この作業は神経通信技術センター（CNCT）、生物医学イメージングとバイオの国立研究所（NIBIB、P41 EB002030）によって資金を供給され、国立衛生研究所（NIH）でサポートされているP41リソースセンターによって資金を供給された。著者らは、手術手技の開発のためのNeuroNexus Technologiesのリオフェッターに感謝したいと、手術中に使用される機器のリストをコンパイルするためにミシガン大学のパテルがパラス。

1。事前にインプラントの手順（共通） <ol><li>にも耳の後ろの目の間から、動物の頭の上を剃る。頭を剃る、電気理髪師のバリカンを使用しています。その後、イソプロピルアルコールで最初のヘッドをスクラブbetadine（3X繰り返す）。眼潤滑剤を適用します。 </li><li>定位耳バーで動物をマウントします。運河に対して頭を横に移動することによって、動物の右耳の外耳道を右に定位耳バーの固定の先端を指示します。細心の注意は、それぞれの耳バーの先端が正しく内耳に配置されることを保証するために注意する必要があります。次に、外耳道に緩んで左耳バーを押すと、わずか内側に圧力をかける。動物の鼻をつかんで、しっかりと前後に小刻み。動物が適切にマウントされている場合はヘッドがしっかりとマウントされた感じになります。 </li></ol> 2。着床前の手術（共通） <ol><li>細かいハサミでは、2つの半CIRCをカットLESは、正中線から外側に向かって、部分的に頭皮を削除する</li><li>骨に付着する骨膜結合組織をこすり削除します。毛細血管がスムーズにカットするのではなく粉砕されるので、そのような外科用メスの刃又は滅菌綿棒の先端の背面として鈍いエッジでこすると出血を最小限に抑えることができます。骨のワックスまたは焼灼鉄を用いた骨の出血を制御します。 </li><li>消毒と過酸化水素を用いた骨の表面を清掃してください。もっとはっきりと見える頭蓋縫合を作るために骨を乾燥させます。ブレグマとラムダの点に注意し、頭を水平に使用する必要があります。開頭術と電極インプラントの場所はブレグマを基準にして測定されるべきであり、一時的にペンで、表面に浅く掘削によってマークすることができます。開頭術の場所は、研究の実験の目的（すなわちモーター、感覚実験1）に依存します。 </li><li>骨にscrewholesを開けると地面に置くとsを安定させる乗組員。常にステンレス鋼またはチタンのネジを使用します。できるだけインチあたりの多数のスレッドが望まれているように、小さな宝石のネジ（例：＃2-56、または＃00から80の場合）。 </li></ol> 3。シリコン電極インプラント手順<ol><li>ネジを注入した後に開頭術を開けます。開頭の角の4つの浅いパイロットマークをドリルダウンします。ガイドとしてパイロットマークを使用すると、開頭の周囲をドリルダウンします。頑丈な鉗子のペアを使用して掘削されていない骨の中心質量を削除します。開いている開頭術のエッジはさらに掘削洗浄する必要があります。骨の薄い部分はmicrorongeursのペアで切り取られることがあります。正中縦縫合糸1の0.5ミリメートル内に現れる可能性があり矢状静脈洞、などの主要血管を避けるために慎重に手術を予定しています。矢状静脈洞を穿刺すると、広範な出血を引き起こす可能性があります。露出硬膜は、ACSFまたは生理食塩水和に保つ必要があります。 </li><li>ピアス硬膜へあなたは、ファッションに小さなゲージの滅菌注射針（例えば、&lt;28ゲージ）を使用して、小さなフックが必要になります。針のベベルを取る時点で90度曲げを形成するためにわずかに傾けてしっかりとした表面（例えば、メスの平らな部分など）に対して平らそれを押してください。硬膜の表面をキャッチするために、このフックを使用して、脳表面から上方に持ち上げます。硬膜は、マイクロはさみを使ってカットすることができますまたは硬フックの横方向の動きを使用して、引き裂かれることがあります。硬フックを使用する場合は、脳の表面の血管を避けるように注意してください。あなたは、CSFは硬膜を切除されたときに漏れると硬膜の表面からの色の違いに気付くはずです。 </li><li>慎重に接地ネジと、歯科用セメントの代わりにタクトのベースの周りにシリコン電極のアース線を接続します。シリコン電極は、電極ケーブルのリード線が開頭の上にアーチしていることを慎重に配置する必要があります。 </li><li>インプラントは、リニアモータアクチュエータPLAを使用して配置されます。定位固定装置にCED。挿入装置に電極を接続するには、ポリエチレングリコール（PEG）は、はんだごてで加熱し、挿入ロッドと記録電極に印加される。 </li><li>電極は固定されて、手動で脳の表面上の所望の定位位置に電極の先端を下げます。 </li><li>リニアアクチュエータへのソフトウェアインタフェースを使用して、脳組織内の所望の深さに電極を移動します。 </li><li>電極ケーブルに隣接するネジからPEGを付加することによって脳内の記録電極を固定します。あなたは今結合したPEGを溶解するために生理食塩水を追加することにより、挿入装置から電極を削除することができます。離れてそれぞれの電極の開頭術、リピートから挿入装置を持ち上げます。 </li><li>生理食塩水に浸したGelfoam露出した開頭手術をカバーしています。シリコーンポリマーは、その後電極ケーブルをカバーし、歯科用アクリルからそれを保護するために使用されます。最終的にコネクタを配置する場所は、その堅牢なヘッドキャップを作るために歯科用アクリルを適用します。 </li></ol> 4。 MicroECoG電極インプラント手順<ol><li> microECoG電極の注入はわずかに大きく開頭術を必要とします。 5×5 mmのデバイスでは、6×6 mmの開頭術を行う必要があります。それはまだ硬膜または軟膜に触れることの危険にさらされ乾燥していない間に開頭術を掘削する前に、UV硬化型歯科用アクリルは、開頭部位の周囲に適用されます。 </li><li>無菌手術のドリルは開頭術の一般的な形状の骨の表面からドリルダウンする＃107バリで使用されます。小さい＃106バリは、その後薄い透明層に骨の残りの部分を下にドリルアウトすることができます。 Rongeursは、骨の残りの部分を持ち上げことができるようになります。 microECoGは、硬膜外またはsubdurally移植することができる。再び、人工脳脊髄液または生理食塩水硬膜は、十分な水分補給を維持。 </li><li> microECoGを埋め込むためには、OPE上の定位腕を置くn個の頭蓋骨、滅菌テープを使用して腕に電極を固定します。電極は現在、開頭術に分け低下させることができる。電極部位が下向きされており、硬膜または軟膜表面との接触を作ることになりますことを確認してください。湿った保存した場合は、電極が組織にスライドしなければなりませんし、場所に留まります。 </li><li>アース線は、少なくとも3倍以上、それ自体の下の周りにラップすることによって、接地ネジに接続されています。アース用ネジは、硬膜と接触したすべてのネジにすることができます。基準線も同様の基準ネジに接続することができます。 </li><li>露出した硬膜またはPIAがある場合には生理食塩水に浸したGelfoamの小片は、電極の周囲に配置する必要があります。生理食塩水に浸したGelfoam少量のも、薄膜電極の上部をカバーする必要があります。 UV硬化型歯科用アクリルは、Gelfoamのトップに適用することができ、安定したヘッドキャップを作成するために使用することができます。アクリルは、薄膜ケーブルCOVERIに直接適用されますNGにコネクタが到達するまで。コネクタキャップの底部を超えてカバーすることがないように注意してください。 </li><li>また、4.3 microECoGの注入は、直ちにmicroECoG基板で製造されて小さな穴を介してシリコン微小電極アレイ（ステップ3.2から3.6）の注入を続けることができます。これは周囲の脳表面からの高解像度のフィールド·レコーディングと一緒に様々な深さの両方で、単一ユニットの同時録画が可能になります。 </li></ol> 5。術後の回復<ol><li>セメントが完全に硬化した後、ヘッドキャップの周りにしっかりと皮膚を縫合し、定位フレームから動物を削除します。 </li><li>おびただしく傷の周り防腐剤粉末や抗生物質軟膏を適用します。耳からの出血がある場合は、外耳道にいくつかの抗生物質も配置します。 </li><li>バルビツール酸塩は、その体温を維持することから動物を防ぐためとして動物を暖かく保つ。ハウスEAChは、慢性的に別のケージに動物を移植した。これは通常、手術から完全に回復するために動物の四から七日かかります。 </li></ol> 6。代表的な結果成功したシリコン電極のインプラント手術は（1kHzで）10kOhm-50kOhmの間に500kOhm-2MOhm間の測定インピーダンス、薄膜電極を持つサイトを記録することになります。神経信号も同様に直後の手術をチェックすることができます。あなたが移植された電極上にスパイクを見て、薄膜の表面電極（図1）徐波の振動を見ることができるはずです。 <img src="/files/ftp_upload/3565/3565fig1.jpg" alt="図1" /> 図1。単一ユニットの活動と皮質振動の代表的な結果。新皮質に移植された慢性的なシリコン電極6が同時に記録されたチャネルから）ワイドバンドのデータ。スパイクが頻繁に記録部位を越えて見ることができることに注意してください。 b）のE脳の表面の16チャネル薄膜uECoG電極アレイの6チャンネルからCOG振動。すべての16の信号の平均値は、各トレース（一般的な平均参照）から削除された、500 Hzのデジタルローパスフィルタを適用した。

