Supported by grants from CIHR (MOP231122) and NSERC (RGPIN-2014-06503). We would like to thank Genevi&#232;ve Cyr for her help preparing experiments and supervising laboratory work. MAL received a scholarship from The Natural Sciences and Engineering Research Council of Canada (NSERC).

NOT: Tüm prosedürler Hayvanları Koruma ve Université de Montréal Etik inceleme kurulu Kanada Konseyi direktifleri doğrultusunda yapıldı. Kayıt enjeksiyon Pipet 1. Meclisi <ol><li> Bir pipet çektirmenin kullanarak yaklaşık 7 cm uzunluğunda bir cam kapiller (1 mm dış çapı) çekin. </li><li> Kılcal ucu kırın ve bir ışık mikroskobu altında diyafram kontrol edin. Iç çapı 40 mikron 30 mikron arasında olduğunu onaylayın. </li><li> Cam pipet olmayan konik ucundan çıkıntılı yaklaşık 1 cm cam kapiller içine 7 cm uzunluğunda gümüş tel yerleştirin. </li><li> Cam kapiller dikey olarak aşırı filament bükün. </li><li> Bir 30 G hipodermik iğnenin mil üzerinde esnek plastik bir yapışkan damlacık uygulanır. </li><li> Şekil 1 &#39;de sunulan şemadaki gibi cam pipet bir 30 G hipodermik iğne takın </stro&gt; ng. </li><li> Cam pipet ve hipodermik iğne arasındaki birleşme gelen uygun bir sızdırmazlık sağlamak için yapıştırıcı bir ikinci kaplama ekleyin. </li><li> Ucu tutkal uygun kür sağlamak için yaklaşık 12 saat boyunca yukarı bakacak şekilde kurumaya pipet bırakın. bitmiş sonucu, Şekil 2&#39;de gösterilmiştir. </li></ol> Not: Bu yordam, akut deneyler ve pipet sterilizasyonu yapılır gerekli değildir. 2. Hayvan hazırlanması <ol><li> Bir anestezi kutusuna sıçan yerleştirin. </li><li> 5 ila 10 dakika süreyle% 4 izofluran kullanarak anestezi neden olur. </li><li> 37 ° C&#39;lik bir vücut ısısını korumak için ısıtma yastığı ve rektal prob ile bir stereotaksik tablo hayvan yerleştirin. % 2 izofluran ile anestezi korumak için burun konisi kullanın. Kulak çubukları ve dişlerin tutucu kullanarak sıçan başını sabitleyin. </li><li> Atropin öğrenci genişlemesi yardımcı olmak için damla% 1 dahil oftalmik merhem ya da göz damlaları, uygulayın. Kuruluğunu önlemek için, damla yağlama uygulamakYaklaşık her 30 dk s. </li><li> Kafa Tıraş ve% 10 povidon-iyot ile temizleyiniz. </li><li> Lokal anestezi için, cildi yukarı kaldırma ve iğne ucu takarak 2-3 yerlerde kafa derisi altında% 2 lidokain 0.5 ml enjekte edilir. </li><li> Bir ayak tutam performans ve hareket eksikliği gözlemleyerek anestezi yeterli düzeyde onaylayın. Buna ek olarak, normal değerler (300-400 atım / dak) içinde olduğundan emin olmak için kalp hızı izleme. </li><li> Koronal ve sagital sütür hem duyurmak için bir 10. neşter bıçağı ile medyan boyunca düz bir çizgi derisini İnsizyon. </li><li> Cerrahi spatula ile kafatası kapsayan doku bir kenara iterek Lambda ve Bregma noktaları açığa vurmak. </li><li> Bregma ve Lambda pozisyonları aynı düzlemde olacak şekilde kafatası Seviye. </li><li> Referans noktasını ayarlamak için, sağ bregmadan yukarıda ayarlamak için bir monte edilmiş cam tüp ile bir stereotaksik cihaz kullanın. Bu &quot;sıfır&quot; için ön-arka ve med olacakial yanal ölçümler koordine eder. </li><li> Stereotaksik gerekli koordinatları montaj hareket ettirerek ilgi noktasını ayarlayın stereotaksik koordinatları not ve kraniyotomi yapılacaktır nerede işaretleri hedef alan etrafında bir kare çizin. </li><li> Yavaş yavaş kemik