Name: an anesthesia, surgery, and harvest method for the evaluation of transpedicular screws using an in vivo porcine lumbar spine model
Uploaded: 2017-05-31T12:30:00.000+00:00
Duration: 9 min 7 s
Description: Watch this Scientific Journal Video about An Anesthesia, Surgery, and Harvest Method for the Evaluation of Transpedicular Screws Using an In Vivo Porcine Lumbar Spine Model at JoVE.com

この研究は、韓国の全北大学病院生物医学研究所（CNUH-BRI）の資金提供による助成金（CNUH-BRI-2012-02-005）によって支持された。

全北国立大学法人化審査委員会がこの研究を承認した。動物の治療、使用、取り扱いは、すべてのガイドラインとポリシーに従った。 24℃で手術室を維持する。 1.麻酔<ol><li>少なくとも1週間、実験単位で12カ月齢の小型ブタを順化させる。呼吸数、心拍数、体温を測定する臨床検査を行う。麻酔処置の12時間前に各小型豚に餌を与えないでください。 </li><li>前投与のために、耳の後ろ側の子宮頸部筋領域にアトロピン（0.05mg / kg）およびケタミン（20mg / kg）/キシラキシン（2mg / kg）を注入する。 </li><li>鎮静後、耳の根元にゴムバンドをしっかりと掛けて、局所アルコールで耳をきれいにしてください。 </li><li>オーバー・ザ・ニードルのプラスチックカテーテルを耳静脈に置き、ゴムバンドを外す。カテーテルが正しく配置されていることを確認します。インフルエンザカテーテルをヘパリン処理した生理食塩水で洗浄し、テープで固定する。 </li><li> 気管内挿管のために、小型豚を胸骨の臥位に配置する。アシスタントの助けを借りて、適切なスリングで豚の顎を持ち、口を開ける。 <ol><li>喉頭鏡の先端を咽頭腔に通して、喉頭蓋を軟口蓋から押し退ける。喉頭鏡の先端を使って声帯を見えるようにし、呼気中気管内チューブを気管に進める。 </li><li>適切な挿管のための空気の自由な通過を感じ、ミニブタの左右両側の呼吸音の胸部聴診を確認する。 </li></ol></li><li> 10 mLの注射器を使用して気管内チューブの空気の適切な容量の空気でカフを満たし、接着テープを使用してチューブをスナウトに固定する。 </li><li>長期間の麻酔のための吸入麻酔剤であるイソフルラン2.0％濃度をエンドトールを通して提供する痛みのある挿管。角膜および眼球の反射を検査して麻酔を確認し、眼に軟膏を使用して乾燥を防ぐ。 </li><li>ミニブタが回復するまで少なくとも5分ごとに麻酔中の心臓血管系、呼吸器系および体温を監視する。 </li><li>脊髄手術の30分前に、セファロスポリン抗生物質の第1世代であるセファゾリンIVを30mg / kgでゆっくりと投与する。 </li><li>恒常性を維持するためにIVラインを用いて5〜10 mL / kg /時の温（37℃）生理食塩水を投与し、痛みを抑制するために50μg/ kg /分のフェンタニルを投与する。 </li><li>脊椎手術の後、強い嚥下反射が明らかな場合には抜管を行う。 </li><li>ミニ豚を部屋に持ち込み、麻酔から回復するまでモニターします。ミニブタが完全に意識しているときには、食べ物と水を提供してください。 </li><li>最初の3日間の疼痛管理のために、毎日4.4mg / kgのカルプロフェンを含む3mg / kgのエンロフロキサシン抗生物質を適用する。 </li><li> Moni縫合糸が取り除かれるまで毎日ブタを服用してください。 </li></ol> 2.脊椎手術<ol><li>製造業者のガイドラインに従って、透過性スクリューおよび後固定器システムをオートクレーブして滅菌する。 </li><li>ブタが仰臥位にある間にシェーバーを使用して、中央から左または右に約10cmの小型ブタの背部を剃る。ポビドンヨード溶液と70％アルコールで皮膚を洗浄する。 </li><li>メスを使用して、腰椎の第2の棘突起から第1の正中仙骨紋に縦中心線切開を行う。皮下組織および筋膜を棘突起の先端まで解剖する。 </li><li> Cobbエレベーターを用いて、下層の小層から傍脊髄筋を腹膜下に上昇させる。棘状突起に沿って解剖し、椎弓板を椎間関節に限定する。 </li><li>エントリーポイントの表在皮質を開く（これはL3から哺乳類のプロセスにちょうど劣る）両側のL5まで）にバリまたはロングガールを付けます。 </li><li>ガイドピンを開いた場所、上端板と平行に、棘突起に対して20°の角度で挿入します。 Cアームまたはポータブルポストロ - 前方/側方X線を使用して理想的な出発点を定義します。 </li><li> X線に従ってペディクルプローブを最大25 mm挿入します。椎弓根の鳴動装置を用いて、椎弓根および体の触診によって完全な骨内軌跡を確認する。 </li><li>準備されたL3〜L5の椎弓根に6つの椎弓根スクリューを差し込み、ネジ頭がしっかりと固定されるようにします。インプラントヘッドの側面開口部を所望の方向に向け、水平位置をロッド軌道と整列させる。 </li><li>茎ネジ頭の両側にそれぞれ2本のロッドを挿入します。ユニバーサルハンドルを使用してペディクルスクリューヘッドのスリーブとナットを入れます。 </li><li>ストレートソケットレンチを使用してナットを緩く締め、カウンタートルクレンチを使用してナットをしっかり締めます。 </li><li>ポジションを確認するポータブルな前後前後のX線を使用した茎ねじ。 注：ミニブタが目を覚ますのを待って、後ろ足の歩行パターンと運動機能をチェックして、ねじがひどく植え込まれているかどうかを判断します。 </li><li>吸引を伴うバルブ洗浄シリンジを使用して、手術部位を通常の生理食塩水3Lで灌注する。 </li><li>手術部位にシリコーンを流し、シリコンチップを取り出します。 1.0メートルの吸収可能な縫合糸を用いて、傍脊髄の筋肉および皮下を閉じる。非吸収性ナイロン縫合糸2.0メートルで皮膚を閉じます。 </li><li>ポビドンヨードで縫合部位を消毒し、滅菌ガーゼおよびテープを用いて包帯を適用する。 </li></ol>収穫手続き<ol><li>術後12週目に、キシラジン（2mg / kg）およびケタミン（10mg / kg）を耳の後ろ側の子宮頸部筋領域に注射し、前投薬する。 </li><li>鎮静後、耳静脈に直接15mg / kgのKClを投与する安楽死のためのカテーテル。 </li><li>以前の手術瘢痕病変で縦中心線切開を行う。軟組織と傍脊髄の筋肉を解剖する。 </li><li>腰椎、薄層板、ロッド、ナット、および横断プロセスの棘突起をL3からL5に露出させる。 </li><li>カウンタートルクレンチとロッドを使用してナットを取り外します。振動刃を使用して、L2-3ディスクスペースとL5-S1ディスクスペースをカットします。 </li><li>コブエレベーターとタワー鉗子でL3-5脊柱の中部と前部の両側を解剖する。収穫後、直ちに脊椎を検査することができない場合は、食塩水を浸したガーゼで包んで-20℃で保存する。 </li></ol>

