作品は、NIH 1 R21NS057718 - 01によってサポートされていました。

1。これは、非生存手術です。最初のステップは、電極の移植です。この例では、電極は、両半球の一次体性感覚野の前足領域に注入されます。 <ol><li> 2パーセントイソフルランと定位手術システム上で所定の位置に修正とラット（雄性SDラット、200〜300 g）を麻酔。手術を開始する前に、動物がよく麻酔で、つま先のピンチへの応答を示さないことを確認してください。頭皮を開く前に毛皮を削除します。頭蓋骨上記の筋肉や他の組織を分離し、cauterizerを使用して骨表面上の任意の出血をブロックする。 </li><li>歯科用セメントを用いて、埋め込み電極のシャフトのための固定点として、頭蓋骨の表面（正中線前方V字型接合部付近）に桟橋を準備します。歯科用セメントを適用する前に、骨に小さなナイロンのネジセットを設定すると、安定性が向上する可能性があります。セメントで形をした桟橋のサイズは、ベース（図1参照）で、面積で約5 mm、高、3 × 5mmの2です。 </li><li>微細な先端電気ドリルを使用して、慎重に頭蓋骨を開いて、各半球の一次体性感覚野における前足の表現上の硬膜を露出。各穴の直径は1mm、前方に配置さ1 mmのブレグマから横4ミリメートル前後となるはずです。顕微鏡下で、あらゆる船の損傷を避けるために注意しながら、注射針の先端を使用して硬膜に小さな開口部をカット。 </li><li>各電極を挿入する前に、出血や滲出液が切開部の近くに存在しないことを確認してください。ガラス微小電極を約3で手術前に準備する必要があります〜4 CM1のシャフトの長さとインピーダンス〜5MΩを。人工CSF（ACSF）と電極のキャピラリーを記入して脳〜定位アームを使用して開かれた硬膜から0.4ミリメートルに（〜45 °、後方から前方に）斜めにそれぞれ電極を挿入する。所定の位置に固定する前に、電気信号​​を確認してください。 chloridized銀線の一端はACSFに浸漬し、入力に接続されているもう一方の端はアンプにつながる必要があります。開いた皮膚の奥に皮下接続されている銀線は、、参照電極として機能します。 </li><li>電極の固定する前に、二重手術のエリアをチェックし、出血や滲出液が発生しないことを確認してから、頭蓋骨に除去皮膚や筋肉を置き換えるために歯磨き粉を適用する。歯磨き粉の使用は、頭蓋骨/空気界面での感受性のミスマッチを減らすことにより、MR画像の品質を向上させます。歯科用セメントと準備桟橋（図1を参照）に、電極シャフトを取り付けます。 </li><li>歯科用セメントが硬化した後、MRIクレードルに動物を譲渡し、代わりに修正。体温、呼吸数、SPO 2、心拍数を含む研究、の残りのためのラットの生理的状態を監視します。 </li><li>コイルの中心から突起電極と、頭の上に表面コイルを（受信/送信）に置きます。電極の固定のためのサポートは、動物の呼吸によって引き起こされる運動を避けるためにつながるとして、クレードルの上に座っている追加のアーチ形のハードカバーが提供しています。同時イメージングと録音に使用リード線〜5メートル（アンプがちょうど磁石の部屋の外に置かれている）に拡張され、受動的なシールドとして機能する導電性プラスチックで覆われている。 </li><li>麻酔は必要に応じて神経活動の抑制を減らすためにメデトミジンにイソフルランから切り替えることができます。磁石に動物を転送する前に電気信号を最終的な時間を調べます。増幅× 1000、0.1Hzの〜5 K Hzのバンドパスフィルタを適用、60 Hzのノッチフィルタをかけ、アナログからデジタルへの変換のための12 kHzのサンプルレートを次のように我々の研究では、記録パラメータであった。 </li></ol> 2。この時点で、動物は、同時イメージングと記録のためのMRIスキャナーに挿入されます。動物は、イメージングの手順を通して麻酔する。 <ol><li> 9.4 T小動物用MRIシステム（ブルカー、ドイツは）私たちの研究に使用されました。録音の前に、画像パラメータを確立する必要があります。 3つの平面のスカウト画像は、fMRIのスキャンを配置するために使用されます。磁場の均一性を向上させるために、関心のボリュームがFASTMAP 1を使用して shimmedている。 fMRIの研究のために、コロナの撮像スライスは、電極が注入された二国間前足一次体性感覚の領域を、含まれている、選ばれました。