malzemeyi çıkarmak için baskı olmadan yavaş yavaş belirgin meydanda boyunca sterilize matkapla bir cerrahi matkap kullanın. Bu korteks üzerindeki lezyonlar ısı üretmek ve neden olacağından, aynı bölgede çok uzun inebilecek dikkatli olun. </li><li> Kraniotomiye sınırlayan kemik yeterince ince eriştiğinde, dikkatlice korteks maruz cımbız ile kranial bölümü çıkarın. </li><li> Sıkça doku kuruma önlemek için yapay beyin omurilik sıvısı ile maruz korteksi sulamak. NOT: injectrode ucu nüfuz kadar sağlam olduğu Dura mater kaldırma sıçan üzerinde gereksiz olduğunu. </li></ol> 3. Doldurma ve Enjeksiyon Sistemi montajı <ol><li> DoldurmakMineral yağ ile aspirasyon ile 5-10 ul mikroenjektör. </li><li> % 0.5 Chicago Sky Blue (CSB) ile 300 uM γ-aminobutirik asit (GABA) ya da% 0.5 CSB 9% 2 lidokain çözeltisi içeren bir çözelti ile, sabit bir cam pipet ile deri altı iğne doldurun. Tuzlu su ile bütün çözümleri seyreltin. <ol><li> Bol bir madde olması halinde, normal bir şırınga kullanılarak girip hava kabarcıklarının oluşumunu önlemek için normal önlemler tekniklerini kullanır. </li><li> Daha pahalı maddeler durumunda, injectrode doldurmak için mineral yağ kullanımı ve kimyasal madde daha sonra aspirasyon ile sokulabilir. Mineral yağ ve su arasındaki yoğunluk farkı nispeten yüksek olduğu için, bu madde, sulu çözeltiler enjeksiyon için iyi bir adaydır. Not: bir boya iki sıvı arasındaki mesafeyi teyit etmek için ilave edilebilir. </li></ol></li><li> Enjeksiyon pipet doldurmak için, aspirasyonla çözeltisi ile 1 ml şırınga girip daha sonra yavaş yavaş solüsyon enjekteEnjeksiyon pipet içine. </li><li> Temiz bu alanları swabbing ve ayrıca şırınga ile yavaşça çözüm enjekte ederek kaçaklar gözlemleyerek Şekil 1&#39;de gösterilen bölgelerde sızıntı olup olmadığını dikkat ediniz. </li><li> 1 ml şırınga çıkarın. Bunu yaparken, vakum enjeksiyon pipet gelen çözüm kaldırmaz, böylece pistonu hafif bir basınç tutmak için emin olun. </li><li> Aspirasyon mineral yağ ile mikroenjektör doldurun ve dolu enjeksiyon pipet sıkıca takın, sonra dikkatlice gazlı bez ile bir araya injectrode herhangi bir aşırı çözüm silerek temizleyin. </li><li> Ucu cam pipet ucu şekillendirme küçük bir damla görmek için yeterli çok küçük bir hacim, enjekte edilerek tıkalı olmadığından emin olun. </li><li> Mikropompa sistemi üzerinde injectrode monte ve iyi sabit olduğundan emin olun. </li><li> Dikkatle hedef koordinatlarında injectrode ucunu yerleştirin ve korteks yüzeyine ucu indirin. </li><li> Yavaşça injectr düşürmekUygun anatomik koordinatları kullanarak hedef yapısına stereotaksik aparatı (bu durumda üstün colliculus) kullanarak gazel. </li><li> Doku kuruma önlemek için sıcak agar ile maruz korteksi örtün. </li></ol> 4. Enjeksiyon ve Tersinir İnaktivasyonu <ol><li> Enjeksiyonu başlatmak için 40 nl / dak ve basın Run 400 nl 800 enjekte mikroenjeksiyon pompasını ayarlayın. O başak oranı enjeksiyon sırasında azalma gösterecektir unutmayın. NOT: deney düzeneği, sinirsel aktivite GABA doğum bitiminden sonra bir saat içinde iyileşti. Herhangi bir kalibre mekanik cihaz enjeksiyonu yapmak amacıyla mikroenjeksiyon şırınga baskı uygulamak için kullanılabilir. </li><li> Elektrofizyolojik veriler kazanılmasından sonra, yerel Hayvan Etik Topluluğu tarafından onaylanmış bir yöntem kullanılarak euthanize. </li></ol>