<ol>
	<li>Greenfield, R. T., Grant, R. E., Bryant, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pedicle+screw+fixation+in+the+management+of+unstable+thoracolumbar+spine+injuries.">Pedicle screw fixation in the management of unstable thoracolumbar spine injuries.</a> Orthop Rev. 21 (6), 701-706 (1992).</li><li>Upasani, V. V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pedicle+screw+surface+coatings+improve+fixation+in+nonfusion+spinal+constructs.">Pedicle screw surface coatings improve fixation in nonfusion spinal constructs.</a> Spine. 34 (4), 335-343 (2009).</li><li>Halvorson, T. L., Kelley, L. A., Thomas, K. A., Whitecloud, T. S., Cook, S. 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Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evaluation+of+Titanium-Coated+Pedicle+Screws:+In+Vivo+Porcine+Lumbar+Spine+Model.">Evaluation of Titanium-Coated Pedicle Screws: In Vivo Porcine Lumbar Spine Model.</a> World Neurosurg. 91, 163-171 (2016).</li><li>Upasani, V. V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pedicle+screw+surface+coatings+improve+fixation+in+nonfusion+spinal+constructs.">Pedicle screw surface coatings improve fixation in nonfusion spinal constructs.</a> Spine. 34 (4), 335-343 (2009).</li><li>Smit, T. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+use+of+a+quadrupted+as+an+in+vivo+model+for+the+study+of+the+spine-biomechanical+considrations.">The use of a quadrupted as an in vivo model for the study of the spine-biomechanical considrations.</a> Eur Spine J. 11 (2), 137-144 (2002).</li><li>Aldini, N. N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pedicular+fixation+in+the+osteoporotic+spine:+a+pilot+in+vivo+study+on+long-term+ovariectomized+sheep.">Pedicular fixation in the osteoporotic spine: a pilot in vivo study on long-term ovariectomized sheep.</a> J Orthop Res. 20 (6), 1217-1224 (2002).</li><li>Fini, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biological+assessment+of+the+bone-screw+interface+after+insertion+of+uncoated+and+hydroxyapatite-coated+pedicular+screws+in+the+osteopenic+sheep.">Biological assessment of the bone-screw interface after insertion of uncoated and hydroxyapatite-coated pedicular screws in the osteopenic sheep.</a> J Biomed Mater Res A. 66 (1), 176-183 (2003).</li><li>Branemark, R., Ohrnell, L. O., Skalak, R., Carlsson, L., Branemark, P. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biomechanical+characterization+of+osseointegration:+an+experimental+in+vivo+investigation+in+the+beagle+dog.">Biomechanical characterization of osseointegration: an experimental in vivo investigation in the beagle dog.</a> J Orthop Res. 16 (1), 61-69 (1998).</li><li>McLain, R. F., Yerby, S. A., Moseley, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparative+morphometry+of+L4+vertebrae:+comparison+of+large+animal+models+for+the+human+lumbar+spine.">Comparative morphometry of L4 vertebrae: comparison of large animal models for the human lumbar spine.</a> Spine. 27 (8), E200-E206 (2002).</li><li>Giavaresi, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+preclinical+evaluation+of+the+influence+of+osteoporosis+on+the+anchorage+of+different+pedicle+screw+designs.">In vivo preclinical evaluation of the influence of osteoporosis on the anchorage of different pedicle screw designs.</a> Eur Spine J. 20 (8), 1289-1296 (2011).</li><li>Hasegawa, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hydroxyapatite-coating+of+pedicle+screws+improves+resistance+against+pull-out+force+in+the+osteoporotic+canine+lumbar+spine+model:+a+pilot+study.">Hydroxyapatite-coating of pedicle screws improves resistance against pull-out force in the osteoporotic canine lumbar spine model: a pilot study.</a> Spine J. 5 (3), 239-243 (2005).</li><li>Smorgick, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Single-+versus+multilevel+fusion+for+single-level+degenerative+spondylolisthesis+and+multilevel+lumbar+stenosis:+four-year+results+of+the+spine+patient+outcomes+research+trial.">Single- versus multilevel fusion for single-level degenerative spondylolisthesis and multilevel lumbar stenosis: four-year results of the spine patient outcomes research trial.</a> Spine. 38 (10), 797-805 (2013).</li><li>Busscher, I., Ploegmakers, J. J., Verkerke, G. J., Veldhuizen, A. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparative+anatomical+dimensions+of+the+complete+human+and+porcine+spine.">Comparative anatomical dimensions of the complete human and porcine spine.</a> Eur Spine J. 19 (7), 1104-1114 (2010).</li></ol>