<ol>
	<li>Paxinos, G. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The Rat Brain in Stereotaxic Coordinates. , (2007).</li><li>Fujisawa, S. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavior-dependent+short-term+assembly+dynamics+in+the+medial+prefrontal+cortex.">Behavior-dependent short-term assembly dynamics in the medial prefrontal cortex.</a> Nat. Neurosci. 11 (7), 823-833 (2008).</li><li>Merrill, D. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electrical+stimulation+of+excitable+tissue:+design+of+efficacious+and+safe+protocols.">Electrical stimulation of excitable tissue: design of efficacious and safe protocols.</a> J. Neurosci. Methods. 141, 171-171 (2005).</li><li>Gage, G. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Naive+coadaptive+cortical+control.">Naive coadaptive cortical control.</a> J. Neural Eng. 2, 52-52 (2005).</li><li>Marzullo, T. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+Closed-Loop+Neural+Interface+Technology+in+a+Rat+Model:+Combining+Motor+Cortex+Operant+Conditioning+With+Visual+Cortex+Microstimulation.">Development of Closed-Loop Neural Interface Technology in a Rat Model: Combining Motor Cortex Operant Conditioning With Visual Cortex Microstimulation.</a> IEEE Trans. Neur. Sys. and Rehab. Eng. 18 (2), 117-126 (2010).</li><li>Maynard, E. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Utah+intracortical+electrode+array:+a+recording+structure+for+potential+brain-computer+interfaces.">The Utah intracortical electrode array: a recording structure for potential brain-computer interfaces.</a> Electroencephalogr. Clin. Neurophysiol. 102, 228-239 (1997).</li><li>Kipke, D. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Silicon-substrate+intracortical+microelectrode+arrays+for+long-term+recording+of+neuronal+spike+activity+in+cerebral+cortex.">Silicon-substrate intracortical microelectrode arrays for long-term recording of neuronal spike activity in cerebral cortex.</a> IEEE Trans. Neural Syst. Rehab. Eng. 11 (2), 151-155 (2003).</li><li>Vetter, R. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronic+neural+recording+using+silicon-substrate+microelectrode+arrays+implanted+in+cerebral+cortex.">Chronic neural recording using silicon-substrate microelectrode arrays implanted in cerebral cortex.</a> IEEE Trans. Biomed. Eng. 51 (6), 896-904 (2004).</li></ol>