行列のサイズ、64 × 64;の面内分解能、0.3 × 0.3ミリメートル2、スライス厚は2mm、TR / TE、15分の500ミリEPIの撮像パラメータは、FOV、1.92 × 1.92 cm 2とした。 </li><li>イメージングセットアップが完了すると、同時録音とfMRIを開始することができます。図2は、代表的なEPI画像と撮影時の生録音を示しています。各スキャンサイクルのごく一部（500分の22ミリ）のために持続飽和録音の画像取得結果、中勾配の急激な切り替え。画像取得後に、電気信号​​のretur非飽和発振の形でベースラインにNS（図3を参照）。組み合わせるfMRIと記録は静止状態（として本研究で示したように）中に、または刺激の間に行うことができる。刺激の研究のために、イメージングパラメータは9 Hzの、1〜現在の4 mAを使用して供給された前足の電気刺激で、静止状態の試験の場合と同じです。ラットは、最後のスキャンの後に安楽死させています。 </li></ol> 3。同時イメージングと録音した後、データが最終的な分析前に事前処理する必要があります。 <ol><li>我々は、電気生理学的記録（図3参照）から勾配アーチファクトの除去から始まります。 <ol><li>スキャンに起因するノイズの構造は、2つの連続した​​fMRIの画像間の間隔に対応するそれぞれが、すべて〜500ミリ秒（TR）のセクションを平均することによって抽出することができます。 </li><li>オリジナル録音から平均ノイズ構造を引きます。このメソッドは、不飽和記録のセグメントを修正。 </li><li>画像を取り込みながら勾配の交代に対応するそれぞれの飽和セグメントは前の時点と勾配誘発性飽和後の時点の間を通過線に置き換えられています。 </li></ol></li><li>局所電場電位（LFPs）のdenoised音声ファイルはその後、fMRIの時間のコースと同じ時間分解能を持つ電源のタイムコース、に変換されます。 2秒ビン内の平均電力は、シータバンド（4〜8ヘルツ）の周波数の1秒ビンと低い周波数（デルタバンド、1〜4 Hz）を、、と〜0.5秒のビンの間を計算するために使用されますより高い周波数（&gt; 8 Hzの、ガンマバンドへのα）のための飽和信号を隣接。すべての周波数帯域のためのスライディングウィンドウは、fMRIのデータのTRをマッチング、0.5秒単位で移動されました。 </li><li>画像データの場合は、標準的なfMRIの前処理は、頭部の動き補正、0.5mmの半値幅を持つ画像の平滑化、および線形ドリフトの除去を含め、実行されます。 </li><li>クロス相関分析は、LFPのパワータイムコースと画像データの各ボクセルからの時間経過との間で行われている。様々なタイムラグが（図4を参照）時間に依存する相関関係の調査を可能にする。 </li></ol>代表的な結果： 例として、この技術は自発的神経活動とBOLD変動との関係を調査するために使用されることがあります。図4一ラットから-2.5の間と9.5秒遅れている時にLFPのパワーとBOLD信号との相関マップを示しています。電極の先端付近の皮質領域から低周波BOLD変動（&lt;0.1 ​​Hz）は2の遅延〜6秒でLFPのパワーの変化（&lt;0.1 ​​Hz）と相関している<img src="/files/ftp_upload/1901/1901fig1.jpg" alt="図1" /> 図1表面コイルと電極の注入と撮像領域の概略構成。 <img src="/files/ftp_upload/1901/1901fig2.jpg" alt="図2" /> 図2電極チップを含む代表的な冠状EPI画像が、、左側のパネルに表示されます。右側のパネルには画像処理の実行前と実行中生の電気生理学的記録を示しています。 <img src="/files/ftp_upload/1901/1901fig3.jpg" alt="図3" /> 図3。1回のスキャンサイクル上で拡大することで、それは撮影時のアーティファクト（緑）は、元の録音（青）から削除される可能性があることがわかる。 denoised時間コース（赤）はさらなる分析のために使用されていました。 <img src="/files/ftp_upload/1901/1901fig4.jpg" alt="図4" /> 図4は、1つの電極からの自発的なデルタのバンド活動の力と時の安静状態のBOLD信号との相関のコロナのマップは、（典型的なラットから）-2.5から9.5秒に遅れているイソフルラン麻酔ラットの約4〜5秒で二国間SIで観察される最大の相関。カラーバーは、ピアソンrを表し、