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ブタ背骨における経直腸スクリューの評価は、多くの時間と労力を必要とする。まず、ミニブタは大きな動物です。動物のケアと麻酔については、研究者は特別なプロトコルが必要です。第二に、手術は人間の手術と同様の環境を維持すべきである。ブタ背骨の椎弓根スクリューを評価する目的は、ヒトに適用できる効率的なスクリューを開発することです。第3に、経直腸ねじの長期安定性...

Transpedicular screw fixation is a gold-standard treatment for degenerative lumbar spine and bursting fracture because it involves three columns of the spine and achieves stabilization1,2. However, most patients who undergo such surgery also have osteoporosis3,4. Many studies have evaluated the fixation strength and the osseointegration status of transpedicular screws, because the loosening of pedicle screws currently in use has been reported in patients with osteoporosis5,6.
The porcine spine is similar to the human spine in terms of size. It is less expensive compared to a primate model7. Furthermore, an in vivo mechanical study has demonstrated that the quadruped porcine spine is essentially loaded in the same way as that of the human spine8, which is why many researchers use porcine spines for studies on the prevention of pedicle screw loosening. However, it takes several months to study pedicle screws in the porcine spine because identifying the long-term stability of pedicle screws takes times. In order to compare different types of screws in the vertebral body, it is necessary to insert the screws in similar positions. Therefore, researchers should be well-acquainted with proper anesthesia techniques, standardized surgical protocols, and harvest procedures before performing any experiments. Here, we describe a detailed method for anesthesia, surgery, and harvest for the evaluation of pedicle screw fixation using a porcine spine model, including ex vivo imaging, histology, and strength testing.