DRKには、NeuroNexus Technologiesの一部所有者です。

4,5 neuroprostheticシステムのための微小刺激3、または制御信号を提供する、脳機能2を研究するための皮質内および表面のマルチチャンネル·レコーディング·インターフェイスを使用しての関心が高まっている。このビデオで説明されているメソッドは、慢性的な浸透と表面電極システムを移植する方法を示しています。他のシリコン慢性電極システムは6を存在するが、我々はNeuroNexus技術7,8によって開発された平面電極を移植に焦点を当てている。他の電極システムの移植のための技術はかなり異なります。 説明した外科手術の手順から、定位耳バーで動物をマウントする最も困難と移植された電極の配置の精度を決定する上で最も重要なステップとなる可能性があります。細心の注意は、それぞれの耳バーの先端が正しく内耳に配置されていることを前提とするため、注意しなければならない。のAssura後左手の親指と人差し指で頭をサポートし、耳バーの先端に対して頭部の圧力を適用するために継続しながらNCEは、耳バーの先端が動物の右耳の内耳にあることを、右手で、動物の首を解放します。右手で、外耳道に緩んで左耳バーを押すと、わずか内側に圧力をかける。あなたが内耳に左耳バーの先端の滑りを感じることができるまで、ダウンして、順方向およびすべての周りに動物の頭の左側に移動します。右手で、左耳バーに圧力をかけ続けています。動物の頭を解放し、あなたの現在無料の左手で、左耳のバーのセットネジを締めます。

<table border="1"><tbody><tr><td> 機器 </td><td> 会社 </td><td> カタログ番号 </td><td> コメント </td></tr><tr><td>骨ねじ - ＃2-56×3/16、なべ頭、18から8 / S </td><td> BoltDepot </td><td> 7650 </td><td></td></tr><tr><td>骨ねじ - ＃2-56×1/8、なべ頭、18から8 / S </td><td> BoltDepot </td><td> 9617 </td><td></td></tr><tr><td>大酸素タンク - サイズK </td><td>極低温ガス</td><td> OXKME </td><td></td></tr><tr><td>小さな酸素ボンベ - サイズE </td><td>極低温ガス</td><td> OXYE-AL </td><td></td></tr><tr><td> Vannas-テュービンゲン春はさみ - 2.5ミリメートルブレード、アングル</td><td>ファイン科学ツール</td><td> 15002から08 </td><td></td></tr><tr><td>デュモン＃5鉗子、テフロン</td><td>ファイン科学ツール</td><td> 11626から11 </td><td></td></tr><tr><td>デュモン＃5鉗子、Dumostar </td><td>ファイン科学ツール</td><td> 11295から10 </td><td></td></tr><tr><td>学生Vannas春はさみ</td><td>ファイン科学ツール</td><td> 91500から09 </td><td></td></tr><tr><td>アドソン鑷子</td><td>ファイン科学ツール</td><td> 11006から12 </td><td></td></tr><tr><td>カーブFriendman-ピアソンRonqeur、 </td><td>ファイン科学ツール</td><td> 16221から14 </td><td></td></tr><tr><td>アイリスはさみ - ToughCut </td><td>ファイン科学ツール</td><td> 14058から11 </td><td></td></tr><tr><td> Vannas-テュービンゲン春はさみ - 5ミリメートルブレード、ストレート</td><td> FiのNE科学ツール</td><td> 15003から08 </td><td></td></tr><tr><td>スカルペルハンドル</td><td>ファイン科学ツール</td><td> 10003から12 </td><td></td></tr><tr><td>プラスチック製の殺菌ボックス</td><td>ファイン科学ツール</td><td> 20810から02 </td><td></td></tr><tr><td>ステンレススチール製の定規</td><td>ファイン科学ツール</td><td> 30086から15 </td><td></td></tr><tr><td>プローブ＆ヘラ - ストレート14センチメートル</td><td>ファイン科学ツール</td><td> 10090から13 </td><td></td></tr><tr><td>ストレート止血剤</td><td>ファイン科学ツール</td><td> 13002から10 </td><td></td></tr><tr><td>止血剤は、湾曲した</td><td>ファイン科学ツール</td><td> 13003から10 </td><td></td></tr><tr valign="bottom"><td>マイクロドリル駅inlessスチールバリ - 0.7ミリメートル</td><td>ファイン科学ツール</td><td> 19008から07 </td><td> 10/Pack </td></tr><tr valign="bottom"><td>マイクロドリルステンレス鋼のバリ - 0.5ミリメートル</td><td>ファイン科学ツール</td><td> 19007から05 </td><td> 10/Pack </td></tr><tr valign="bottom"><td>セルフタッピング骨ねじ - 1.17ミリメートル（直径） - 4.7ミリメートル（長さ） </td><td>ファイン科学ツール</td><td> 19010から00 </td><td> 100/Pack </td></tr><tr valign="bottom"><td>人工涙液</td><td>フィッシャー</td><td> NC9860842 </td><td>各</td></tr><tr valign="bottom"><td> Betadine </td><td>フィッシャー</td><td> 19-066452 </td><td>ボトル</td></tr><tr valign="bottom"><td> F-AIRカーボンフィルター</td><td>フィッシャー</td><td> NC9112250 </td><td>キャニスター</td></tr><tr valign="bottom"><td>アプリケーターコットンは、先端不稔の非</td> &lt;TD&gt;資材·サービス</td><td> 1104 </td><td> 10Pack/Case </td></tr><tr valign="bottom"><td>ガーゼ2 &quot;×2&quot; </td><td>資材·サービス</td><td> 1630 </td><td> 25Pack/Case </td></tr><tr valign="bottom"><td>針先、23 GA×1 &quot; </td><td>資材·サービス</td><td> 39412 </td><td>ケース販売</td></tr><tr valign="bottom"><td>針の先端、27 GA×1.25 &quot; </td><td>資材·サービス</td><td> 25251 </td><td></td></tr><tr valign="bottom"><td>針先は、30 GA×0.5 &quot; </td><td>資材·サービス</td><td> 22023 </td><td> 10Box/Case </td></tr><tr valign="bottom"><td>軟膏、三重の抗生物質0.6グラム</td><td>資材·サービス</td><td> 2528 </td><td></td></tr><tr valign="bottom"><td>ポーチ、セルフシール滅菌Pouche<html> S、3.5 &quot;×8&quot; </td><td>資材·サービス</td><td> 1023 </td><td> 10Box/Case </td></tr><tr valign="bottom"><td>ソリューション千ミリリットルのリンガ</td><td>資材·サービス</td><td> 5263 </td><td> 12Bag/Case </td></tr><tr valign="bottom"><td>手術用メスの刃使い捨て無菌第15位</td><td>資材·サービス</td><td> 1975 </td><td> 3Box/Case </td></tr><tr valign="bottom"><td>縫合糸、ナイロンw/P-3ニードル、5-0、18インチ</td><td>資材·サービス</td><td> 4618 </td><td> 12/Box </td></tr><tr valign="bottom"><td> Underpad、使い捨て23 &quot;×36&quot; </td><td>資材·サービス</td><td> 2545 </td><td> 15Pack/Case </td></tr><tr valign="bottom"><td>潤滑剤、ゼリー外科滅菌</td><td>資材·サービス</td><td> 2538 </td><td> 12Box/Case </td></tr>&quot;&gt; <td>吸収発泡ゲル2cmx6cmx7mm </td><td>資材·サービス</td><td> 3107 </td><td></td></tr><tr valign="bottom"><td>シックスピースセットスクリュードライバーセット</td><td>国立宝石用品</td><td> ETSCR98001 </td><td></td></tr><tr valign="bottom"><td>クイック·シル接着</td><td> WPI </td><td> KWIK-SIL </td><td></td></tr><tr valign="bottom"><td>クイック·シルチューブ</td><td> WPI </td><td> 600009 </td><td></td></tr><tr valign="top"><td>慢性的なシリコン電極16から64のサイト</td><td> NeuroNexus技術</td><td>プローブの設計によって異なります</td><td></td></tr><tr valign="top"><td>慢性的な薄膜の表面電極（microECoG）16から32のサイト</td><td> NeuroNexus技術</td><td>プローブの設計によって異なります</td><td>&nbsp;
 </td>
 </tr>
 </tbody>
</table>

