çerçeve dezenfekte ve onun% 70 etil alkol kullanarak çevredeki. </li></ol></li><li> Stereotaksik implantasyon ameliyatı <ol><li> Intraperitonal ketamin 10 ml kokteyl karışımının enjeksiyonu (2 ml / kg) (50 mg / ml), ksilazin hidroklorür 1.5 ml (23.32 mg / ml) ve tuzlu su 2.5 mi, bir hayvan anestezisi. Hayvan tamamen uyuşturulduktan ve kuyruk ve ayak tutam yanıt vermiyor emin olun. Gerektiğinde, ameliyat sırasında, 1 ml / kg bir kokteyl karışımı enjekte edilir. </li><li> Stereotaksik aparat hayvan sabitleyin. Ameliyat sırasında kuruluğunu önlemek için gözleri oftalmik merhem sürün. Tıraş ve% 70 etanol ile kafa derisi temizleyin. % 2 lidokain subkutan enjeksiyon vermek ve SCA insizyonKafatasının orta hat boyunca lp. Tamamen periost kazıyın. Tuzlu birkaç kez durulayın. </li><li> Kafatasının kenarına yakın Burr-hole sondaj yapın. İmplant vidaları kafatası yüzeyinden 4-5 tur (1 mm kadar derinlik) dura mater ihlal önlemek için. Vida implantasyon sonrası kafatası yüzeyinde CSF ve kan emin hiçbir sızıntı olun. Tuzlu su ile durulayın ve özellikle vida etrafında, hava puf kullanarak kafatası yüzeyi kurulayın. </li><li> &quot;Dental yapıştırıcı (Süper Bond C &amp; B)&quot; ince bir tabaka ile kaplayın kafatası yüzeyi. Tamamen ek elektrot implantasyonu planlanan eğer bregma dahil kafatasının yüzeyini örtmeyin. Kraniyotomi koordinatları işaretleme için bregma kullanın. İmplant elektrotlar ve / veya Microdrive ve çevresinde akrilik reçine uygulayarak kafatası sabitleyin. </li><li> Diş çimentosu en ek akrilik reçine uygulanır. Bar kaplı reçine arasında yeterli dolgu-out boşluk bırakmak için akrilik reçine uygun miktarda kullanınd İlk montaj reçine vida ve / veya elektrotların (Şekil 2A) sabitleme zaman. </li></ol></li><li> Baş-sabitleme çubuğunu kafatası yüzeyi üzerinde 5 mm veya daha fazla koyun ve merkezi (Şekil 2A -Sol) hizalayın. </li><li> Kafa tabanı ve ilk montaj reçineler arasındaki boşluğa akrilik reçine uygulayın. (Şekil 2A -Center). Sert olmak reçine kadar bekleyin. </li><li> Kulak-bar çıkarın ve sabitleme çubuğu liman dışında kalıp Mounter çekin. (Şekil 2A -Sağ). Dikkatle kalıplanmış mounter gerektiğinde kenar rayları etrafında ek bir akrilik reçine uygulanır. </li><li> Steril serum fizyolojik ile kafa derisi yara yüzeyini durulayın. Ameliyattan hemen sonra bir antibiyotik (enrofloksasin, 5 mg / kg sc) ve bir analjezik (butorfanol tartrat, 2 mg / kg sc) enjekte edilir. Tek bir kafeste tek hayvan