<table><tbody><tr><td>Miniature pig</td><td>OrientBio</td><td></td><td></td></tr><tr><td>Atropine</td><td>Jeil pharmaceutical</td><td>A04900241</td><td>Anesthesia</td></tr><tr><td>Over-the needle plastic catheter</td><td>BD</td><td>REF382412</td><td>Maintenance of IV line</td></tr><tr><td>Ketamine</td><td>Yuhan</td><td>A04502441</td><td>Anesthesia</td></tr><tr><td>Xylazine</td><td>Bayer Korea</td><td>A00800071</td><td>Anesthesia</td></tr><tr><td>Laryngoscope</td><td>Karl storz</td><td></td><td>Intubation</td></tr><tr><td>Endotracheal tube</td><td>Covidien</td><td></td><td>Intubation</td></tr><tr><td>Isoflurane</td><td>JW pharmaceutical Co</td><td>A02104781</td><td>Anesthesia</td></tr><tr><td>Eye ointment</td><td>Hanlim pharma</td><td>A37851721</td><td>Protection of pig&#039;s eye</td></tr><tr><td>Cefazolin</td><td>Donga pharma</td><td>A01503951</td><td>Antibiotics</td></tr><tr><td>Saline</td><td>JW pharmaceutical Co</td><td>A02151392</td><td>Maintenance of fluid homeostasis</td></tr><tr><td>Fentanyl</td><td>Hana pharm</td><td>C03200032</td><td>Pain control</td></tr><tr><td>Enrofloxacin</td><td>Bayer</td><td>93106-60-6&amp;nbsp;</td><td>Antibiotics</td></tr><tr><td>Morphine</td><td>Myungmoon pharma</td><td>C03700091</td><td>Pain control</td></tr><tr><td>Meloxicam</td><td>Boehringer Ingelheim</td><td>A07600711</td><td>Antibiotics</td></tr><tr><td>Povidone-iodine</td><td>Hyundai pharma</td><td></td><td>Wound dressing</td></tr><tr><td>Scalpel blade size 15</td><td>Braun</td><td>&amp;nbsp;I1 BB515</td><td>Skin incision</td></tr><tr><td>Cobb elevator</td><td>Codman</td><td>65-2546</td><td>Dissection of muscle</td></tr><tr><td>Burr</td><td>Medtronic</td><td></td><td>Making of starting point of screw</td></tr><tr><td>Rongeur</td><td>Aesculap</td><td>FO515R</td><td>Making of starting point of screw</td></tr><tr><td>Guide pin (K-wire)</td><td>CE</td><td>01067803</td><td>Guidance of screw trajectory</td></tr><tr><td>C-arm</td><td>GE</td><td>OEC 9800 plus</td><td>Guidance of screw trajectory</td></tr><tr><td>Portable X-ray</td><td>Siemens</td><td>Mobile XP hybrid</td><td>Guidance of screw trajectory</td></tr><tr><td>Pedicle probe</td><td>OtisBiotech</td><td>SPI-02-01</td><td>Guidance of screw trajectory</td></tr><tr><td>Pedicle sounding device</td><td>OtisBiotech</td><td>SPI-03-01</td><td>Guidance of screw trajectory</td></tr><tr><td>Pedicle screw</td><td>OtisBiotech</td><td>MS-40025</td><td></td></tr><tr><td>Posterior fixator systems</td><td>OtisBiotech</td><td></td><td></td></tr><tr><td>Rod&amp;nbsp;</td><td>OtisBiotech</td><td>ROD-60140</td><td>Rigid fixation between screws</td></tr><tr><td>Universal handle</td><td>OtisBiotech</td><td>SPI-08-01</td><td>To fix the screws to the rod</td></tr><tr><td>Straight socket wrench</td><td>OtisBiotech</td><td>SPI-06-01</td><td>To fix the screws to the rod</td></tr><tr><td>counter torque wrench</td><td>OtisBiotech</td><td>SPI-07-01</td><td>To fix the screws to the rod</td></tr><tr><td>Bulb irrigation syringe</td><td>Hyupsug medical</td><td>HS-IR-140</td><td>Irrigation</td></tr><tr><td>Silicone drain</td><td>Sewon medical</td><td>2205-006</td><td>To drain the fluid at the surgical site</td></tr><tr><td>3.0 metric absorbable suture</td><td>Ethicon</td><td>BA1673H</td><td>Muscle suture</td></tr><tr><td>2.0 metric nonabsorbable nylon suture</td><td>Ethicon</td><td>W1626T</td><td>Skin suture</td></tr><tr><td>Gauze</td><td>Kingphar Korea</td><td>KP120-06</td><td></td></tr><tr><td>Pentobarbital</td><td>Hanlim pharma</td><td>645301221</td><td>Euthanasia</td></tr><tr><td>Oscillating saw</td><td>Zimmer</td><td></td><td>Harvest spine</td></tr><tr><td>Tower forceps</td><td>Aesculap</td><td>BF461R</td><td>Harvest spine</td></tr></tbody></table>

an anesthesia, surgery, and harvest method for the evaluation of transpedicular screws using an in vivo porcine lumbar spine model

椎弓根スクリュー固定は、脊髄疾患の処置のためのゴールドスタンダードです。しかし、脊髄手術後の茎ねじ緩みの問題が多くの研究によって報告されており、これは深刻な問題である。この問題に対処するために、様々なタイプの椎弓根スクリューを検査し、脊椎骨における良好な固定強度および骨結合を有するものを同定した。ブタの脊椎は、解剖学的サイズ、機械的特性、およびコストのために椎弓根スクリューの評価における人間の背骨のための良い選択肢です。いくつかの研究では、椎弓根スクリューはブタモデルにおいて効率的であると報告されているが、ブタモデルを用いた椎弓根スクリューの評価のための詳細なプロトコールは記載されていない。ここでは、インビボブタ腰椎モデルを使用して経椎骨ねじを評価するための詳細な方法を説明する。ここで提供される麻酔、背骨手術、収穫の技術的な詳細は、ranspedicularスクリュー固定モデル。

インビボでのブタの腰椎モデルを用いた経椎間膜スクリューの評価のための麻酔、手術、および収穫の詳細なプロトコールをここに記載する。このプロトコルは、機械的試験（ 図1 ）、定量的マイクロCT評価（ 図2 ）、および組織学（ 図3 ）を含む多くの下流分析に適しています。代表的な機械的試験（ 図1 ?...