surgical implantation of chronic neural electrodes for recording single unit activity and electrocorticographic signals

長期的な電気生理学的記録の成功は、多くの場合、移植手術の品質に依存します。ここでは、電極系を注入するプロセスを学習している外科医のための有用な情報を提供します。我々は貫通と表面電極両方の注入手順を示しています。手術のプロセスは、プロシージャ全体で各ステップの詳細な説明を含めて、最初から最後まで説明されています。それはこのビデオガイドは齧歯類モデルやその他の小さな動物モデルで実施手続きに向かって集中されていることにも留意すべきである。説明する手順の変更が他の動物モデルの実現可能である。

Watch this Scientific Journal Video about Surgical Implantation of Chronic Neural Electrodes for Recording Single Unit Activity and Electrocorticographic Signals at JoVE.com

Surgical Implantation of Chronic Neural Electrodes for Recording Single Unit Activity and Electrocorticographic Signals

我々は、慢性神経記録電極を注入するプロセスを学習している外科医のための有用な情報を提供します。浸透と表面の両方電極システムの技術は、げっ歯類の動物モデルで説明されています。

単一ユニット活動とElectrocorticographic信号を記録するための慢性神経電極の外科的移植

The success of long-term electrophysiological recordings often depends on the quality of the implantation surgery.  Here we provide useful information for ...

surgical-implantation-chronic-neural-electrodes-for-recording-single

Nanyang Technological University

Research

JoVE Journal

Neuroscience

47.2K ビュー.  University of Michigan. 我々は、慢性神経記録電極を注入するプロセスを学習している外科医のための有用な情報を提供します。浸透と表面の両方電極システムの技術は、げっ歯類の動物モデルで説明されています。

ビデオ: Surgical Implantation of Chronic Neural Electrodes for Recording Single Unit Activity and Electrocorticographic Signals

Retrograde Fluorescent Labeling Allows for Targeted Extracellular Single-unit Recording from Identified Neurons In vivo

