1）準備<ol><li>開始するには、、NeuroStarシステムを起動してログインし、自動化されたコイルのテストを実行します。 </li><li>次に、利用可能なリスト（NeuroStarのシステムを使用して治療を受けたすべての患者が中央のコンピュータにログインしている）から適切な患者のファイルにアクセスする</li><li>最後に、椅子の設定が制限されていないことを確認し、それが簡単に患者が入ると座ってできるようにするために、椅子のアームを持ち上げます。 </li></ol> 2）患者の座席と整列<ol><li>患者が到着、座席彼/椅子で快適に彼女。 </li><li>患者と技術者のインサートの耳栓の両方を確認してください。 </li><li>粘着テープを使用し、単に患者の眉毛の上にヘッドの位置決めのストラップを取り付けます。中心点は[後]頭骨の上に置かれていることを確認してください。 </li><li>椅子を倒して、必要に応じてヘッドのサポートを調整する。 </li><li>軽く、患者の[後]頭骨上記のベルクロに上部ヘッドストラップを取り付けます。 </li><li>レーザーをアクティブにし、それが患者の[後]頭骨を二等分することを確認してください。 </li><li>それは、眼球とレベルになるまで/ Pバーを壁上の点を凝視して調整されることを求める。 </li><li>患者が完全に中心にされていることを確認し、治療の椅子に側のヘッドの位置決めのストラップを固定するマジックテープを使用してください。 </li><li>軽く運動しきい値の決定の際に頭部の安定性を確保するために側頭部のパッドを従事する。 </li><li>最後に、タッチスクリーンのインターフェースを使用してヘッド支援システムの設定を記録。 </li></ol> 3）モーターのしきい値を決定する<ol><li>モーターのしきい値を決定するために、患者の頭部にコイルが頂点にあると盛士達のタブが耳の上端より上に整列されるようにコイルを下に移動。 </li><li>それはコイルの面と揃うまで前方/ Pのバーを移動します。 </li><li>コイルの角度指標で0度マークでコイルにセンターラインを合わせますと、/ Pのバーを戻す。 </li><li>コンタクトとポジションが確立されたら、ヒッチハイカーの位置を制定する対象を求める：腕は肘で90度曲げ、こぶしは、大まかに親指拡張外側を開催。 </li><li>刺激と親指のあらゆる動きに焦点を始める。 </li><li>親指または一致した指の動きが見られるまで徐々に0.68から1.10にMTのレベルを増加させる。 </li><li>動きが確立されると、モーターのホットスポットの検索を開始。必要に応じてMTのレベルを調整し、パターンのようなグリッドの中をコイルが移動する。あなたが10の連続パルスのうち正確に5親指のけいれんを誘発する場所と電力レベルの設定が見つかるまでこの検索を続けます。 </li><li>モーターのホットスポットが決定されるとそれは、再び、コイルの面と揃うまで、前方に/ Pバーを移動。 </li><li> / Pの位置、コイルの角度、SOAの位置、およびパワーレベルを記録するためにタッチスクリーンインターフェイスを使用します。 </li><li>最後に、見つかったMTのボタンを押してください。我々は現在、治療のための準備が整いました。 </li></ol> 4）患者の治療<ol><li>開始するには、MTの位置に前方コイル5.5センチメートルを移動する。これは、治療の場所になります。 </li><li>コイルは、患者の頭部と接触をしていることを確認し、治療の期間はまだ残っているために患者を求める。 </li><li>治療のパラメータは、デバイスにプリロードされます。刺激は26第二沈黙期間が続く4秒間、10 Hzのパルスシーケンスと120％モーターのしきい値、で生成されます。治療は、これは3000パルスの合計である37.5分の合計のために持続する。 </li><li>確認のパルスシーケンスのボタンを押して[スタート]ボタンを押してください。 </li><li>治療中、患者の部屋に残り、定期的にコイルが患者に接触することを確認して下さい。 </li><li>パルスシーケンスが完了すると、コイルを上に移動し、離れて患者の頭部からと側頭部のサポートパッドを離してください。 </li><li>椅子の後部を持ち上げ、椅子のアームを持ち上げます。 </li><li>彼/彼女が立ち上がるように患者を支援する。 </li><li>耳栓を外し、治療セッションを終了するには、ログアウトボタンをタッチします。 </li></ol> 5）代表者の結果 TMSは、通常4-6週間の期間のために毎日適用されます。最近のデータは、一部の患者は4-6ヶ月後に症状の再発を起こしやすいことを示唆している。8は、この問題が発生した場合は、実務家は効果を延長するためのメンテナンスセッションを検討する必要があります。この治療は、治療を受けた患者の約60％で有効であった。 <img src="/files/ftp_upload/2345/2345fig1.jpg" alt="表5" />