Watch this Scientific Journal Video about An Anesthesia, Surgery, and Harvest Method for the Evaluation of Transpedicular Screws Using an In Vivo Porcine Lumbar Spine Model at JoVE.com

An Anesthesia, Surgery, and Harvest Method for the Evaluation of Transpedicular Screws Using an In Vivo Porcine Lumbar Spine Model

ここでは、 インビボでのブタの腰椎モデルを用いて経椎骨のネジを評価する方法を紹介する。

麻酔、手術および収縮法を用いた経肛門用ねじの評価<em&gt;インビボ</em&gt;豚の腰椎モデル

Pedicle screw fixation is the gold standard for the treatment of spinal diseases. However, many studies have reported the issue of loosening pedicle ...

an-anesthesia-surgery-harvest-method-for-evaluation-transpedicular

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Unit 1

An Anesthesia, Surgery, and Harvest Method for the Evaluation of Transpedicular Screws Using an In Vivo Porcine Lumbar Spine Model

Local Anesthetics

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7.6K ビュー.  Chonbuk National University School of Medicine, Chonbuk National University Hospital. ここでは、 インビボでのブタの腰椎モデルを用いて経椎骨のネジを評価する方法を紹介する。 

ビデオ: An Anesthesia, Surgery, and Harvest Method for the Evaluation of Transpedicular Screws Using an In Vivo Porcine Lumbar Spine Model

In vivo Imaging of the Mouse Spinal Cord Using Two-photon Microscopy

In vivo imaging using two-photon microscopy 1 in mice that have been genetically engineered to express fluorescent proteins in specific cell types 2-3 has significantly broadened our knowledge of physiological and pathological processes in numerous tissues in vivo 4-7. In studies of the central nervous system (CNS), there has been a broad application of in vivo imaging in the brain, which has produced a plethora of novel and often unexpected findings about the behavior of cells such as neurons, astrocytes, microglia, under physiological or pathological conditions 8-17. However, mostly technical complications have limited the implementation of in vivo imaging in studies of the living mouse spinal cord. In particular, the anatomical proximity of the spinal cord to the lungs and heart generates significant movement artifact that makes imaging the living spinal cord a challenging task.
We developed a novel method that overcomes the inherent limitations of spinal cord imaging by stabilizing the spinal column, reducing respiratory-induced movements and thereby facilitating the use of two-photon microscopy to image the mouse spinal cord in vivo. This is achieved by combining a customized spinal stabilization device with a method of deep anesthesia, resulting in a significant reduction of respiratory-induced movements. This video protocol shows how to expose a small area of the living spinal cord that can be maintained under stable physiological conditions over extended periods of time by keeping tissue injury and bleeding to a minimum. Representative raw images acquired in vivo detail in high resolution the close relationship between microglia and the vasculature. A timelapse sequence shows the dynamic behavior of microglial processes in the living mouse spinal cord. Moreover, a continuous scan of the same z-frame demonstrates the outstanding stability that this method can achieve to generate stacks of images and/or timelapse movies that do not require image alignment post-acquisition. Finally, we show how this method can be used to revisit and reimage the same area of the spinal cord at later timepoints, allowing for longitudinal studies of ongoing physiological or pathological processes in vivo.

In vivo Imaging of the Mouse Spinal Cord Using Two-photon Microscopy

A minimally invasive protocol to stabilize the mouse spinal column and perform repetitive in vivo spinal cord imaging using two-photon microscopy is described. This method combines a spinal stabilization device and an anesthetic regimen to minimize respiratory-induced movements and produce raw imaging data that require no alignment or other post-processing.

二光子顕微鏡を用いたマウス脊髄の in vivoイメージングで

In vivo imaging using two-photon microscopy 1 in mice that have been genetically engineered to express fluorescent proteins in specific cell types 2-3 has ...

神経科学

A Procedure for Implanting a Spinal Chamber for Longitudinal In Vivo Imaging of the Mouse Spinal Cord

Studies in the mammalian neocortex have enabled unprecedented resolution of cortical structure, activity, and response to neurodegenerative insults by repeated, time-lapse in vivo imaging in live rodents. These studies were made possible by straightforward surgical procedures, which enabled optical access for a prolonged period of time without repeat surgical procedures. In contrast, analogous studies of the spinal cord have been previously limited to only a few imaging sessions, each of which required an invasive surgery. As previously described, we have developed a spinal chamber that enables continuous optical access for upwards of 8 weeks, preserves mechanical stability of the spinal column, is easily stabilized externally during imaging, and requires only a single surgery. Here, the design of the spinal chamber with its associated surgical implements is reviewed and the surgical procedure is demonstrated in detail. Briefly, this video will demonstrate the preparation of the surgical area and mouse for surgery, exposure of the spinal vertebra and appropriate tissue debridement, the delivery of the implant and vertebral clamping, the completion of the chamber, the removal of the delivery system, sealing of the skin, and finally, post-operative care. The procedure for chronic in vivo imaging using nonlinear microscopy will also be demonstrated. Finally, outcomes, limitations, typical variability, and a guide for troubleshooting are discussed.

A Procedure for Implanting a Spinal Chamber for Longitudinal In Vivo Imaging of the Mouse Spinal Cord

In this video, we describe a procedure for implanting a chronic optical imaging chamber over the dorsal spinal cord of a live mouse. The chamber, surgical procedure, and chronic imaging are reviewed and demonstrated.