<ol>
	<li>Gruetter, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Automatic,+localized+in+vivo+adjustment+of+all+first-+and+second-order+shim+coils.">Automatic, localized in vivo adjustment of all first- and second-order shim coils.</a> Magn Reson Med. 29 (6), 804-804 (1993).</li><li>Logothetis, N. K., Pauls, J., Augath, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neurophysiological+investigation+of+the+basis+of+the+fMRI+signal.">Neurophysiological investigation of the basis of the fMRI signal.</a> Nature. 412 (6843), 150-150 (2001).</li><li>Nir, Y., Fisch, L., Mukamel, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Coupling+between+neuronal+firing+rate,+gamma+LFP,+and+BOLD+fMRI+is+related+to+interneuronal+correlations.">Coupling between neuronal firing rate, gamma LFP, and BOLD fMRI is related to interneuronal correlations.</a> Curr Biol. 17 (15), 1275-1275 (2007).</li></ol>

電気生理学的記録とBOLD fMRIの両方が別々によく発達した手法です。しかし、同時に録音し、イメージングには2つのモダリティの相互干渉2のため困難である。ここでは、げっ歯類での複合実験のための可能な解決策を提供します。電極の注入の変更方法は、画像の品質に影響を最小限に抑え、電気録音のアーティファクト除去は、画像取得により誘導されるノイズを除去する必要があ...

simultaneous fmri and electrophysiology in the rodent brain

血中酸素レベル依存（BOLD）磁気共鳴画像（MRI）信号の神経基盤を調べるために、我々は、機能的MRIデータの齧歯類モデルを開発し、<em&gt; in vivoで</em&gt;皮質内の記録を同時に行うことができます。録音のために使用される電極は、MRI画像とMRI買収を歪めるためMRIと電気録音の組み合わせが技術的に困難であり、電気録音にノイズを誘導する。 twoモダリティの相互干渉を最小限に抑えるために、ガラス微小電極は、金属ではなく、使用され、ノイズ除去アルゴリズムは、電気生理学的データのために実装されました。我々の研究では、2つの微小電極を別々にラットの二国間の一次体性感覚野（SI）に移植したとの場所に固定。電極チップを覆う1つの冠状スライスは、機能的MRIに選ばれた。電極シャフトと固定位置は、画像アーチファクトを避けるために、画像のスライスに含まれていませんでした。削除された頭皮は、感受性のミスマッチを低減し、画像にギブスリンギングアーチファクトを防ぐために歯磨き粉に置き換えられました。画像取得中に急速にスイッチング磁場によって電気録音で誘導される成果物の構造は、実行するたびにスキャンのすべてのサイクルを平均することにより特徴づけられた。撮影時のノイズの構造は、元の録音から減算した。 denoised時間のコースは、fMRIのデータと組み合わせて、さらなる分析のために使用されていました。例として、同時取得は自発的なfMRIのBOLD信号と帯域制限された皮質の電​​気的活動との関係を決定するために使用されていました。同時fMRIとげっ歯類における電気生理学的記録は、fMRIのBOLD信号と神経活動の関係を解明することに加えて神経科学の多くのエキサイティングなアプリケーションのためのプラットフォームを提供します。

Watch this Scientific Journal Video about Simultaneous fMRI and Electrophysiology in the Rodent Brain at JoVE.com

Simultaneous fMRI and Electrophysiology in the Rodent Brain

我々は、神経活動と血中酸素レベル依存（BOLD）MRI信号との関係の調査のためのプラットフォームを提供し、齧歯類の脳における同時機能的磁気共鳴イメージングと電気生理学的記録法を開発した。