<ol>
	<li>Pascual-Leone, A., Davey, M., Wassermann, E. M., Rothwell, J., Puri, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Handbook of Transcranial Magnetic Stimulation. , (2002).</li><li>Walsh, V., Pascual-Leone, 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> Transcranial Magnetic Stimulation: A Neurochronometrics of Mind. , (2005).</li><li>Horvath, J. C., Perez, J., Forrow, L., Fregni, F., Pascual-Leone, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transcranial+Magnetic+Stimulation:+An+Historical+Evaluation+and+Future+Prognosis+of+Therapeutically+Relevant+Ethical+Concerns.">Transcranial Magnetic Stimulation: An Historical Evaluation and Future Prognosis of Therapeutically Relevant Ethical Concerns.</a> , .</li><li>O'Reardon, J. P., Solvason, H. B., Janicak, P. G., Sampson, S., Isenberg, K. E., Nahas, Z., McDonald, W. M., Avery, D., Fitzgerald, P. B., Loo, C., Demitrack, M. A., George, M. S., Sackheim, H. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Efficacy+and+safety+of+transcranial+magnetic+stimulation+in+the+acute+treatment+of+major+depression:+a+multisite+randomized+control+trial.">Efficacy and safety of transcranial magnetic stimulation in the acute treatment of major depression: a multisite randomized control trial.</a> Biological Psychiatry. 62, 1208-1216 (2007).</li><li>George, M. S., Lisanby, S. H., Avery, D., McDonald, W. M., Durkalski, V., Pavlicova, M., Anderson, B., Nahas, Z., Bulow, P., Zarkowski, P., Holtzheimer, P. D., Schwartz, T., Sackheim, H. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Daily+prefrontal+transcranial+magnetic+stimulation+therapy+for+major+depressive+disorder:+a+sham-controlled+randomized+study.">Daily prefrontal transcranial magnetic stimulation therapy for major depressive disorder: a sham-controlled randomized study.</a> Arch Gen Psychiatry. 67, 507-516 (2010).</li><li>Lisanbym, S. H., Husain, M. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Daily+Left+Prefrontal+Repetitive+Transcranial+Magnetic+Stimulation+(rTMS)+in+the+Acute+Treatment+of+Major+Depression:+Clinical+Predictors+of+Outcome+in+a+Multisite,+Randomized+Controlled+Clinical+Trial.">Daily Left Prefrontal Repetitive Transcranial Magnetic Stimulation (rTMS) in the Acute Treatment of Major Depression: Clinical Predictors of Outcome in a Multisite, Randomized Controlled Clinical Trial.</a> Neuropsychopharmacology. 34, 522-534 (2008).</li><li>Thase, M., Demitrack, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evaluating+clinical+significance+of+treatment+outcomes+in+studies+of+resistant+major+depression.">Evaluating clinical significance of treatment outcomes in studies of resistant major depression.</a> Biological Psychiatry. 63, 138-138 (2008).</li><li>Demirtas-Tatlidede, A., Mechanic-Hamilton, D., Press, D. Z., Pearlman, C., Stern, W. M., Thall, M., Pascual-Leone, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+open-label,+prospective+study+of+repetitive+transcranial+magnetic+stimulation+(rTMS)+in+the+long-term+treatment+of+refractory+depression:+reproducibility+and+duration+of+the+antidepressant+effect+in+medication-free+patients.">An open-label, prospective study of repetitive transcranial magnetic stimulation (rTMS) in the long-term treatment of refractory depression: reproducibility and duration of the antidepressant effect in medication-free patients.</a> Journal of Clinical Psychiatry. 69, 930-934 (2008).</li></ol>