barındıracak. </li></ol> 3. alıştırmak Hayvanlar Baş-tespit etmek Not: Bu adım DEYazarların ampirik başarılı değerlendirmelere dayalı alışkanlık programları scribes. Baş-sabitleme için alışkanlık prosedürünün genel bir bakış için, Schwartz&#39;ın 10 veya Guo&#39;nun 11 iş görmek. <ol><li> Hayvanlara yeterli iyileşme süresini (bir hafta veya daha fazla) verin. Gün 1, sık sık bölünmüş kısa oturumlarında hayvanları taşıyınız. </li><li> Ertesi gün (Gün 2) üzerinde sabitleme ortamı (kutu) hayvan maruz bırakmaktadır. Tespit çubuğunun portuna onların baş Mounter takmayın. Çevreye alışması için hayvanları bırakın. Güvenli 1 ila 2 saniye elinizi kullanarak kafa Mounter kapmak ve hemen bırakın. El-kapmak birkaç kez kısa süreli fiksasyon tekrarlayın. </li><li> Gün 3, baş-tespitin hayvanları alıştırarak. <ol><li> Ev kafeslerden hayvanları getirin ve sabitleme çubuğu limanına doğru başlarını yönetmenlik kutuya yerleşmek. Baş Mounter tut ve tespit çubuğunun bağlantı noktasına takın. </li><li> Hayvan en (Mounter Bindbir lastik bant kullanarak tespit çubuğunun kilitleme kancasına baş). 5 dakika bekleyin. NOT: gıcırtı ve hayvanın strese artan mücadele hayvan açar tespit sonrasında hayvanın gövde veya kuyruk dokunmayın. Hayvan fazla 1 dk mücadele devam ederse, bırakın ve daha sonra tekrar deneyin. Tespit kafa maruz ancak bu yavaş yavaş alışkanlık ilerledikçe azaltılmış olur zaman bu adımı itibaren, hayvanlar idrar ve dışkı atılımı göstermektedir. </li><li> Kilitleme lastik bant alın ve liman dışında Mounter çıkarın. Eve kafesine hayvan koyun ve tahıl bir parça veya bir ödül olarak ev kafesin içine bir ayçiçeği tohumu sağlamaktadır. Bir sonraki duruşma 30 dakika başlar. Bir gün içinde 3.4.1 beş 3.4.4 ya da daha fazla tekrarlayın. </li></ol></li><li> 7 Gün 4, Protokolün 3.4 anlatıldığı gibi tespit kafa hayvan maruz. Yavaş yavaş (10 dk, 20 dk, 30 dk, ve alışkanlık ilerledikçe 50 dakika) baş sabitleme süresini artırır. NOT: Hayvanlar yaparsanız hiçbirt stres düzeyi yüksek bir işareti olabilir, gıda ödül yem. Fiksasyon süresini artırmak etmeyin. Hayvanlar davranışı mücadele tekrarlanan göstermek ise, daha uzun bir süre onları alıştırmak. </li><li> Günlük hayvanın vücut ağırlığını kontrol edin. Vücut ağırlığı azalıyor başarısız alışkanlık düşünün. Başarılı alışkanlık sonra baş sabitleme ile davranışsal görevlere devam edin. NOT: Başarılı alışkanlık sonraki davranış eğitimi (Şekil 3E ve F) kolaylaştırır. </li></ol>