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	<li>Martin, J. H., Ghez, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pharmacological+inactivation+in+the+analysis+of+the+central+control+of+movement.">Pharmacological inactivation in the analysis of the central control of movement.</a> Journal Of Neuroscience Methods. 86 (2), 145-159 (1999).</li><li>Ponce, C. R., Hunter, J. N., Pack, C. C., Lomber, S. G., Born, R. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contributions+of+indirect+pathways+to+visual+response+properties+in+macaque+middle+temporal+area+MT.">Contributions of indirect pathways to visual response properties in macaque middle temporal area MT.</a> The Journal Of Neuroscience The Official Journal Of The Society For Neuroscience. 31 (10), 3894-3903 (2011).</li><li>Lomber, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+advantages+and+limitations+of+permanent+or+reversible+deactivation+techniques+in+the+assessment+of+neural+function.">The advantages and limitations of permanent or reversible deactivation techniques in the assessment of neural function.</a> Journal Of Neuroscience Methods. 86 (2), 109-117 (1999).</li><li>Malpeli, J., Schiller, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+method+of+reversible+inactivation+of+small+regions+of+brain+tissue.">A method of reversible inactivation of small regions of brain tissue.</a> Journal Of Neuroscience Methods. 1 (2), 143-151 (1979).</li><li>Lomber, S. 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(46), (2010).</li><li>Hup&#233;, J., Chouvet, G., Bullier, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spatial+and+temporal+parameters+of+cortical+inactivation+by+GABA.">Spatial and temporal parameters of cortical inactivation by GABA.</a> Journal Of Neuroscience Methods. 86 (2), 129-143 (1999).</li><li>Casanova, C., McKinley, P., Molotchnikofff, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Responsiveness+of+Reorganized+Primary+Somatosensory+(SI)+Cortex+after+Local+Inactivation+of+Normal+SI+Cortex+in+Chronic+Spinal+Cats.">Responsiveness of Reorganized Primary Somatosensory (SI) Cortex after Local Inactivation of Normal SI Cortex in Chronic Spinal Cats.</a> Somatosensory &amp; Motor Research. 8 (1), 65-76 (1991).</li><li>Malpeli, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reversible+inactivation+of+subcortical+sites+by+drug+injection.">Reversible inactivation of subcortical sites by drug injection.</a> Journal Of Neuroscience Methods. 86 (2), 119-128 (1999).</li><li>Minville, K., Casanova, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spatial+frequency+processing+in+posteromedial+lateral+suprasylvian+cortex+does+not+depend+on+the+projections+from+the+striate-recipient+zone+of+the+cat&#8217;s+lateral+posterior-pulvinar+complex.">Spatial frequency processing in posteromedial lateral suprasylvian cortex does not depend on the projections from the striate-recipient zone of the cat&#8217;s lateral posterior-pulvinar complex.</a> Neuroscience. 84 (3), 699-711 (1998).</li><li>Diao, Y., Wang, Y., Xiao, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Representation+of+the+binocular+visual+field+in+the+superior+colliculus+of+the+albino+rat.">Representation of the binocular visual field in the superior colliculus of the albino rat.</a> Experimental Brain Research. 52 (1), 67-72 (1983).</li></ol>

We have nothing to disclose.

Önerilen protokol mevcut geri dönüşümlü etkisizleştirme yöntemleri kaynaklanan sorunları çözmek için tasarlanmıştır. Özellikle bu proje özellikle derin beyin yapılarında nöral aktiviteyi modüle maddelerin kimyasal microinjections için kullanılan yöntemleri rafine yönelik. Kurulum bu tür ortaya çıkan bir teknik zorluk, her iki sondalar enjeksiyon yerinde kesin kayıtları elde etmek amacıyla in vivo aynı kısıtlı alanda kolokalize edilmesi ihtiyacıdır. Bu sorun, aynı sitede Enj...