この記事のビデオ制作は、これらの研究で使用される楽器を作り出すNeuronetics後援した。

うつ病のための潜在的な治療的介入としてTMSを検討する際に、それがFDA承認されたプロトコルが高度に指定されて認識することが重要です。具体的には、FDAの承認は、10 Hzの閾値上刺激に制限されているものから満足のいく改善を達成するために失敗した患者に基づいNeuroStarのデバイスを使用して4-6週間は毎日適用されるが、現在の抑うつの間に1つしか、十分な抗うつ薬の臨床試験エピソード。この時点で、任意の代替刺激パターン、別の時間の経過、代替デバイス、または別の患者集団の使用率は適応外とみなされます。 結果は、TMS治療5,6,7の有効性をサポートするために生成され続けるように、新しいデータは、治療中止後4-6ヶ月に発生する症状の再発の可能性を明らかにしている。この出版物のように、承認されたメンテナンスプロトコルはありません。個々の専門家によってそのため、追加の治療および/または効果の維持管理に関する問題を考慮する必要がありますし、厳重に監視。

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<td>Neuronetics Neurostar TMS Device</td>
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<td>SenStar Treatment Link</td>
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<td>Ear Plugs</td>
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<td>Head Pad Sanitary Sheets</td>
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<td>Alignment Head Strap</td>
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<td>Motor Threshold Arm Position Pad</td>
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the neurostar tms device: conducting the fda approved protocol for treatment of depression

Neuronetics NeuroStar経頭蓋磁気刺激（TMS）システムが短期間に生成するクラスII医療機器であり、磁場をパルス。これらの急速に交互のフィールドは様々な生理的および機能的な脳の変化に関連付けられている大脳皮質のローカライズされた、対象地域内で電気電流を誘発する。1,2,3 2007年に、O&#39;Reardon ら 、これまででへの応答に失敗していた大うつ病の患者301人、業界主催の、マルチサイト、無作為、偽刺激対照臨床試験の結果を発表し、NeuroStar装置を利用している少なくとも一つの適切な抗うつ薬治療の臨床試験は、左背外側前頭前野（DLPFC）上のアクティブまたは偽TMSのいずれかを受けた。調査の時点で薬剤フリーであった患者は、4〜6週間にわたって週に5回TMSを受けた。4 結果は、患者の亜集団は、（薬に比較的少ない耐性であった人々、つ以上の良好な薬理学的試験ではない失敗したこと）モンゴメリー- Asberg抑うつ尺度（MADRS）、ハミルトンうつ病評価に関する統計的に有意な改善を示したことが明らかにスケール（HAMD）、および他の様々なアウトカム指標。 2008年10月には、これらおよび他の同様の結果が5,6,7でサポートされている、Neuroneticsは（TMSの治療装置を使用して薬物治療が難治性うつ病の特定の形態の臨床治療のための最初で唯一の食品医薬品局（FDA）の承認を得FDAの承認K061053）。 本稿では、（どちらか、または外来での唯一の処方箋のもと、ライセンスを取得した医療従事者によって投与すること）指定されたFDA承認NeuroStarうつ病の治療のプロトコールを検討します。

Watch this Scientific Journal Video about The NeuroStar TMS Device: Conducting the FDA Approved Protocol for Treatment of Depression at JoVE.com

The NeuroStar TMS Device: Conducting the FDA Approved Protocol for Treatment of Depression