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The authors have nothing to disclose.

This protocol demonstrates a simple and convenient head fixation method using materials at hand. For physically successful head fixation, the first critical steps during surgery is to rinse and dry the skull surface clearly before applying the dental adhesive resin cement. Second, this method does not uses screws implanting under temporal muscles and only depends on the screws on the skull, their perpendicular implantation against the surface of skull rim is crucial10. This reduces the possibility of accidenta...

Kafa tespit veya kafa kısıtlama sert sıkma için kafatası üzerinde bir çerçeve ya da plaka takarak hayvanın başını hareketsizleştirir bir tekniktir. Teknik, hassas duyusal stimülasyon, ya da uyanık hayvanlarda 1-3 nöral kayıt olmadan ve uyku-uyanıklık döngüsü 4 sırasında juxtacellular etiketleme için davranışsal algılama için kabul edilmiştir. Aynı zamanda, in vivo beyin görüntüleme 5,6 kullanılır. Şimdi kafa sabitleme tekniği nörobilim ve davranışsal araştırmalarda yaygın olarak kullanılan bir araçtır. Temelde, geleneksel kafa sabitleme kafatası 6-8 üstündeki metal baş mesajların cerrahi eki gerektirir. Ek parça daha sonra baş immobilizasyonu ile sonuçlanan sabit bir platforma kenetlenir. Böyle nöral ve / veya optik kayıt, beyin devreleri müdahale olarak uyanık hayvanlarda nörofizyolojik araştırma, beyin görüntüleme diş akrilik kullanarak implantasyon cerrahisi eşlik ediyor. Bu nedenle,o deneysel gerektiğinde bu laboratuvarlarda kafa tespiti uygulamak için basit bir görev gibi görünüyor. Çerçeve evlerinde kafeslerde hayvanın doğal davranışlarını rahatsız değil, hafif ve yeterince küçük olması dizayn edilmelidir. Sabit platforma kenetlenmiş zaman da mekanik sertlik sağlar. Ayrıca deneysel amaçlarla bağlı biyouyumluluk vardır. Bu nedenle, tasarım ve genel laboratuar ekipmanları gibi baş mesajları imal etmek teknik olarak zordur. Bu çalışmada, yeni bir baş tespit yöntemi, metal kafa Mesajları kullanmadan uygun sıkıştırma yoluyla mekanik sertlik sağlarken, tarif edilmektedir. Yöntemi kullanarak, kafa sabitleme implantasyonu genellikle beyin çalışma için kemirgenlerde gerçekleştirilen genel laboratuar koşulları altında yapılabilir. Metal baş mesajlar implan sırasında yapılan bir ısmarlama kafa sabitleme çubuğu ve kalıplama işlemi kısmen kalıplı akrilik reçine çimento ile değiştirilirtation cerrahisi. Sabitleme çubuğu gibi nöral kayıt sondaları, stimülasyon elektrotları gibi diğer implantlar, yüksek uzaysal uyumluluk sağlar. Ayrıca vidalı bağlama kaldırarak sabitleme platformunda hayvanın kafasını sıkma adımlarını kolaylaştırır. Biz önceki davranışsal çalışmada bu yöntem kullanılır ve bu yöntem pratik deneyler 9 uygulamak kolay ve rahat olduğu sonucuna varıldı.

<table border="0" cellpadding="0" cellspacing="0" width="901">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Dental acrylic resin</td>
			<td style="width:199px;">Lang Dental Manufacturing</td>
			<td style="width:129px;">Ortho-Jet</td>
			<td style="width:343px;">to make molded mounter</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Paraffin film</td>
			<td style="width:199px;">Bemis</td>
			<td style="width:129px;">Parafilm</td>
			<td>to prevent adhesion of acrylic resin to the bar</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Dental adhesive</td>
			<td style="width:199px;">Sun Medical</td>
			<td style="width:129px;">Super-Bond C&#38;B</td>
			<td>to bond between acrylic resin and skull</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Plexglass or other acrylic plate</td>
			<td style="width:199px;"></td>
			<td style="width:129px;"></td>
			<td>to make fixation bar and box platform</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:231px;">Cyanoacrylate</td>
			<td style="width:199px;"></td>
			<td style="width:129px;"></td>
			<td>to bond acrylic plates</td>
		</tr>
	</tbody>
</table>

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.

Bu kafa sabitleme yöntemi, farelerde ve sıçanlarda da uygulanabilir. Toplam sekiz sıçan medyal ön beyin demetine uyarıcı elektrot bilateral implante aldı. Bunların dördünde için özelleştirilmiş tungsten tel elektrotlar (Microdrives) ek M1 ön ayakları alanına (Şekil 2C) implante edildi. Fareler (n = 2), dört adet paslanmaz çelik tel elektrot frontal / Oksipital kortikal alanlara implante edildi. Ameliyattan çıktıktan sonra Protokol bölümünde 2. tarif olarak Ameliyat sırasınd...

Watch this Scientific Journal Video about Acrylic Resin Molding Based Head Fixation Technique in Rodents at JoVE.com

Acrylic Resin Molding Based Head Fixation Technique in Rodents

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 ...

acrylic-resin-molding-based-head-fixation-technique-in-rodents

Research

JoVE Journal

Neuroscience

11.7K Görüntüleme.  Kyungpook National University. 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.

Video: Acrylic Resin Molding Based Head Fixation Technique in Rodents

Preparing Undercut Model of Posttraumatic Epileptogenesis in Rodents

Partially isolated cortex ("undercut") is an animal model of posttraumatic epileptogenesis. The surgical procedure involves cutting through the sensorimotor cortex and the underneath white matter (undercut) so that a specific region of the cerebral cortex is largely isolated from the neighboring cortex and subcortical regions1-3. After a latency of two or more weeks following the surgery, epileptiform discharges can be recorded in brain slices from rodents1; and electrical or behavior seizures can be observed in vivo from other species such as cat and monkey4-6. This well established animal model is efficient to generate and mimics several important characteristics of traumatic brain injury. However, it is technically challenging attempting to make precise cortical lesions in the small rodent brain with a free hand. Based on the procedure initially established in Dr. David Prince's lab at the Stanford University1, here we present an improved technique to perform a surgery for the preparation of this model in mice and rats. We demonstrate how to make a simple surgical device and use it to gain a better control of cutting depth and angle to generate more precise and consistent results. The device is easy to make, and the procedure is quick to learn. The generation of this animal model provides an efficient system for study on the mechanisms of posttraumatic epileptogenesis.