The overall goal of this method is to record single-unit responses from an identified population of neurons. In vivo electrophysiological recordings from individual neurons are critical for understanding how neural circuits function under natural conditions. Traditionally, these recordings have been performed 'blind', meaning the identity of the recorded cell is unknown at the start of the recording. Cellular identity can be subsequently determined via intracellular1, juxtacellular2 or loose-patch3 iontophoresis of dye, but these recordings cannot be pre-targeted to specific neurons in regions with functionally heterogeneous cell types. Fluorescent proteins can be expressed in a cell-type specific manner permitting visually-guided single-cell electrophysiology4-6. However, there are many model systems for which these genetic tools are not available. Even in genetically accessible model systems, the desired promoter may be unknown or genetically homogenous neurons may have varying projection patterns. Similarly, viral vectors have been used to label specific subgroups of projection neurons7, but use of this method is limited by toxicity and lack of trans-synaptic specificity. Thus, additional techniques that offer specific pre-visualization to record from identified single neurons in vivo are needed. Pre-visualization of the target neuron is particularly useful for challenging recording conditions, for which classical single-cell recordings are often prohibitively difficult8-11. The novel technique described in this paper uses retrograde transport of a fluorescent dye applied using tungsten needles to rapidly and selectively label a specific subset of cells within a particular brain region based on their unique axonal projections, thereby providing a visual cue to obtain targeted electrophysiological recordings from identified neurons in an intact circuit within a vertebrate CNS.
The most significant novel advancement of our method is the use of fluorescent labeling to target specific cell types in a non-genetically accessible model system. Weakly electric fish are an excellent model system for studying neural circuits in awake, behaving animals12. We utilized this technique to study sensory processing by "small cells" in the anterior exterolateral nucleus (ELa) of weakly electric mormyrid fish. "Small cells" are hypothesized to be time comparator neurons important for detecting submillisecond differences in the arrival times of presynaptic spikes13. However, anatomical features such as dense myelin, engulfing synapses, and small cell bodies have made it extremely difficult to record from these cells using traditional methods11, 14. Here we demonstrate that our novel method selectively labels these cells in 28% of preparations, allowing for reliable, robust recordings and characterization of responses to electrosensory stimulation.

Retrograde Fluorescent Labeling Allows for Targeted Extracellular Single-unit Recording from Identified Neurons In vivo

Retrograde transport of fluorescent dye labels a sub-population of neurons based on anatomical projection. Labeled axons can be visually targeted in vivo, permitting extracellular recording from identified axons. This technique facilitates recording when neurons cannot be labeled through genetic manipulation or are difficult to isolate using 'blind' in vivo approaches.

逆行性蛍光標識は、特定された神経細胞から標的細胞外シングルユニット記録を可能にします<em生体内で&gt;</em

The overall goal of this method is to record single-unit responses from an identified population of neurons. In vivo electrophysiological recordings from ...

神経科学

Recording Human Electrocorticographic (ECoG) Signals for Neuroscientific Research and Real-time Functional Cortical Mapping

Neuroimaging studies of human cognitive, sensory, and motor processes are usually based on noninvasive techniques such as electroencephalography (EEG), magnetoencephalography or functional magnetic-resonance imaging. These techniques have either inherently low temporal or low spatial resolution, and suffer from low signal-to-noise ratio and/or poor high-frequency sensitivity. Thus, they are suboptimal for exploring the short-lived spatio-temporal dynamics of many of the underlying brain processes. In contrast, the invasive technique of electrocorticography (ECoG) provides brain signals that have an exceptionally high signal-to-noise ratio, less susceptibility to artifacts than EEG, and a high spatial and temporal resolution (i.e., &lt;1 cm/&lt;1 millisecond, respectively). ECoG involves measurement of electrical brain signals using electrodes that are implanted subdurally on the surface of the brain. Recent studies have shown that ECoG amplitudes in certain frequency bands carry substantial information about task-related activity, such as motor execution and planning1, auditory processing2 and visual-spatial attention3. Most of this information is captured in the high gamma range (around 70-110 Hz). Thus, gamma activity has been proposed as a robust and general indicator of local cortical function1-5. ECoG can also reveal functional connectivity and resolve finer task-related spatial-temporal dynamics, thereby advancing our understanding of large-scale cortical processes. It has especially proven useful for advancing brain-computer interfacing (BCI) technology for decoding a user's intentions to enhance or improve communication6 and control7. Nevertheless, human ECoG data are often hard to obtain because of the risks and limitations of the invasive procedures involved, and the need to record within the constraints of clinical settings. Still, clinical monitoring to localize epileptic foci offers a unique and valuable opportunity to collect human ECoG data. We describe our methods for collecting recording ECoG, and demonstrate how to use these signals for important real-time applications such as clinical mapping and brain-computer interfacing. Our example uses the BCI2000 software platform8,9 and the SIGFRIED10 method, an application for real-time mapping of brain functions. This procedure yields information that clinicians can subsequently use to guide the complex and laborious process of functional mapping by electrical stimulation.
Prerequisites and Planning:
Patients with drug-resistant partial epilepsy may be candidates for resective surgery of an epileptic focus to minimize the frequency of seizures. Prior to resection, the patients undergo monitoring using subdural electrodes for two purposes: first, to localize the epileptic focus, and second, to identify nearby critical brain areas (i.e., eloquent cortex) where resection could result in long-term functional deficits. To implant electrodes, a craniotomy is performed to open the skull. Then, electrode grids and/or strips are placed on the cortex, usually beneath the dura. A typical grid has a set of 8 x 8 platinum-iridium electrodes of 4 mm diameter (2.3 mm exposed surface) embedded in silicon with an inter-electrode distance of 1cm. A strip typically contains 4 or 6 such electrodes in a single line. The locations for these grids/strips are planned by a team of neurologists and neurosurgeons, and are based on previous EEG monitoring, on a structural MRI of the patient's brain, and on relevant factors of the patient's history. Continuous recording over a period of 5-12 days serves to localize epileptic foci, and electrical stimulation via the implanted electrodes allows clinicians to map eloquent cortex. At the end of the monitoring period, explantation of the electrodes and therapeutic resection are performed together in one procedure.
In addition to its primary clinical purpose, invasive monitoring also provides a unique opportunity to acquire human ECoG data for neuroscientific research. The decision to include a prospective patient in the research is based on the planned location of their electrodes, on the patient's performance scores on neuropsychological assessments, and on their informed consent, which is predicated on their understanding that participation in research is optional and is not related to their treatment. As with all research involving human subjects, the research protocol must be approved by the hospital's institutional review board. The decision to perform individual experimental tasks is made day-by-day, and is contingent on the patient's endurance and willingness to participate. Some or all of the experiments may be prevented by problems with the clinical state of the patient, such as post-operative facial swelling, temporary aphasia, frequent seizures, post-ictal fatigue and confusion, and more general pain or discomfort.
At the Epilepsy Monitoring Unit at Albany Medical Center in Albany, New York, clinical monitoring is implemented around the clock using a 192-channel Nihon-Kohden Neurofax monitoring system. Research recordings are made in collaboration with the Wadsworth Center of the New York State Department of Health in Albany. Signals from the ECoG electrodes are fed simultaneously to the research and the clinical systems via splitter connectors. To ensure that the clinical and research systems do not interfere with each other, the two systems typically use separate grounds. In fact, an epidural strip of electrodes is sometimes implanted to provide a ground for the clinical system. Whether research or clinical recording system, the grounding electrode is chosen to be distant from the predicted epileptic focus and from cortical areas of interest for the research. Our research system consists of eight synchronized 16-channel g.USBamp amplifier/digitizer units (g.tec, Graz, Austria). These were chosen because they are safety-rated and FDA-approved for invasive recordings, they have a very low noise-floor in the high-frequency range in which the signals of interest are found, and they come with an SDK that allows them to be integrated with custom-written research software. In order to capture the high-gamma signal accurately, we acquire signals at 1200Hz sampling rate-considerably higher than that of the typical EEG experiment or that of many clinical monitoring systems. A built-in low-pass filter automatically prevents aliasing of signals higher than the digitizer can capture. The patient's eye gaze is tracked using a monitor with a built-in Tobii T-60 eye-tracking system (Tobii Tech., Stockholm, Sweden). Additional accessories such as joystick, bluetooth Wiimote (Nintendo Co.), data-glove (5th Dimension Technologies), keyboard, microphone, headphones, or video camera are connected depending on the requirements of the particular experiment.
Data collection, stimulus presentation, synchronization with the different input/output accessories, and real-time analysis and visualization are accomplished using our BCI2000 software8,9. BCI2000 is a freely available general-purpose software system for real-time biosignal data acquisition, processing and feedback. It includes an array of pre-built modules that can be flexibly configured for many different purposes, and that can be extended by researchers' own code in C++, MATLAB or Python. BCI2000 consists of four modules that communicate with each other via a network-capable protocol: a Source module that handles the acquisition of brain signals from one of 19 different hardware systems from different manufacturers; a Signal Processing module that extracts relevant ECoG features and translates them into output signals; an Application module that delivers stimuli and feedback to the subject; and the Operator module that provides a graphical interface to the investigator.
A number of different experiments may be conducted with any given patient. The priority of experiments will be determined by the location of the particular patient's electrodes. However, we usually begin our experimentation using the SIGFRIED (SIGnal modeling For Realtime Identification and Event Detection) mapping method, which detects and displays significant task-related activity in real time. The resulting functional map allows us to further tailor subsequent experimental protocols and may also prove as a useful starting point for traditional mapping by electrocortical stimulation (ECS).
Although ECS mapping remains the gold standard for predicting the clinical outcome of resection, the process of ECS mapping is time consuming and also has other problems, such as after-discharges or seizures. Thus, a passive functional mapping technique may prove valuable in providing an initial estimate of the locus of eloquent cortex, which may then be confirmed and refined by ECS. The results from our passive SIGFRIED mapping technique have been shown to exhibit substantial concurrence with the results derived using ECS mapping10.
The protocol described in this paper establishes a general methodology for gathering human ECoG data, before proceeding to illustrate how experiments can be initiated using the BCI2000 software platform. Finally, as a specific example, we describe how to perform passive functional mapping using the BCI2000-based SIGFRIED system.