この記事では、我々はNeuronetics NeuroStar TMSデバイスを使用してFDA承認されたうつ病の治療プロトコルに関連する方法論と考慮事項を検討する。

NeuroStar TMSデバイス：うつ病の治療薬としてFDA承認されたプロトコルを実施

The Neuronetics NeuroStar Transcranial Magnetic Stimulation (TMS) System is a class II medical device that produces brief duration, pulsed magnetic ...

the-neurostar-tms-device-conducting-fda-approved-protocol-for

Nanyang Technological University

Research

JoVE Journal

Neuroscience

39.5K ビュー.  Beth Israel Deaconess Medical Center. この記事では、我々はNeuronetics NeuroStar TMSデバイスを使用してFDA承認されたうつ病の治療プロトコルに関連する方法論と考慮事項を検討する。

ビデオ: The NeuroStar TMS Device: Conducting the FDA Approved Protocol for Treatment of Depression

TMS: Using the Theta-Burst Protocol to Explore Mechanism of Plasticity in Individuals with Fragile X Syndrome and Autism

Fragile X Syndrome (FXS), also known as Martin-Bell Syndrome, is a genetic abnormality found on the X chromosome.1,2 Individuals suffering from FXS display abnormalities in the expression of FMR1 - a protein required for typical, healthy neural development.3 Recent data has suggested that the loss of this protein can cause the cortex to be hyperexcitable thereby affecting overall patterns of neural plasticity.4,5
In addition, Fragile X shows a strong comorbidity with autism: in fact, 30% of children with FXS are diagnosed with autism, and 2 - 5% of autistic children suffer from FXS.6
Transcranial Magnetic Stimulation (a non-invasive neurostimulatory and neuromodulatory technique that can transiently or lastingly modulate cortical excitability via the application of localized magnetic field pulses 7,8) represents a unique method of exploring plasticity and the manifestations of FXS within affected individuals. More specifically, Theta-Burst Stimulation (TBS), a specific stimulatory protocol shown to modulate cortical plasticity for a duration up to 30 minutes after stimulation cessation in healthy populations, has already proven an efficacious tool in the exploration of abnormal plasticity.9,10 
Recent studies have shown the effects of TBS last considerably longer in individuals on the autistic spectrum - up to 90 minutes.11 This extended effect-duration suggests an underlying abnormality in the brain's natural plasticity state in autistic individuals - similar to the hyperexcitability induced by Fragile X Syndrome.
In this experiment, utilizing single-pulse motor-evoked potentials (MEPs) as our benchmark, we will explore the effects of both intermittent and continuous TBS on cortical plasticity in individuals suffering from FXS and individuals on the Autistic Spectrum.

In this article, we examine the effects of Theta-Burst TMS stimulation on cortical plasticity in individuals suffering from Fragile X syndrome and individuals on the autistic spectrum.

TMS：脆弱X症候群と自閉症の可塑性のMechasnismを探検するシータバーストプロトコルを使用する

Fragile X Syndrome (FXS), also known as Martin-Bell Syndrome, is a genetic abnormality found on the X chromosome.1,2 Individuals suffering from FXS ...

神経科学

Modeling Neural Immune Signaling of Episodic and Chronic Migraine Using Spreading Depression In Vitro