kortikal alan ve alt kortikal çekirdeklerin inaktivasyonu, çeşitli beyin yapıları 2-4 arasındaki fonksiyonel ilişkiler çalışmalarında önemlidir. Son literatür beyin yapılarının 2,5 rolünü incelemek için kayıp fonksiyon-kimyasal veya kriyojenik teknikleri istihdam vardır. Çevre doku 6,7 kollateral hasarı en aza indirerek ilaç microinjections ilgili olarak, ilaçların küçük miktarlar kontrollü bir hızda, bir beyin bölgesine uygulanabilir. Bu teknik, nöronal aktivitenin farklı farmakolojik hedeflerin etkisini incelemek için spesifik agonistleri, ters agonistleri veya antagonistleri sunmak için de kullanılabilir. Bu tür etkileri de araştırmacılar, farklı kortikal ve subkortikal yapılar arasındaki ilişkileri incelemek için izin uzak yerlerden nöronal yanıtları değişiklikleri ölçerek ele alınabilir. Burada, biz bo yeteneğine sahip bir cihazın montajını, injectrode, göstermekelektrofizyolojik sinyaller kayıt ve hedef konumda ilaçların küçük miktarlarda teslim inci. Biz sıçan üstün colliculus, GABA, nöronal aktivitenin ortak bir inhibitörü enjekte edilerek bu sistemin yeteneklerini göstermek. Bu bölge bize injectrode yerelleştirme onaylamak için görsel uyarılmış multiunit aktiviteyi kullanmak için izin görsel uyarı, duyarlıdır. inaktivasyon reversibilitesinin GABA enjeksiyonu bitimini izleyen, normal nöronal aktivitenin iyileşme ile değerlendirildi. enjeksiyon bölgesinden çok üniteli etkinliğini izlemek için yeteneği istenen farmakodinamik yanıtı elde etmek için gerekli enjeksiyon oranları ve hacimlerin ince ayar için izin verir. En küçük etkili miktarlar enjekte edilir çünkü Bu nedenle, bu tekniğin bir avantajı, Şekilde, mikro neden olduğu doku hasarının sınırlayıcı potansiyelidir. önerilen protokol tek bir donanım necessar üretilmesi için bir maliyet açısından verimli bir yöntem sağlarilaç verme ve yerel nöronal etkinliği kayıt istendiği deneyleri yürütmek için y.

<table><tbody><tr><td>Injection pump (UltraMicroPump III)</td><td>WPI</td><td>#UMP3</td><td></td></tr><tr><td>Injection console (Micro4 Controller)</td><td>WPI</td><td>#SYS-MICRO4</td><td></td></tr><tr><td>Hamilton syringe</td><td>Hamliton</td><td>(80301) 701LT 10 &amp;micro;L SYR</td><td>Syringes between 5 and 10 &amp;mu;l used</td></tr><tr><td>Flexible plastic adhesive</td><td>Lepage</td><td>393915</td><td>The gel form is easier to apply to the shaft of the 30&amp;nbsp;G hypodermic needle forming a waterproof seal when dry.</td></tr><tr><td>Glass pipettes</td><td>WPI</td><td>#TW100F-4</td><td>Thin wall, 1&amp;nbsp;mm OD, 0.75&amp;nbsp;mm ID with filament pipettes used</td></tr><tr><td>720 Needle Pipette Puller</td><td>Kopf</td><td>720</td><td></td></tr><tr><td>Silver wire</td><td>A-M Systems, Inc.</td><td>782500</td><td>Bare 0.010&amp;rdquo;</td></tr></tbody></table>

simultaneous electrophysiological recording and micro-injections of inhibitory agents in the rodent brain

Burada ticari olarak erişilebilir ve uygun fiyatlı parçalar kullanarak tek kullanımlık &quot;injectrode&quot; yapımı için bir yöntem açıklanmaktadır. Bir tarama sisteminin etkilenen nöronal popülasyonundan elektrofizyolojik sinyalleri kayıt sırasında bir ilacın enjeksiyonu sağlar geliştirildi. Bu yöntem, ticari çözümler için basit ve ekonomik bir alternatif sağlar. Bir cam pipet bir hipodermik iğne ve gümüş filament ile birleştirilmesi suretiyle modifiye edildi. injectrode ilaç iletimi için ticari bir mikroenjektör pompasına bağlanmıştır. Bu ilaç dağıtım sitesinden kaynaklanan çoklu ünite dışı sinyalleri aracılığıyla gerçek zamanlı farmakodinamikler geri bildirim sağlayan teknikle sonuçlanır. Kavramının bir kanıtı olarak, eş zamanlı olarak injectrode yoluyla ilaçların teslimi ile, sıçanlarda ışık yanıp söner tarafından ortaya üstün colliculus gelen nöronal aktivitenin kaydetti. injectrode kayıt kapasitesi inj fonksiyonel karakterizasyonu izin veririlaç dağıtım lokalizasyonu üzerinde hassas kontrol lehine ection sitesi. Injectrode yüklü kimyasal madde seçimi anatomik deneyler için işaretleyiciler izleme dahil olmak üzere, geniş bir şekilde, bu yöntemin uygulanması, aynı zamanda, buradan çok gösterilmiştir ötesine uzanır.

injectrode inşaat tel bükülmüş bir kısmı ile bir cam pipet (D) içine beslenir ve açıklıktan dışarı doğru çıkıntı yapan, Şekil 1 bir gümüş tel (C) &#39;de gösterilmiştir. A 30 G iğne (B) bağlı ve yapıştırıcı ile cam pipet açılmasına kapatılır. Pipet püskürtme maddesi ile doldurulduktan sonra, cam mikro şırınga (A), bir iğne takılır. Mikro şırınga iğnesi (E) ve nerede gümüş tel cam pipet (F) çıkıntı bağlanır iyi bir mühür var olması önemlidir....