縦断のために脊髄商工を移植するための手順<em&gt;インビボ</emマウス脊髄の&gt;イメージング

Studies in the mammalian neocortex have enabled unprecedented resolution of cortical structure, activity, and response to neurodegenerative insults by ...

Laminotomy for Lumbar Dorsal Root Ganglion Access and Injection in Swine

Dorsal root ganglia (DRG) are anatomically well defined structures that contain all primary sensory neurons below the head. This fact makes DRG attractive targets for injection of novel therapeutics aimed at treating chronic pain. In small animal models, laminectomy has been used to facilitate DRG injection because it involves surgical removal of the vertebral bone surrounding each DRG. We demonstrate a technique for intraganglionic injection of lumbar DRG in a large animal species, namely, swine. Laminotomy is performed to allow direct access to DRG using standard neurosurgical techniques, instruments, and materials. Compared with more extensive bone removal via laminectomy, we implement laminotomy to conserve spinal anatomy while achieving sufficient DRG access. Intraoperative progress of DRG injection is monitored using a non-toxic dye. Following euthanasia on post-operative day 21, the success of injection is determined by histology for intraganglionic distribution of 4&#39;,6-diamidino-2-phenylindole (DAPI). We inject a biologically inactive solution to demonstrate the protocol. This method could be applied in future preclinical studies to target therapeutic solutions to DRG. Our methodology should facilitate testing the translatability of intraganglionic small animal paradigms in a large animal species. Additionally, this protocol may serve as a key resource for those planning preclinical studies of DRG injection in swine.

We describe a method for laminotomy in swine that provides access to lumbar dorsal root ganglia (DRG) for intraganglionic injection. Injection progress is monitored intraoperatively and histologically confirmed up to 21 days after surgery. This protocol could be used for future preclinical studies involving DRG injection.

腰部後根神経節のアクセスと豚注入の椎

Dorsal root ganglia (DRG) are anatomically well defined structures that contain all primary sensory neurons below the head. This fact makes DRG attractive ...

Laminectomy and Spinal Cord Window Implantation in the Mouse

This protocol describes a method for spinal cord laminectomy and glass window implantation for in vivo imaging of the mouse spinal cord. An integrated digital vaporizer is utilized to achieve a stable plane of anesthesia at a low-flow rate of isoflurane. A single vertebral spine is removed, and a commercially available cover-glass is overlaid on a thin agarose bed. A 3D-printed plastic backplate is then affixed to the adjacent vertebral spines using tissue adhesive and dental cement. A stabilization platform is used to reduce motion artifact from respiration and heartbeat. This rapid and clamp-free method is well-suited for acute multi-photon fluorescence microscopy. Representative data are included for an application of this technique to two-photon microscopy of the spinal cord vasculature in transgenic mice expressing eGFP:Claudin-5 &#8212; a tight junction protein.

This protocol describes implantation of a glass window onto the spinal cord of a mouse to facilitate visualization by intravital microscopy.

マウスにおける線込みおよび脊髄窓移植

This protocol describes a method for spinal cord laminectomy and glass window implantation for in vivo imaging of the mouse spinal cord. An integrated ...

In Vivo Mouse Model of Spinal Implant Infection

Spine implant infections portend poor outcomes as diagnosis is challenging and surgical eradication is at odds with mechanical spinal stability. The purpose of this method is to describe a novel mouse model of spinal implant infection (SII) that was created to provide an inexpensive, rapid, and accurate in vivo tool to test potential therapeutics and treatment strategies for spinal implant infections.
In this method, we present a model of posterior-approach spinal surgery in which a stainless-steel k-wire is transfixed into the L4 spinous process of 12-week old C57BL/6J wild-type mice and inoculated with 1 x 103 CFU of a bioluminescent strain of Staphylococcus aureus Xen36 bacteria. Mice are then longitudinally imaged for bioluminescence in vivo on post-operative days 0, 1, 3, 5, 7, 10, 14, 18, 21, 25, 28, and 35. Bioluminescence imaging (BLI) signals from a standardized field of view are quantified to measure in vivo bacterial burden.
To quantify bacteria adhering to implants and peri-implant tissue, mice are euthanized and the implant and surrounding soft tissue are harvested. Bacteria are detached from the implant by sonication, cultured overnight and then colony forming units (CFUs) are counted. The results acquired from this method include longitudinal bacterial counts as measured by in vivo S. aureus bioluminescence (mean maximum flux) and CFU counts following euthanasia.
While prior animal models of instrumented spine infection have involved invasive, ex vivo tissue analysis, the mouse model of SII presented in this paper leverages noninvasive, real time in vivo optical imaging of bioluminescent bacteria to replace static tissue study. Applications of the model are broad and may include utilizing alternative bioluminescent bacterial strains, incorporating other types of genetically engineered mice to contemporaneously study host immune response, and evaluating current or investigating new diagnostic and therapeutic modalities such as antibiotics or implant coatings.