Migraine and its transformation to chronic migraine are healthcare burdens in need of improved treatment options. We seek to define how neural immune signaling modulates the susceptibility to migraine, modeled in vitro using spreading depression (SD), as a means to develop novel therapeutic targets for episodic and chronic migraine. SD is the likely cause of migraine aura and migraine pain. It is a paroxysmal loss of neuronal function triggered by initially increased neuronal activity, which slowly propagates within susceptible brain regions. Normal brain function is exquisitely sensitive to, and relies on, coincident low-level immune signaling. Thus, neural immune signaling likely affects electrical activity of SD, and therefore migraine. Pain perception studies of SD in whole animals are fraught with difficulties, but whole animals are well suited to examine systems biology aspects of migraine since SD activates trigeminal nociceptive pathways. However, whole animal studies alone cannot be used to decipher the cellular and neural circuit mechanisms of SD. Instead, in vitro preparations where environmental conditions can be controlled are necessary. Here, it is important to recognize limitations of acute slices and distinct advantages of hippocampal slice cultures. Acute brain slices cannot reveal subtle changes in immune signaling since preparing the slices alone triggers: pro-inflammatory changes that last days, epileptiform behavior due to high levels of oxygen tension needed to vitalize the slices, and irreversible cell injury at anoxic slice centers. 
In contrast, we examine immune signaling in mature hippocampal slice cultures since the cultures closely parallel their in vivo counterpart with mature trisynaptic function; show quiescent astrocytes, microglia, and cytokine levels; and SD is easily induced in an unanesthetized preparation. Furthermore, the slices are long-lived and SD can be induced on consecutive days without injury, making this preparation the sole means to-date capable of modeling the neuroimmune consequences of chronic SD, and thus perhaps chronic migraine. We use electrophysiological techniques and non-invasive imaging to measure neuronal cell and circuit functions coincident with SD. Neural immune gene expression variables are measured with qPCR screening, qPCR arrays, and, importantly, use of cDNA preamplification for detection of ultra-low level targets such as interferon-gamma using whole, regional, or specific cell enhanced (via laser dissection microscopy) sampling. Cytokine cascade signaling is further assessed with multiplexed phosphoprotein related targets with gene expression and phosphoprotein changes confirmed via cell-specific immunostaining. Pharmacological and siRNA strategies are used to mimic and modulate SD immune signaling.

Modeling Neural Immune Signaling of Episodic and Chronic Migraine Using Spreading Depression In Vitro

Migraine and its transformation to chronic migraine are immense healthcare burdens in need of improved treatment options. We seek to define how neural immune signaling modulates the susceptibility to migraine, modeled in vitro using spreading depression in hippocampal slice cultures, as a means to develop novel therapeutic targets.

拡散うつ病を使用してエピソードと慢性片頭痛の神経免疫シグナルのモデリングインビトロ</em

Migraine and its transformation to chronic migraine are healthcare burdens in need of improved treatment options.  We seek to define how neural immune ...

A Comprehensive Protocol for Manual Segmentation of the Medial Temporal Lobe Structures

The present paper describes a comprehensive protocol for manual tracing of the set of brain regions comprising the medial temporal lobe (MTL): amygdala, hippocampus, and the associated parahippocampal regions (perirhinal, entorhinal,&#160;and parahippocampal proper). Unlike most other tracing protocols available, typically focusing on certain MTL areas (e.g., amygdala and/or hippocampus), the integrative perspective adopted by the present tracing guidelines allows for clear localization of all MTL subregions. By integrating information from a variety of sources, including extant tracing protocols separately targeting various MTL structures, histological reports, and brain atlases, and with the complement of illustrative visual materials, the present protocol provides an accurate, intuitive, and convenient guide for understanding the MTL anatomy. The need for such tracing guidelines is also emphasized by illustrating possible differences between automatic and manual segmentation protocols. This knowledge can be applied toward research involving not only structural MRI investigations but also structural-functional colocalization and fMRI signal extraction from anatomically defined ROIs, in healthy and clinical groups alike.

The present work provides a comprehensive set of guidelines for manually tracing the medial temporal lobe (MTL) structures. This protocol can be applied to research involving structural and/or combined structural-functional magnetic resonance imaging (MRI) investigations of the MTL, in both healthy and clinical groups.

内側側頭葉構造の手動分割のための総合的なプロトコル

The present paper describes a comprehensive protocol for manual tracing of the set of brain regions comprising the medial temporal lobe (MTL): amygdala, ...