Watch this Scientific Journal Video about Simultaneous Electrophysiological Recording and Micro-injections of Inhibitory Agents in the Rodent Brain at JoVE.com

Simultaneous Electrophysiological Recording and Micro-injections of Inhibitory Agents in the Rodent Brain

Here, we craft a glass pipette with dual functions: inhibition of deep brain structures by microinjections of drugs and real-time monitoring of their effects through simultaneous electrophysiological recordings.

Eşzamanlı Elektrofizyolojik Kayıt ve kemirgen Beyindeki inhibitör Ajanların Mikro-enjeksiyon

Here we describe a method for the construction of a single-use &#8220;injectrode&#8221; using commercially accessible and affordable parts. A probing ...

simultaneous-electrophysiological-recording-micro-injections

Universidad de Cartagena UDEC

Research

JoVE Journal

Neuroscience

8.0K Görüntüleme.  Université de Montréal. Here, we craft a glass pipette with dual functions: inhibition of deep brain structures by microinjections of drugs and real-time monitoring of their effects through simultaneous electrophysiological recordings.

Video: Simultaneous Electrophysiological Recording and Micro-injections of Inhibitory Agents in the Rodent Brain

Simultaneous fMRI and Electrophysiology in the Rodent Brain

To examine the neural basis of the blood oxygenation level dependent (BOLD) magnetic resonance imaging (MRI) signal, we have developed a rodent model in which functional MRI data and in vivo intracortical recording can be performed simultaneously. The combination of MRI and electrical recording is technically challenging because the electrodes used for recording distort the MRI images and the MRI acquisition induces noise in the electrical recording. To minimize the mutual interference of the two modalities, glass microelectrodes were used rather than metal and a noise removal algorithm was implemented for the electrophysiology data. In our studies, two microelectrodes were separately implanted in bilateral primary somatosensory cortices (SI) of the rat and fixed in place. One coronal slice covering the electrode tips was selected for functional MRI. Electrode shafts and fixation positions were not included in the image slice to avoid imaging artifacts. The removed scalp was replaced with toothpaste to reduce susceptibility mismatch and prevent Gibbs ringing artifacts in the images. The artifact structure induced in the electrical recordings by the rapidly-switching magnetic fields during image acquisition was characterized by averaging all cycles of scans for each run. The noise structure during imaging was then subtracted from original recordings. The denoised time courses were then used for further analysis in combination with the fMRI data. As an example, the simultaneous acquisition was used to determine the relationship between spontaneous fMRI BOLD signals and band-limited intracortical electrical activity. Simultaneous fMRI and electrophysiological recording in the rodent will provide a platform for many exciting applications in neuroscience in addition to elucidating the relationship between the fMRI BOLD signal and neuronal activity.

We have developed a method for simultaneous functional magnetic resonance imaging and electrophysiological recording in the rodent brain, providing a platform for the investigation of the relationship between neural activity and the blood oxygenation level dependent (BOLD) MRI signal.

Kemirgen Beyin aynı anda fMRI ve Elektrofizyoloji

To examine the neural basis of the blood oxygenation level dependent (BOLD) magnetic resonance imaging (MRI) signal, we have developed a rodent model in ...

Nörobilim

Simultaneous Electroencephalography, Real-time Measurement of Lactate Concentration and Optogenetic Manipulation of Neuronal Activity in the Rodent Cerebral Cortex