齧歯類の脳における同時fMRIと電気生理学

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

simultaneous-fmri-and-electrophysiology-in-the-rodent-brain

Research

JoVE Journal

Neuroscience

13.3K ビュー.  Emory University. 我々は、神経活動と血中酸素レベル依存（BOLD）MRI信号との関係の調査のためのプラットフォームを提供し、齧歯類の脳における同時機能的磁気共鳴イメージングと電気生理学的記録法を開発した。

ビデオ: Simultaneous fMRI and Electrophysiology in the Rodent Brain

Design and Construction of a Cost Effective Headstage for Simultaneous Neural Stimulation and Recording in the Water Maze

Headstage preamplifiers and source followers are commonly used to study neural activity in behavioral neurophysiology experiments. Available commercial products are often expensive, not easily customized, and not submersible. Here we describe a method to design and build a customized, integrated circuit headstage for simultaneous 4-channel neural recording and 2-channel simulation in awake, behaving animals. The headstage is designed using a free, commercially available CAD-type design package, and can be modified easily to accommodate different scales (e.g. to add channels). A customized printed circuit board is built using surface mount resistors, capacitors and operational amplifiers to construct the unity gain source follower circuit. The headstage is made water-proof with a combination of epoxy, parafilm and a synthetic rubber putty. We have successfully used this device to record local field potentials and stimulate different brain regions simultaneously via independent channels in rats swimming in a water maze. The total cost is &lt; $30/unit and can be manufactured readily.

We present a low-cost method to design and construct a light headstage pre-amplifier system with simultaneous neural recording and stimulation capability. This device can be waterproofed for use in swimming animals.

水迷路における同時神経刺激および記録のためのコスト効率のヘッドステージの設計と構築

Headstage preamplifiers and source followers are commonly used to study neural activity in behavioral neurophysiology experiments. Available commercial ...

神経科学

Combining Transcranial Magnetic Stimulation and fMRI to Examine the Default Mode Network

The default mode network is a group of brain regions that are active when an individual is not focused on the outside world and the brain is at "wakeful rest."1,2,3 It is thought the default mode network corresponds to self-referential or "internal mentation".2,3
It has been hypothesized that, in humans, activity within the default mode network is correlated with certain pathologies (for instance, hyper-activation has been linked to schizophrenia 4,5,6 and autism spectrum disorders 7 whilst hypo-activation of the network has been linked to Alzheimer's and other neurodegenerative diseases 8). As such, noninvasive modulation of this network may represent a potential therapeutic intervention for a number of neurological and psychiatric pathologies linked to abnormal network activation. One possible tool to effect this modulation is Transcranial Magnetic Stimulation: a non-invasive neurostimulatory and neuromodulatory technique that can transiently or lastingly modulate cortical excitability (either increasing or decreasing it) via the application of localized magnetic field pulses.9
In order to explore the default mode network's propensity towards and tolerance of modulation, we will be combining TMS (to the left inferior parietal lobe) with functional magnetic resonance imaging (fMRI). Through this article, we will examine the protocol and considerations necessary to successfully combine these two neuroscientific tools.

In this article, we examine the methodology and considerations relevant to the combination of TMS and fMRI to examine the effects of brain stimulation on the default network.

デフォルトモードのネットワークを検査する経頭蓋磁気刺激とfMRIを組み合わせ

The default mode network is a group of brain regions that are active when an individual is not focused on the outside world and the brain is at "wakeful ...