Telomerase Activity in the Various Regions of Mouse Brain: Non-Radioactive Telomerase Repeat Amplification Protocol (TRAP) Assay

Telomerase, a ribonucleoprotein, is responsible for maintaining the telomere length and therefore promoting genomic integrity, proliferation, and lifespan. In addition, telomerase protects the mitochondria from oxidative stress and confers resistance to apoptosis, suggesting its possible importance for the surviving of non-mitotic, highly active cells such as neurons. We previously demonstrated the ability of novel telomerase activators to increase telomerase activity and expression in the various mouse brain regions and to protect motor neurons cells from oxidative stress. These results strengthen the notion that telomerase is involved in the protection of neurons from various lesions. To underline the role of telomerase in the brain, we here compare the activity of telomerase in male and female mouse brain and its dependence on age. TRAP assay is a standard method for detecting telomerase activity in various tissues or cell lines. Here we demonstrate the analysis of telomerase activity in three regions of the mouse brain by non-denaturing protein extraction using CHAPS lysis buffer followed by modification of the standard TRAP assay.
In this 2-step assay, endogenous telomerase elongates a specific telomerase substrate (TS primer) by adding TTAGGG 6 bp repeats (telomerase reaction). The telomerase reaction products are amplified by PCR reaction creating a DNA ladder of 6 bp increments. The analysis of the DNA ladder is made by 4.5% high resolution agarose gel electrophoresis followed by staining with highly sensitive nucleic acid stain.
Compared to the traditional TRAP assay that utilize 32P labeled radioactive dCTP&#39;s for DNA detection and polyacrylamide gel electrophoresis for resolving the DNA ladder, this protocol offers a non-toxic time saving TRAP assay for evaluating telomerase activity in the mouse brain, demonstrating the ability to detect differences in telomerase activity in the various female and male mouse brain region.

Telomerase Activity in the Various Regions of Mouse Brain: Non-Radioactive Telomerase Repeat Amplification Protocol TRAP Assay

Telomerase is expressed in the neonatal brain and also in distinct regions of the adult brain. We present a non-toxic time saving TRAP assay for the analysis of telomerase activity in various regions of the mouse brain and detection of differences in telomerase activity between male and female mouse brains.

マウス脳のさまざまな領域におけるテロメラーゼ活性：非放射性テロメラーゼリピート増幅プロトコル（TRAP）アッセイ

Telomerase, a ribonucleoprotein, is responsible for maintaining the telomere length and therefore promoting genomic integrity, proliferation, and ...

Preparation of Acute Brain Slices Using an Optimized N-Methyl-D-glucamine Protective Recovery Method

This protocol is a practical guide to the N-methyl-D-glucamine (NMDG) protective recovery method of brain slice preparation. Numerous recent studies have validated the utility of this method for enhancing neuronal preservation and overall brain slice viability. The implementation of this technique by early adopters has facilitated detailed investigations into brain function using diverse experimental applications and spanning a wide range of animal ages, brain regions, and cell types. Steps are outlined for carrying out the protective recovery brain slice technique using an optimized NMDG artificial cerebrospinal fluid (aCSF) media formulation and enhanced procedure to reliably obtain healthy brain slices for patch clamp electrophysiology. With this updated approach, a substantial improvement is observed in the speed and reliability of gigaohm seal formation during targeted patch clamp recording experiments while maintaining excellent neuronal preservation, thereby facilitating challenging experimental applications. Representative results are provided from multi-neuron patch clamp recording experiments to assay synaptic connectivity in neocortical brain slices prepared from young adult transgenic mice and mature adult human neurosurgical specimens. Furthermore, the optimized NMDG protective recovery method of brain slicing is compatible with both juvenile and adult animals, thus resolving a limitation of the original methodology. In summary, a single media formulation and brain slicing procedure can be implemented across various species and ages to achieve excellent viability and tissue preservation.

Preparation of Acute Brain Slices Using an Optimized N-Methyl-D-glucamine Protective Recovery Method

This protocol demonstrates the implementation of an optimized N-methyl-D-glucamine (NMDG) protective recovery method of brain slice preparation. A single media formulation is used to reliably obtain healthy brain slices from animals of any age and for diverse experimental applications.