Recording Human Electrocorticographic ECoG Signals for Neuroscientific Research and Real-time Functional Cortical Mapping

We present a method for collecting electrocorticographic signals for research purposes from humans who are undergoing invasive epilepsy monitoring. We show how to use the BCI2000 software platform for data collection, signal processing and stimulus presentation. Specifically, we demonstrate SIGFRIED, a BCI2000-based tool for real-time functional brain mapping.

神経科学の研究とリアルタイム機能皮質マッピングのためのヒトElectrocorticographic（ECOG）信号を記録

Neuroimaging studies of human cognitive, sensory, and motor processes are usually based on noninvasive techniques such as electroencephalography (EEG), ...

Optical Recording of Suprathreshold Neural Activity with Single-cell and Single-spike Resolution

Signaling of information in the vertebrate central nervous system is often carried by populations of neurons rather than individual neurons. Also propagation of suprathreshold spiking activity involves populations of neurons. Empirical studies addressing cortical function directly thus require recordings from populations of neurons with high resolution. Here we describe an optical method and a deconvolution algorithm to record neural activity from up to 100 neurons with single-cell and single-spike resolution. This method relies on detection of the transient increases in intracellular somatic calcium concentration associated with suprathreshold electrical spikes (action potentials) in cortical neurons. High temporal resolution of the optical recordings is achieved by a fast random-access scanning technique using acousto-optical deflectors (AODs)1. Two-photon excitation of the calcium-sensitive dye results in high spatial resolution in opaque brain tissue2. Reconstruction of spikes from the fluorescence calcium recordings is achieved by a maximum-likelihood method. Simultaneous electrophysiological and optical recordings indicate that our method reliably detects spikes (&gt;97% spike detection efficiency), has a low rate of false positive spike detection (&lt; 0.003 spikes/sec), and a high temporal precision (about 3 msec) 3. This optical method of spike detection can be used to record neural activity in vitro and in anesthetized animals in vivo3,4.

Understanding the function of the vertebrate central nervous system requires recordings from many neurons because cortical function arises on the level of populations of neurons. Here we describe an optical method to record suprathreshold neural activity with single-cell and single-spike resolution, dithered random-access scanning. This method records somatic fluorescence calcium signals from up to 100 neurons with high temporal resolution. A maximum-likelihood algorithm deconvolves the underlying suprathreshold neural activity from the somatic fluorescence calcium signals. This method reliably detects spikes with high detection efficiency and a low rate of false positives and can be used to study neural populations in vitro and in vivo.