Although the brain represents less than 5% of the body by mass, it utilizes approximately one quarter of the glucose used by the body at rest1. The function of non rapid eye movement sleep (NREMS), the largest portion of sleep by time, is uncertain. However, one salient feature of NREMS is a significant reduction in the rate of cerebral glucose utilization relative to wakefulness2-4. This and other findings have led to the widely held belief that sleep serves a function related to cerebral metabolism. Yet, the mechanisms underlying the reduction in cerebral glucose metabolism during NREMS remain to be elucidated.
	One phenomenon associated with NREMS that might impact cerebral metabolic rate is the occurrence of slow waves, oscillations at frequencies less than 4 Hz, in the electroencephalogram5,6. These slow waves detected at the level of the skull or cerebral cortical surface reflect the oscillations of underlying neurons between a depolarized/up state and a hyperpolarized/down state7. During the down state, cells do not undergo action potentials for intervals of up to several hundred milliseconds. Restoration of ionic concentration gradients subsequent to action potentials represents a significant metabolic load on the cell8; absence of action potentials during down states associated with NREMS may contribute to reduced metabolism relative to wake.
	Two technical challenges had to be addressed in order for this hypothetical relationship to be tested. First, it was necessary to measure cerebral glycolytic metabolism with a temporal resolution reflective of the dynamics of the cerebral EEG (that is, over seconds rather than minutes). To do so, we measured the concentration of lactate, the product of aerobic glycolysis, and therefore a readout of the rate of glucose metabolism in the brains of mice. Lactate was measured using a lactate oxidase based real time sensor embedded in the frontal cortex. The sensing mechanism consists of a platinum-iridium electrode surrounded by a layer of lactate oxidase molecules. Metabolism of lactate by lactate oxidase produces hydrogen peroxide, which produces a current in the platinum-iridium electrode. So a ramping up of cerebral glycolysis provides an increase in the concentration of substrate for lactate oxidase, which then is reflected in increased current at the sensing electrode. It was additionally necessary to measure these variables while manipulating the excitability of the cerebral cortex, in order to isolate this variable from other facets of NREMS.
	We devised an experimental system for simultaneous measurement of neuronal activity via the elecetroencephalogram, measurement of glycolytic flux via a lactate biosensor, and manipulation of cerebral cortical neuronal activity via optogenetic activation of pyramidal neurons. We have utilized this system to document the relationship between sleep-related electroencephalographic waveforms and the moment-to-moment dynamics of lactate concentration in the cerebral cortex. The protocol may be useful for any individual interested in studying, in freely behaving rodents, the relationship between neuronal activity measured at the electroencephalographic level and cellular energetics within the brain.

A procedure is described for manipulating the activity of cerebral cortical pyramidal neurons optogenetically while the electroencephalogram, electromyogram, and cerebral lactate concentration are monitored. Experimental recordings are performed on cable-tethered mice while they undergo spontaneous sleep/wake cycles. Optogenetic equipment is assembled in our laboratory; recording equipment is commercially available.

Eşzamanlı Elektroensefalografi, Kemirgen Serebral Korteks Laktat Konsantrasyonu ve Nöronal Faaliyet Optogenetic Manipülasyon Gerçek zamanlı ölçüm

Although the brain represents less than 5% of the body  by mass, it utilizes approximately one quarter of the glucose used by the body  at rest1. The ...

Extracellular Recording of Neuronal Activity Combined with Microiontophoretic Application of Neuroactive Substances in Awake Mice

Differences in the activity of neurotransmitters and neuromodulators, and consequently different neural responses, can be found between anesthetized and awake animals. Therefore, methods allowing the manipulation of synaptic systems in awake animals are required in order to determine the contribution of synaptic inputs to neuronal processing unaffected by anesthetics. Here, we present methodology for the construction of electrodes to simultaneously record extracellular neural activity and release multiple neuroactive substances at the vicinity of the recording sites in awake mice. By combining these procedures, we performed microiontophoretic injections of gabazine to selectively block GABAA receptors in neurons of the inferior colliculus of head-restrained mice. Gabazine successfully modified neural response properties such as the frequency response area and stimulus-specific adaptation. Thus, we demonstrate that our methods are suitable for recording single-unit activity and for dissecting the role of specific neurotransmitter receptors in auditory processing.
The main limitation of the described procedure is the relatively short recording time (~3 hr), which is determined by the level of habituation of the animal to the recording sessions. On the other hand, multiple recording sessions can be performed in the same animal. The advantage of this technique over other experimental procedures used to manipulate the level of neurotransmission or neuromodulation (such as systemic injections or the use of optogenetic models), is that the drug effect is confined to the local synaptic inputs to the target neuron. In addition, the custom-manufacture of electrodes allows adjustment of specific parameters according to the neural structure and type of neuron of interest (such as the tip resistance for improving the signal-to-noise ratio of the recordings).

We present methods for the construction of electrodes to simultaneously record extracellular neural activity and release multiple neuroactive substances at the vicinity of the recording sites in awake mice. This technique allows the detailed analysis of putative local synaptic inputs to the neuron of interest.