Partially isolated cortex (&#x201C;undercut&#x201D;) is an efficient animal model of posttraumatic epileptogenesis. Here we demonstrate how to make a novel surgical device and use it to make more precise and consistent lesions to generate this model.

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 ...

Nörobilim

Whole Animal Perfusion Fixation for Rodents

The goal of fixation is to rapidly and uniformly preserve tissue in a life-like state. While placing tissue directly in fixative works well for small pieces of tissue, larger specimens like the intact brain pose a problem for immersion fixation because the fixative does not reach all regions of the tissue at the same rate 5,7. Often, changes in response to hypoxia begin before the tissue can be preserved 12. The advantage of directly perfusing fixative through the circulatory system is that the chemical can quickly reach every corner of the organism using the natural vascular network. In order to utilize the circulatory system most effectively, care must be taken to match physiological pressures 3. It is important to note that physiological pressures are dependent on the species used. Techniques for perfusion fixation vary depending on the tissue to be fixed and how the tissue will be processed following fixation. In this video, we describe a low-cost, rapid, controlled and uniform fixation procedure using 4% paraformaldehyde perfused via the vascular system: through the heart of the rat to obtain the best possible preservation of the brain for immunohistochemistry. The main advantage of this technique (vs. gravity-fed systems) is that the circulatory system is utilized most effectively.

Here we describe a low-cost, rapid, controlled and uniform fixation procedure using 4% paraformaldehyde perfused via the vascular system: through the heart of the rat to obtain the best possible preservation of the brain.

Kemirgenler için Tüm Hayvan Perfüzyon Fiksasyon

The goal of fixation is to rapidly and uniformly preserve tissue in a life-like state. While placing tissue directly in fixative works well for small ...

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 ...

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, ...

Transplantation of Olfactory Ensheathing Cells to Evaluate Functional Recovery after Peripheral Nerve Injury

Olfactory ensheathing cells (OECs) are neural crest cells which allow growth and regrowth of the primary olfactory neurons. Indeed, the primary olfactory system is characterized by its ability to give rise to new neurons even in adult animals. This particular ability is partly due to the presence of OECs which create a favorable microenvironment for neurogenesis. This property of OECs has been used for cellular transplantation such as in spinal cord injury models. Although the peripheral nervous system has a greater capacity to regenerate after nerve injury than the central nervous system, complete sections induce misrouting during axonal regrowth in particular after facial of laryngeal nerve transection. Specifically, full sectioning of the recurrent laryngeal nerve (RLN) induces aberrant axonal regrowth resulting in synkinesis of the vocal cords. In this specific model, we showed that OECs transplantation efficiently increases axonal regrowth.
OECs are constituted of several subpopulations present in both the olfactory mucosa (OM-OECs) and the olfactory bulbs (OB-OECs). We present here a model of cellular transplantation based on the use of these different subpopulations of OECs in a RLN injury model. Using this paradigm, primary cultures of OB-OECs and OM-OECs were transplanted in Matrigel after section and anastomosis of the RLN. Two months after surgery, we evaluated transplanted animals by complementary analyses based on videolaryngoscopy, electromyography (EMG), and histological studies. First, videolaryngoscopy allowed us to evaluate laryngeal functions, in particular muscular cocontractions phenomena. Then, EMG analyses demonstrated richness and synchronization of muscular activities. Finally, histological studies based on toluidine blue staining allowed the quantification of the number and profile of myelinated fibers.
All together, we describe here how to isolate, culture, identify and transplant OECs from OM and OB after RLN section-anastomosis and how to evaluate and analyze the efficiency of these transplanted cells on axonal regrowth and laryngeal functions.