単一セルと単スパイク分解能で閾値上神経活動の光記録

Signaling of information in the vertebrate central nervous system is often carried by populations of neurons rather than individual neurons. Also ...

Fiber-optic Implantation for Chronic Optogenetic Stimulation of Brain Tissue

Elucidating patterns of neuronal connectivity has been a challenge for both clinical and basic neuroscience. Electrophysiology has been the gold standard for analyzing patterns of synaptic connectivity, but paired electrophysiological recordings can be both cumbersome and experimentally limiting. The development of optogenetics has introduced an elegant method to stimulate neurons and circuits, both in vitro1 and in vivo2,3. By exploiting cell-type specific promoter activity to drive opsin expression in discrete neuronal populations, one can precisely stimulate genetically defined neuronal subtypes in distinct circuits4-6. Well described methods to stimulate neurons, including electrical stimulation and/or pharmacological manipulations, are often cell-type indiscriminate, invasive, and can damage surrounding tissues. These limitations could alter normal synaptic function and/or circuit behavior. In addition, due to the nature of the manipulation, the current methods are often acute and terminal. Optogenetics affords the ability to stimulate neurons in a relatively innocuous manner, and in genetically targeted neurons. The majority of studies involving in vivo optogenetics currently use a optical fiber guided through an implanted cannula6,7; however, limitations of this method include damaged brain tissue with repeated insertion of an optical fiber, and potential breakage of the fiber inside the cannula. Given the burgeoning field of optogenetics, a more reliable method of chronic stimulation is necessary to facilitate long-term studies with minimal collateral tissue damage. Here we provide our modified protocol as a video article to complement the method effectively and elegantly described in Sparta et al.8 for the fabrication of a fiber optic implant and its permanent fixation onto the cranium of anesthetized mice, as well as the assembly of the fiber optic coupler connecting the implant to a light source. The implant, connected with optical fibers to a solid-state laser, allows for an efficient method to chronically photostimulate functional neuronal circuitry with less tissue damage9 using small, detachable, tethers. Permanent fixation of the fiber optic implants provides consistent, long-term in vivo optogenetic studies of neuronal circuits in awake, behaving mice10 with minimal tissue damage.

The development of optogenetics now provides the means to precisely stimulate genetically defined neurons and circuits, both in vitro and in vivo. Here we describe the assembly and implantation of a fiber optic for chronic photostimulation of brain tissue.

脳組織の慢性Optogenetic刺激のための光ファイバ注入

Elucidating patterns of neuronal connectivity has been a challenge for both clinical and basic neuroscience. Electrophysiology has been the gold standard ...

Multi-unit Recording Methods to Characterize Neural Activity in the Locust (Schistocerca Americana) Olfactory Circuits

Detection and interpretation of olfactory cues are critical for the survival of many organisms. Remarkably, species across phyla have strikingly similar olfactory systems suggesting that the biological approach to chemical sensing has been optimized over evolutionary time1. In the insect olfactory system, odorants are transduced by olfactory receptor neurons (ORN) in the antenna, which convert chemical stimuli into trains of action potentials. Sensory input from the ORNs is then relayed to the antennal lobe (AL; a structure analogous to the vertebrate olfactory bulb). In the AL, neural representations for odors take the form of spatiotemporal firing patterns distributed across ensembles of principal neurons (PNs; also referred to as projection neurons)2,3. The AL output is subsequently processed by Kenyon cells (KCs) in the downstream mushroom body (MB), a structure associated with olfactory memory and learning4,5. Here, we present electrophysiological recording techniques to monitor odor-evoked neural responses in these olfactory circuits.
First, we present a single sensillum recording method to study odor-evoked responses at the level of populations of ORNs6,7. We discuss the use of saline filled sharpened glass pipettes as electrodes to extracellularly monitor ORN responses. Next, we present a method to extracellularly monitor PN responses using a commercial 16-channel electrode3. A similar approach using a custom-made 8-channel twisted wire tetrode is demonstrated for Kenyon cell recordings8. We provide details of our experimental setup and present representative recording traces for each of these techniques.

Multi-unit Recording Methods to Characterize Neural Activity in the Locust Schistocerca Americana Olfactory Circuits

We demonstrate variations of the extracellular multi-unit recording technique to characterize odor-evoked responses in the first three stages of the invertebrate olfactory pathway. These techniques can easily be adapted to examine ensemble activity in other neural systems as well.

ローカスの神経活動を特徴づけるためにマルチユニット記録方法（<em&gt; Schistocercaアメリカーナ</em&gt;）嗅覚回路

Detection and interpretation of olfactory cues are critical for the survival  of many organisms. Remarkably, species across phyla have strikingly similar ...

A Method for Systematic Electrochemical and Electrophysiological Evaluation of Neural Recording Electrodes

New materials and designs for neural implants are typically tested separately, with a demonstration of performance but without reference to other implant characteristics.&#160;This precludes a rational selection of a particular implant as optimal for a particular application and the development of new materials based on the most critical performance parameters.&#160;This article develops a protocol for in vitro and in vivo testing of neural recording electrodes.&#160;Recommended parameters for electrochemical and electrophysiological testing are documented with the key steps and potential issues discussed.&#160;This method eliminates or reduces the impact of many systematic errors present in simpler in vivo testing paradigms, especially variations in electrode/neuron distance and between animal models.&#160;The result is a strong correlation between the critical in vitro and in vivo responses, such as impedance and signal-to-noise ratio.&#160;This protocol can easily be adapted to test other electrode materials and designs.&#160;The in vitro techniques can be expanded to any other nondestructive method to determine further important performance indicators.&#160;The principles used for the surgical approach in the auditory pathway can also be modified to other neural regions or tissue.