The protocol describes a novel in vivo mouse model of spinal implant infection where a stainless-steel k-wire implant is infected with bioluminescent Staphylococcus aureus Xen36. Bacterial burden is monitored longitudinally with bioluminescent imaging and confirmed with colony forming unit counts after euthanasia.

脊椎インプラント感染のin vivoマウスモデル

Spine implant infections portend poor outcomes as diagnosis is challenging and surgical eradication is at odds with mechanical spinal stability. The ...

医学

Real-Time Assessment of Spinal Cord Microperfusion in a Porcine Model of Ischemia/Reperfusion

Spinal cord injury is a devastating complication of aortic repair. Despite developments for the prevention and treatment of spinal cord injury, its incidence is still considerably high and therefore, influences patient outcome. Microcirculation plays a key role in tissue perfusion and oxygen supply and is often dissociated from macrohemodynamics. Thus, direct evaluation of spinal cord microcirculation is essential for the development of microcirculation-targeted therapies and the evaluation of existing approaches in regard to spinal cord microcirculation. However, most of the methods do not provide real-time assessment of spinal cord microcirculation. The aim of this study is to describe a standardized protocol for real-time spinal cord microcirculatory evaluation using laser-Doppler needle probes directly inserted in the spinal cord. We used a porcine model of ischemia/reperfusion to induce deterioration of the spinal cord microcirculation. In addition, a fluorescent microsphere injection technique was used. Initially, animals were anesthetized and mechanically ventilated. Thereafter, laser-Doppler needle probe insertion was performed, followed by the placement of cerebrospinal fluid drainage. A median sternotomy was performed for exposure of the descending aorta to perform aortic cross-clamping. Ischemia/reperfusion was induced by supra-celiac aortic cross-clamping for a total of 48 min, followed by reperfusion and hemodynamic stabilization. Laser-Doppler Flux was performed in parallel with macrohemodynamic evaluation. In addition, automated cerebrospinal fluid drainage was used to maintain a stable cerebrospinal pressure. After completion of the protocol, animals were sacrificed, and the spinal cord was harvested for histopathological and microsphere analysis. The protocol reveals the feasibility of spinal cord microperfusion measurements using laser-Doppler probes and shows a marked decrease during ischemia as well as recovery after reperfusion. Results showed comparable behavior to fluorescent microsphere evaluation. In conclusion, this new protocol might provide a useful large animal model for future studies using real-time spinal cord microperfusion assessment in ischemia/reperfusion conditions.

Spinal cord microcirculation plays a pivotal role in spinal cord injury. Most methods do not allow real-time assessment of spinal cord microcirculation, which is essential for the development of microcirculation-targeted therapies. Here, we propose a protocol using Laser-Doppler-Flow Needle probes in a large animal model of ischemia/reperfusion.

イシュ血症/再灌流の豚モデルにおける脊髄微小灌流のリアルタイム評価

Spinal cord injury is a devastating complication of aortic repair. Despite developments for the prevention and treatment of spinal cord injury, its ...

Surgical Technique for Lumbar Spinal Catheter Insertion in Pigs Enabling Continuous Access to the Thecal Sac in a Terminal Setup

Pigs are increasingly used as a large animal model for pharmacologic CNS research due to the anatomical and physiological similarities between the porcine and human central nervous systems (CNS). However, accessing the cerebrospinal fluid (CSF) in larger pig breeds by conventional lumbar puncture techniques can be challenging due to an oblique orientation of the spinal spinous processes and a limited interlaminar space. Accordingly, an open surgical procedure for inserting a lumbar spinal catheter for continuous CSF sampling at the L4/L5 level in pigs is thoroughly described in this work. After positioning the pig and identifying the anatomical landmarks, a dorsal midline surgical incision is made to expose the spinous processes. By advancing the introducer needle, the spinal catheter is inserted inside the thecal sac of the spinal canal while leaving the bone structures of the spine intact. This method allows continuous infusion into or sampling from the porcine thecal sac with minimal bleeding or CSF leakage. The procedure is simple, time-efficient, and reproducible across different experimental setups, offering significant potential for various pre-clinical studies, including pharmacokinetic research, surgical training, and spinal cord injury models.

We present a technique for inserting a lumbar spinal catheter at the L4-L5 level in a 3-month-old Danish Landrace pig as part of a terminal research protocol, enabling continuous infusion or CSF sampling from the thecal sac.

ピグの腰椎カテーテル挿入のための外科的技術により、ターミナルセットアップで髄腔嚢への継続的なアクセスが可能

Pigs are increasingly used as a large animal model for pharmacologic CNS research due to the anatomical and physiological similarities between the porcine ...