Olfactory ensheathing cells (OECs) are neural crest cells which allow growth of the primary olfactory neurons. This specific property can be used for cellular transplantation. We present here a model of cellular transplantation based on the use of OECs in a laryngeal nerve injury model.

Periferik Sinir Hasarı Sonrası Fonksiyonel Kurtarma Değerlendirmesinde Koku Ensheathing Hücre Nakli

Olfactory ensheathing cells (OECs) are neural crest cells which allow growth and regrowth of the primary olfactory neurons. Indeed, the primary olfactory ...

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 ...

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 ...

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 ...

The Power of Interstimulus Interval for the Assessment of Temporal Processing in Rodents

Temporal processing deficits have been implicated as a potential elemental dimension of higher-level cognitive processes, commonly observed in neurocognitive disorders. Despite the popularization of prepulse inhibition (PPI) in recent years, many current protocols promote using a percent of control measure, thereby precluding the assessment of temporal processing. The present study used cross-modal PPI and gap prepulse inhibition (gap-PPI) to demonstrate the benefits of employing a range of interstimulus intervals (ISIs) to delineate effects of sensory modality, psychostimulant exposure, and age. Assessment of sensory modality, psychostimulant exposure, and age reveals the utility of an approach varying the interstimulus interval (ISI) to establish the shape of the ISI function, including increases (sharper curve inflections) or decreases (flattening of the response amplitude curve) in startle amplitude. Additionally, shifts in peak response inhibition, suggestive of a differential sensitivity to the manipulation of ISI, are often revealed. Thus, the systematic manipulation of ISI affords a critical opportunity to evaluate temporal processing, which may reveal the underlying neural mechanisms involved in neurocognitive disorders.

Temporal processing, a preattentive process, may underlie deficits in higher-level cognitive processes, including attention, commonly observed in neurocognitive disorders. Using prepulse inhibition as an exemplar paradigm, we present a protocol for manipulating interstimulus interval (ISI) to establish the shape of the ISI function to provide an assessment of temporal processing.

Interstimulus Aralık kemirgenler zamansal işleme değerlendirilmesi için güç

Temporal processing deficits have been implicated as a potential elemental dimension of higher-level cognitive processes, commonly observed in ...

A Revised Surgical Approach to Induce Endolymphatic Hydrops in the Guinea Pig

Endolymphatic hydrops is an enlargement of scala media that is most often associated with Meniere&#39;s disease, though the pathophysiologic mechanism(s) remain unclear. In order to adequately study the attributes of endolymphatic hydrops, such as the origins of low-frequency hearing loss, a reliable model is needed. The guinea pig is a&#160;good model because it hears in the low-frequency regions that are putatively affected by endolymphatic hydrops. Previous research has demonstrated that endolymphatic hydrops can be induced surgically via intradural or extradural approaches that involve drilling on the endolymphatic duct and sac. However, whether it was possible to create an endolymphatic hydrops model using an extradural approach that avoided dangerous drilling on the endolymphatic duct and sac was unknown. The objective of this study was to demonstrate a revised extradural approach to induce experimental endolymphatic hydrops at 30 days post-operatively by obliterating the endolymphatic sac and injuring the endolymphatic duct with a fine pick. The sample size consisted of seven guinea pigs. Functional measurements of hearing were made and temporal bones were subsequently harvested for histologic analysis. The approach had a success rate of 86% in achieving endolymphatic hydrops. The risk of cerebrospinal fluid leak was minimal. No perioperative deaths or injuries to the posterior semicircular canal occurred in the sample. The presented method demonstrates a safe and reliable way to induce endolymphatic hydrops at a relatively quick time point of 30 days. The clinical implications are that the presented method provides a reliable model to further explore the origins of low-frequency hearing loss that can be associated endolymphatic hydrops.

This article demonstrates an extradural approach to obliterate the guinea pig endolymphatic sac and injure the endolymphatic duct with a fine pick in order to induce experimental endolymphatic hydrops.

Kobayda Endolenfatik Hidrops İndüklemek için Gözden Geçirilmiş Cerrahi Yaklaşım

Endolymphatic hydrops is an enlargement of scala media that is most often associated with Meniere&#39;s disease, though the pathophysiologic mechanism(s) ...