Co-analysis of Brain Structure and Function using fMRI and Diffusion-weighted Imaging

The study of complex computational systems is facilitated by network maps, such as circuit diagrams. Such mapping is particularly informative when studying the brain, as the functional role that a brain area fulfills may be largely defined by its connections to other brain areas. In this report, we describe a novel, non-invasive approach for relating brain structure and function using magnetic resonance imaging (MRI). This approach, a combination of structural imaging of long-range fiber connections and functional imaging data, is illustrated in two distinct cognitive domains, visual attention and face perception. Structural imaging is performed with diffusion-weighted imaging (DWI) and fiber tractography, which track the diffusion of water molecules along white-matter fiber tracts in the brain (Figure 1). By visualizing these fiber tracts, we are able to investigate the long-range connective architecture of the brain. The results compare favorably with one of the most widely-used techniques in DWI, diffusion tensor imaging (DTI). DTI is unable to resolve complex configurations of fiber tracts, limiting its utility for constructing detailed, anatomically-informed models of brain function. In contrast, our analyses reproduce known neuroanatomy with precision and accuracy. This advantage is partly due to data acquisition procedures: while many DTI protocols measure diffusion in a small number of directions (e.g., 6 or 12), we employ a diffusion spectrum imaging (DSI)1, 2 protocol which assesses diffusion in 257 directions and at a range of magnetic gradient strengths. Moreover, DSI data allow us to use more sophisticated methods for reconstructing acquired data. In two experiments (visual attention and face perception), tractography reveals that co-active areas of the human brain are anatomically connected, supporting extant hypotheses that they form functional networks. DWI allows us to create a "circuit diagram" and reproduce it on an individual-subject basis, for the purpose of monitoring task-relevant brain activity in networks of interest.

We describe a novel approach for simultaneous analysis of brain function and structure using magnetic resonance imaging (MRI). We assess brain structure with high-resolution diffusion-weighted imaging and white-matter fiber tractography. Unlike standard structural MRI, these techniques allow us to directly relate anatomical connectivity to functional properties of brain networks.

fMRIと拡散強調画像を用いた脳の構造と機能の同時分析

The study of complex computational systems is facilitated by network maps, such as circuit diagrams. Such mapping is particularly informative when ...

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.

同時脳波、リアルタイム乳酸濃度の測定と齧歯類大脳皮質ニューロン活動のOptogeneticマニピュレーション

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 ...

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.

げっ歯類での同時のfMRIと脳深部刺激

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

In vivo Optogenetic Stimulation of the Rodent Central Nervous System

The ability to probe defined neural circuits in awake, freely-moving animals with cell-type specificity, spatial precision, and high temporal resolution has been a long sought tool for neuroscientists in the systems-level search for the neural circuitry governing complex behavioral states. Optogenetics is a cutting-edge tool that is revolutionizing the field of neuroscience and represents one of the first systematic approaches to enable causal testing regarding the relation between neural signaling events and behavior. By combining optical and genetic approaches, neural signaling can be bi-directionally controlled through expression of light-sensitive ion channels (opsins) in mammalian cells. The current protocol describes delivery of specific wavelengths of light to opsin-expressing cells in deep brain structures of awake, freely-moving rodents for neural circuit modulation. Theoretical principles of light transmission as an experimental consideration are discussed in the context of performing in vivo optogenetic stimulation. The protocol details the design and construction of both simple and complex laser configurations and describes tethering strategies to permit simultaneous stimulation of multiple animals for high-throughput behavioral testing.

In vivo Optogenetic Stimulation of the Rodent Central Nervous System

Optogenetics has become a powerful tool for use in behavioral neuroscience experiments. This protocol offers a step-by-step guide to the design and set-up of laser systems, and provides a full protocol for carrying out multiple and simultaneous in vivo optogenetic stimulations compatible with most rodent behavioral testing paradigms.

齧歯類の中枢神経系の生体光遺伝学的刺激

The ability to probe defined neural circuits in awake, freely-moving animals with cell-type specificity, spatial precision, and high temporal resolution ...

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

Here we describe a method for the construction of a single-use &#8220;injectrode&#8221; using commercially accessible and affordable parts. A probing system was developed that allows for the injection of a drug while recording electrophysiological signals from the affected neuronal population. This method provides a simple and economical alternative to commercial solutions. A glass pipette was modified by combining it with a hypodermic needle and a silver filament. The injectrode is attached to commercial microsyringe pump for drug delivery. This results in a technique that provides real-time pharmacodynamics feedback through multi-unit extracellular signals originating from the site of drug delivery. As a proof of concept, we recorded neuronal activity from the superior colliculus elicited by flashes of light in rats, concomitantly with delivery of drugs through the injectrode. The injectrode recording capacity permits the functional characterization of the injection site favoring precise control over the localization of drug delivery. Application of this method also extends far beyond what is demonstrated here, as the choice of chemical substance loaded into the injectrode is vast, including tracing markers for anatomic experiments.