A Mouse Model of Lumbar Spine Instability

Intervertebral disc degeneration (IDD) is a common pathological change leading to low back pain. Appropriate animal models are desired for understanding the pathological processes and evaluating new drugs. Here, we introduced a surgically induced lumbar spine instability (LSI) mouse model that develops IDD starting from 1 week post operation. In detail, the mouse under anesthesia was operated by low back skin incision, L3&#8211;L5 spinous processes exposure, detachment of paraspinous muscles, resection of processes and ligaments, and skin closure. L4&#8211;L5 IVDs were chosen for the observation. The LSI model develops lumbar IDD by porosity and hypertrophy in endplates at an early stage, decrease in intervertebral disc volume, shrinkage in nucleus pulposus at an intermediate stage, and bone loss in lumbar vertebrae (L5) at a later stage. The LSI mouse model has the advantages of strong operability, no requirement of special equipment, reproducibility, inexpensive, and relatively short period of IDD development. However, LSI operation is still a trauma that causes inflammation within the first week post operation. Thus, this animal model is suitable for study of lumbar IDD.

We developed a lumbar intervertebral disc degeneration mouse model by resection of L3&#8211;L5 spinous processes along with supra- and inter-spinous ligaments and detachment of paraspinous muscles.

腰椎不安定性のマウスモデル

Intervertebral disc degeneration (IDD) is a common pathological change leading to low back pain. Appropriate animal models are desired for understanding ...

生物学

Pedicle Screw Placement Using an Augmented Reality Head-Mounted Display in a Porcine Model

This protocol helps assess the accuracy and workflow of an augmented reality (AR) hybrid navigation system using the Magic Leap head-mounted display (HMD) for minimally invasive pedicle screw placement. The cadaveric porcine specimens were placed on a surgical table and draped with sterile covers. The levels of interest were identified using fluoroscopy, and a dynamic reference frame was attached to the spinous process of a vertebra in the region of interest. Cone beam computerized tomography (CBCT) was performed, and a 3D rendering was automatically generated, which was used for the subsequent planning of the pedicle screw placements. Each surgeon was fitted with an HMD that was individually eye-calibrated and connected to the spinal navigation system.
Navigated instruments, tracked by the navigation system and displayed in 2D and 3D in the HMD, were used for 33 pedicle cannulations, each with a diameter of 4.5 mm. Postprocedural CBCT scans were assessed by an independent reviewer to measure the technical (deviation from the planned path) and clinical (Gertzbein grade) accuracy of each cannulation. The navigation time for each cannulation was measured. The technical accuracy was 1.0 mm &#177; 0.5 mm at the entry point and 0.8 mm &#177; 0.1 mm at the target. The angular deviation was 1.5&#176; &#177; 0.6&#176;, and the mean insertion time per cannulation was 141 s &#177; 71 s. The clinical accuracy was 100% according to the Gertzbein grading scale (32 grade 0; 1 grade 1). When used for minimally invasive pedicle cannulations in a porcine model, submillimeter technical accuracy and 100% clinical accuracy could be achieved with this protocol.

The augmented reality head-mounted display, Magic Leap, was used in combination with a conventional navigation system to place pedicle screws in a porcine model by adhering to a novel workflow. With a median insertion time of &lt;2.5 min, submillimeter technical accuracy and 100% clinical accuracy were achieved according to Gertzbein.

ブタモデルにおける拡張現実ヘッドマウントディスプレイを用いた椎弓根スクリュー配置

This protocol helps assess the accuracy and workflow of an augmented reality (AR) hybrid navigation system using the Magic Leap head-mounted display (HMD) ...

<meta charset="utf8"></head><body>胸腰椎破裂骨折のための矢状調整スクリューシステムの技術

Minimally Invasive Treatment for Thoracolumbar Burst Fracture Using Sagittal Alignment Screws and A Trauma Reduction Device

Thoracolumbar (TL) burst fracture is one of the most common indications for minimally invasive percutaneous pedicle screw fixation. Although the indication for surgical treatment of neurologically intact TL fractures remains under debate, studies have demonstrated that posttraumatic malalignment may lead to a deterioration in the patient&#39;s quality of life. For burst fractures with malalignment or fragments in the spinal canal, a reduction technique using ligamentotaxis is commonly used to improve long-term outcomes.
The sagittal adjusting screw (SAS) system is a monoaxial screw system with a fixed head and concave sliding saddle that allows lordotic sliding of the rod in the sagittal plane after screw insertion. SAS also has a percutaneous option and has been used for TL spine fractures. Notably, the SAS only allows motion on the sagittal plane, allowing both secure fixation and angular reduction. The SAS has certain advantages over the conventional Schanz screw system or normal mono-/multiaxial pedicle screws for TL spine fracture treatment. In addition, specialized trauma reduction devices are available for the SAS system. In this video protocol, we discuss the indication for the SAS system in TL burst fracture and describe a technique of TL burst fracture reduction and fixation using the SAS system. Additionally, we describe our recent case series with radiological evaluation, including regional kyphotic angle and percent loss of anterior vertebral body height, to evaluate the newly introduced trauma reduction device.

<meta charset="utf8"></head><body>矢状アライメントスクリューと外傷軽減装置を用いた胸腰椎破裂骨折の低侵襲治療

The sagittal adjusting screw (SAS) system has been widely used for thoracolumbar (TL) spinal trauma. Percutaneous options and specialized trauma reduction devices are also available for the SAS system. We describe a technique for reducing TL burst fractures using an SAS system and a newly introduced trauma reduction device.

麻酔、手術および収縮法を用いた経肛門用ねじの評価

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