Uyanık Farelerde Nöroaktif Maddelerin Microiontophoretic Uygulaması ile birlikte Nöronal Faaliyet dışı Kaydı

Differences in the activity of neurotransmitters and neuromodulators, and consequently different neural responses, can be found between anesthetized and ...

Acute In Vivo Electrophysiological Recordings of Local Field Potentials and Multi-unit Activity from the Hyperdirect Pathway in Anesthetized Rats

Converging evidence shows that many neuropsychiatric diseases should be understood as disorders of large-scale neuronal networks. To better understand the pathophysiological basis of these diseases, it is necessary to precisely characterize in which way the processing of information is disturbed between the different neuronal parts of the circuit. Using extracellular in vivo electrophysiological recordings, it is possible to accurately delineate neuronal activity within a neuronal network. The application of this method has several advantages over alternative techniques, e.g., functional magnetic resonance imaging and calcium imaging, as it allows a unique temporal and spatial resolution and does not rely on genetically engineered organisms. However, the use of extracellular in vivo recordings is limited since it is an invasive technique that cannot be universally applied. In this article, a simple and easy to use method is presented with which it is possible to simultaneously record extracellular potentials such as local field potentials and multiunit activity at multiple sites of a network. It is detailed how a precise targeting of subcortical nuclei can be achieved using a combination of stereotactic surgery and online analysis of multi-unit recordings. Thus, it is demonstrated, how a complete network such as the hyperdirect cortico-basal ganglia loop can be studied in anesthetized animals in vivo.

Acute In Vivo Electrophysiological Recordings of Local Field Potentials and Multi-unit Activity from the Hyperdirect Pathway in Anesthetized Rats

In this study, the methodology is presented on how to perform multi-site in vivo electrophysiological recordings from the hyperdirect pathway under urethane anesthesia.

Akut<emIn vivo</em&gt; Anestezi Uygulanan Sıçanlarda Hiperdirect Pathway&#39;den Lokal Alan Potansiyellerinin Elektrofizyolojik Kayıtları ve Çoklu Ünite Aktivitesi

Converging evidence shows that many neuropsychiatric diseases should be understood as disorders of large-scale neuronal networks. To better understand the ...

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

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

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

Co yerelleştirilmiş elektroansefalografi ve yerel alan potansiyel kemirgen eşzamanlı kayıt

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

Recording and Modulation of Epileptiform Activity in Rodent Brain Slices Coupled to Microelectrode Arrays

Temporal lobe epilepsy (TLE) is the most common partial complex epileptic syndrome and the least responsive to medications. Deep brain stimulation (DBS) is a promising approach when pharmacological treatment fails or neurosurgery is not recommended. Acute brain slices coupled to microelectrode arrays (MEAs) represent a valuable tool to study neuronal network interactions and their modulation by electrical stimulation. As compared to conventional extracellular recording techniques, they provide the added advantages of a greater number of observation points and a known inter-electrode distance, which allow studying the propagation path and speed of electrophysiological signals. However, tissue oxygenation may be greatly impaired during MEA recording, requiring a high perfusion rate, which comes at the cost of decreased signal-to-noise ratio and higher oscillations in the experimental temperature. Electrical stimulation further stresses the brain tissue, making it difficult to pursue prolonged recording/stimulation epochs. Moreover, electrical modulation of brain slice activity needs to target specific structures/pathways within the brain slice, requiring that electrode mapping be easily and quickly performed live during the experiment. Here, we illustrate how to perform the recording and electrical modulation of 4-aminopyridine (4AP)-induced epileptiform activity in rodent brain slices using planar MEAs. We show that the brain tissue obtained from mice outperforms rat brain tissue and is thus better suited for MEA experiments. This protocol guarantees the generation and maintenance of a stable epileptiform pattern that faithfully reproduces the electrophysiological features observed with conventional field potential recording, persists for several hours, and outlasts sustained electrical stimulation for prolonged epochs. Tissue viability throughout the experiment is achieved thanks to the use of a small-volume custom recording chamber allowing for laminar flow and quick solution exchange even at low (1 mL/min) perfusion rates. Quick MEA mapping for real-time monitoring and selection of stimulating electrodes is performed by a custom graphic user interface (GUI).

We illustrate how to perform recording and electrical modulation of 4-aminopyridine-induced epileptiform activity in rodent brain slices using microelectrode arrays. A custom recording chamber maintains tissue viability throughout prolonged experimental sessions. Live electrode mapping and selection of stimulating pairs are performed by a custom graphical user interface.