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.

同時電気生理学的記録およびげっ歯類の脳における阻害剤のマイクロ注射

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

Personalized Needles for Microinjections in the Rodent Brain

Microinjections have been used for a long time for the delivery of drugs or toxins within specific brain areas and, more recently, they have been used to deliver gene or cell therapy products. Unfortunately, current microinjection techniques use steel or glass needles that are suboptimal for multiple reasons: in particular, steel needles may cause tissue damage, and glass needles may bend when lowered deeply into the brain, missing the target region. In this article, we describe a protocol to prepare and use quartz needles that combine a number of useful features. These needles do not produce detectable tissue damage and, being very rigid, ensure reliable delivery in the desired brain region even when using deep coordinates. Moreover, it is possible to personalize the design of the needle by making multiple holes of the desired diameter. Multiple holes facilitate the injection of large amounts of solution within a larger area, whereas large holes facilitate the injection of cells. In addition, these quartz needles can be cleaned and re-used, such that the procedure becomes cost-effective.

We describe here a protocol for microinjection in the rodent brain that uses quartz needles. These needles do not produce detectable tissue damage and ensure reliable delivery even in deep regions. Moreover, they can be adapted to research needs by personalized designs and can be re-used.

パーソナライズされた齧歯動物の脳でのマイクロインジェクション針

Microinjections have been used for a long time for the delivery of drugs or toxins within specific brain areas and, more recently, they have been used to ...

Simultaneous Cryosectioning of Multiple Rodent Brains

Histology and immunohistochemistry are routine methods of analysis to visualize microscopic anatomy and localize proteins within biological tissue. In neuroscience, as well as a plethora of other scientific fields, these techniques are used. Immunohistochemistry can be done on slide mounted tissue or free-floating sections. Preparing slide-mounted samples is a time intensive process. The following protocol for a technique, called the Megabrain, reduced the time taken to cryosection and mount brain tissue by up to 90% by combining multiple brains into a single frozen block. Furthermore, this technique reduced variability seen between staining rounds, in a large histochemical study. The current technique has been optimized for using rodent brain tissue in downstream immunohistochemical analyses; however, it can be applied to different scientific fields that use cryosectioning.

Here, we present a protocol to freeze and section brain tissue from multiple animals as a timesaving alternative to processing single brains. This reduces staining variability during immunohistochemistry and reduces time cryosectioning and imaging.

複数の齧歯動物の頭脳の同時セクショニング

Histology and immunohistochemistry are routine methods of analysis to visualize microscopic anatomy and localize proteins within biological tissue. In ...

Using Near-infrared Fluorescence and High-resolution Scanning to Measure Protein Expression in the Rodent Brain

Neuroscience is the study of how cells in the brain mediate various functions. Measuring protein expression in neurons and glia is critical for the study of neuroscience as cellular function is determined by the composition and activity of cellular proteins. In this article, we describe how immunocytochemistry can be combined with near-infrared high-resolution scanning to provide a semi-quantitative measure of protein expression in distinct brain regions. This technique can be used for single or double protein expression in the same brain region. Measuring proteins in this fashion can be used to obtain a relative change in protein expression with an experimental manipulation, molecular signature of learning and memory, activity in molecular pathways, and neural activity in multiple brain regions. Using the correct proteins and statistical analysis, functional connectivity among brain regions can be determined as well. Given the ease of implementing immunocytochemistry in a laboratory, using immunocytochemistry with near-infrared high-resolution scanning can expand the ability of the neuroscientist to examine neurobiological processes at a systems level.

Here, we present a protocol that uses near-infrared dyes in conjunction with immunohistochemistry and high-resolution scanning to assay proteins in brain regions.

げっ歯類の脳におけるタンパク質発現を測定するために近赤外蛍光と高解像度走査を用いること

Neuroscience is the study of how cells in the brain mediate various functions. Measuring protein expression in neurons and glia is critical for the study ...