Different electrode coatings affect neural recording performance through changes to electrochemical, chemical and mechanical properties. Comparison of electrodes in vitro is relatively simple, however comparison of in vivo response is typically complicated by variations in electrode/neuron distance and between animals. This article provides a robust method to compare neural recording electrodes.

神経電位電極の体系的電気化学的および電気生理学的評価方法

New materials and designs for neural implants are typically tested separately, with a demonstration of performance but without reference to other implant ...

A Guide to In vivo Single-unit Recording from Optogenetically Identified Cortical Inhibitory Interneurons

A major challenge in neurophysiology has been to characterize the response properties and function of the numerous inhibitory cell types in the cerebral cortex. We here share our strategy for obtaining stable, well-isolated single-unit recordings from identified inhibitory interneurons in the anesthetized mouse cortex using a method developed by Lima and colleagues1. Recordings are performed in mice expressing Channelrhodopsin-2 (ChR2) in specific neuronal subpopulations. Members of the population are identified by their response to a brief flash of blue light. This technique &#8211; termed &#8220;PINP&#8221;, or Photostimulation-assisted Identification of Neuronal Populations &#8211; can be implemented with standard extracellular recording equipment. It can serve as an inexpensive and accessible alternative to calcium imaging or visually-guided patching, for the purpose of targeting extracellular recordings to genetically-identified cells. Here we provide a set of guidelines for optimizing the method in everyday practice. We refined our strategy specifically for targeting parvalbumin-positive (PV+) cells, but have found that it works for other interneuron types as well, such as somatostatin-expressing (SOM+) and calretinin-expressing (CR+) interneurons.

A Guide to In vivo Single-unit Recording from Optogenetically Identified Cortical Inhibitory Interneurons

Here we describe our strategy for obtaining stable, well-isolated single-unit recordings from identified inhibitory interneurons in the anesthetized mouse cortex. Neurons expressing ChR2 are identified by their response to blue light. The method uses standard extracellular recording equipment, and serves as an inexpensive alternative to calcium imaging or visually-guided patching.

へのガイド<em&gt;インビボ</emOptogenetically同定された皮質抑制性介在から&gt;シングル単位記録

A major challenge in neurophysiology has been to characterize the response properties and function of the numerous inhibitory cell types in the cerebral ...

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.

覚醒マウスにおける神経活性物質のMicroiontophoreticアプリケーションと組み合わせることで神経活動の細胞外記録

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

Fabrication of High Contact-Density, Flat-Interface Nerve Electrodes for Recording and Stimulation Applications

Many attempts have been made to manufacture multi-contact nerve cuff electrodes that are safe, robust and reliable for long term neuroprosthetic applications. This protocol describes a fabrication technique of a modified cylindrical nerve cuff electrode to meet these criteria. Minimum computer-aided design and manufacturing (CAD and CAM) skills are necessary to consistently produce cuffs with high precision (contact placement 0.51 &#177; 0.04 mm) and various cuff sizes. The precision in spatially distributing the contacts and the ability to retain a predefined geometry accomplished with this design are two criteria essential to optimize the cuff&#39;s interface for selective recording and stimulation. The presented design also maximizes the flexibility in the longitudinal direction while maintaining sufficient rigidity in the transverse direction to reshape the nerve by using materials with different elasticities. The expansion of the cuff&#39;s cross sectional area as a result of increasing the pressure inside the cuff was observed to be 25% at 67 mm Hg. This test demonstrates the flexibility of the cuff and its response to nerve swelling post-implant. The stability of the contacts&#39; interface and recording quality were also examined with contacts&#39; impedance and signal-to-noise ratio metrics from a chronically implanted cuff (7.5 months), and observed to be 2.55 &#177; 0.25 k&#8486; and 5.10 &#177; 0.81 dB respectively.

This article provides a detailed description on the fabrication process of a high contact-density flat interface nerve electrode (FINE). This electrode is optimized for recording and stimulating neural activity selectively within peripheral nerves.

記録と刺激アプリケーションの高接触密度、フラットインターフェイスの神経電極の作製

Many attempts have been made to manufacture multi-contact nerve cuff electrodes that are safe, robust and reliable for long term neuroprosthetic ...

Implantation of Chronic Silicon Probes and Recording of Hippocampal Place Cells in an Enriched Treadmill Apparatus

An important requisite for understanding brain function is the identification of behavior and cell activity correlates. Silicon probes are advanced electrodes for large-scale electrophysiological recording of neuronal activity, but the procedures for their chronic implantation are still underdeveloped. The activity of hippocampal place cells is known to correlate with an animal&#39;s position in the environment, but the underlying mechanisms are still unclear. To investigate place cells, here we describe a set of techniques which range from the fabrication of devices for chronic silicon probe implants to the monitoring of place field activity in a cue-enriched treadmill apparatus. A micro-drive and a hat are built by fitting and fastening together 3D-printed plastic parts. A silicon probe is mounted on the micro-drive, cleaned, and coated with dye. A first surgery is performed to fix the hat on the skull of a mouse. Small landmarks are fabricated and attached to the belt of a treadmill. The mouse is trained to run head-fixed on the treadmill. A second surgery is performed to implant the silicon probe in the hippocampus, following which broadband electrophysiological signals are recorded. Finally, the silicon probe is recovered and cleaned for reuse. The analysis of place cell activity in the treadmill reveals a diversity of place field mechanisms, outlining the benefit of the approach.

We describe the diverse steps to implant chronic silicon probes and to record place cells in mice that are running head-fixed on a cue-enriched treadmill apparatus.

慢性的なシリコン プローブと豊かなトレッドミル装置海馬場所細胞の記録

An important requisite for understanding brain function is the identification of behavior and cell activity correlates. Silicon probes are advanced ...