Kayıt ve modülasyon Epileptiform aktivite elektrot dizilerin birleştiğinde kemirgen beyin dilimleri içinde

Temporal lobe epilepsy (TLE) is the most common partial complex epileptic syndrome and the least responsive to medications. Deep brain stimulation (DBS) ...

Microinjectrode System for Combined Drug Infusion and Electrophysiology

This microinjectrode system is designed for drug infusion, electrophysiology, and delivery and retrieval of experimental probes, such as microelectrodes and nanosensors, optimized for repeated use in awake, behaving animals. The microinjectrode system can be configured for multiple purposes: (1) simple arrangement of the cannula for placement of an experimental probe that would otherwise be too fragile to penetrate the dura mater, (2) microfluidic infusion of a drug, either independently or coupled to a cannula containing an experimental probe (i.e., microelectrode, nanosensor). In this protocol we explain the step by step construction of the microinjectrode, its coupling to microfluidic components, and the protocol for use of the system in vivo. The microfluidic components of this system allow for delivery of volumes on the nanoliter scale, with minimal penetration damage. Drug infusion can be performed independently or simultaneously with experimental probes such as microelectrodes or nanosensors in an awake, behaving animal. Applications of this system range from measuring the effects of a drug on cortical electrical activity and behavior, to understanding the function of a specific region of cortex in the context of behavioral performance based on probe or nanosensor measurements. To demonstrate some of the capabilities of this system, we present an example of muscimol infusion for reversible inactivation of the frontal eye field (FEF) in rhesus macaque during a working memory task.

We present a microinjectrode system designed for electrophysiology and assisted delivery of experimental probes (i.e., nanosensors, microelectrodes), with optional drug infusion. Widely available microfluidic components are coupled to a cannula containing the probe. A step-by-step protocol for microinjectrode construction is included, with results during muscimol infusion in macaque cortex.

Kombine İlaç İnfüzyonu ve Elektrofizyolojisi için Mikroenjekrode Sistemi

This microinjectrode system is designed for drug infusion, electrophysiology, and delivery and retrieval of experimental probes, such as microelectrodes ...

Drug Microinjection with Electrophysiological Recording in a Rat Brain

Source: Lai, J., et al. <a href="https://appjove.unicartagenaproxy.elogim.com/v/52271">Simultaneous electrophysiological recording and micro-injections of inhibitory agents in the rodent brain.</a> J. Vis. Exp. (2015)
This protocol outlines the assembly and use of an injectrode for precise drug delivery into targeted brain regions of an anesthetized rat, followed by monitoring of neuronal activity through recorded electrical signals.

Sıçan Beyninde Elektrofizyolojik Kayıt ile İlaç Mikroenjeksiyonu

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review ...

JoVE Encyclopedia of Experiments

Neurophysiology

Electrophysiology Techniques

In Vivo Electrophysiological Recordings of Brain Activities from Multiple Sites in Rats

Source: Haumesser, J. K., et al. <a href="https://appjove.unicartagenaproxy.elogim.com/v/55940">Acute In Vivo Electrophysiological Recordings of Local Field Potentials and Multi-unit Activity from the Hyperdirect Pathway in Anesthetized Rats.</a> J. Vis. Exp.(2017).
This video demonstrates the detailed procedure for preparing an anesthetized rat for electrophysiological recordings of brain activity from multiple sites. The scalp is dissected to access the skull, and holes are drilled at precise locations to install and secure electrodes. Tungsten microwire electrodes are then inserted into the targeted location to record the data.

Sıçanlarda Birden Fazla Bölgeden Beyin Aktivitelerinin İn Vivo Elektrofizyolojik Kayıtları

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. ...

A Micro-drive Array Method for Electrophysiological Recording from Multiple Brain Regions

Source: Shikano, Y., et al. <a href="https://appjove.unicartagenaproxy.elogim.com/v/56980">Simultaneous Recordings of Cortical Local Field Potentials, Electrocardiogram, Electromyogram, and Breathing Rhythm from a Freely Moving Rat.</a> J. Vis. Exp. (2018).
The video demonstrates simultaneous electrophysiological recording from multiple brain regions in a rat using an integrative micro-drive array. Recording electrodes are placed in the olfactory bulb, reference electrodes in the frontal cortex, and micro-drive array electrodes in the hippocampus. The rat is allowed to forage freely, and simultaneous recording from all the implanted electrodes is recorded using the array.

Eşzamanlı Elektrofizyolojik Kayıt ve kemirgen Beyindeki inhibitör Ajanların Mikro-enjeksiyon

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