最適化されたNを使用して急性脳スライスの準備-メチル-D-glucamine 保護回復法

This protocol is a practical guide to the N-methyl-D-glucamine (NMDG) protective recovery method of brain slice preparation. Numerous recent studies have ...

Receptor Autoradiography Protocol for the Localized Visualization of Angiotensin II Receptors

This protocol describes receptor binding patterns for Angiotensin II (Ang II) in the rat brain using a radioligand specific for Ang II receptors to perform receptor autoradiographic mapping.
Tissue specimens are harvested and stored at -80 &#176;C. A cryostat is used to coronally section the tissue (brain) and thaw-mount the sections onto charged slides. The slide-mounted tissue sections are incubated in 125I-SI-Ang II to radiolabel Ang II receptors. Adjacent slides are separated into two sets: &#39;non-specific binding&#39; (NSP) in the presence of a receptor saturating concentration of non-radiolabeled Ang II, or an AT1 Ang II receptor subtype (AT1R) selective Ang II receptor antagonist, and &#39;total binding&#39; with no AT1R antagonist. A saturating concentration of AT2 Ang II receptor subtype (AT2R) antagonist (PD123319, 10 &#181;M) is also present in the incubation buffer to limit 125I-SI-Ang II binding to the AT1R subtype. During a 30 min pre-incubation at ~22 &#176;C, NSP slides are exposed to 10 &#181;M PD123319 and losartan, while &#39;total binding&#39; slides are exposed to 10 &#181;M PD123319. Slides are then incubated with 125I-SI-Ang II in the presence of PD123319 for &#39;total binding&#39;, and PD123319 and losartan for NSP in assay buffer, followed by several &#39;washes&#39; in buffer, and water to remove salt and non-specifically bound radioligand. The slides are dried using blow-dryers, then exposed to autoradiography film using a specialized film and cassette. The film is developed and the images are scanned into a computer for visual and quantitative densitometry using a proprietary imaging system and a spreadsheet. An additional set of slides are thionin-stained for histological comparisons.
The advantage of using receptor autoradiography is the ability to visualize Ang II receptors in situ, within a section of a tissue specimen, and anatomically identify the region of the tissue by comparing it to an adjacent histological reference section.

Here we present a protocol to describe the localization of angiotensin II Type 1 receptors in the rat brain by quantitative, densitometric, in vitro receptor autoradiography using an iodine-125 labeled analog of angiotensin II.

アンジオテンシンII受容体のローカライズされた可視化のための受容体オートラジオグラフィープロトコル

This protocol describes receptor binding patterns for Angiotensin II (Ang II) in the rat brain using a radioligand specific for Ang II receptors to ...

Protocol for Isolating the Mouse Circle of Willis

The cerebral arterial circle (circulus arteriosus cerebri) or circle of Willis (CoW) is a circulatory anastomosis surrounding the optic chiasma and hypothalamus that supplies blood to the brain and surrounding structures. It has been implicated in several cerebrovascular disorders, including cerebral amyloid angiopathy (CAA)-associated vasculopathies, intracranial atherosclerosis and intracranial aneurysms. Studies of the molecular mechanisms underlying these diseases for the identification of novel drug targets for their prevention require animal models. Some of these models may be transgenic, whereas others will involve isolation of the cerebro-vasculature, including the CoW.The method described here is suitable for CoW isolation in any mouse lineage and has considerable potential for screening (expression of genes, protein production, posttranslational protein modifications, secretome analysis, etc.) studies on the large vessels of the mouse cerebro-vasculature. It can also be used for ex vivo studies, by adapting the organ bath system developed for isolated mouse olfactory arteries.

We describe here a reproducible protocol for isolating the mouse circle of Willis.

ウィリスのマウスサークルを単離するためのプロトコル

The cerebral arterial circle (circulus arteriosus cerebri) or circle of Willis (CoW) is a circulatory anastomosis surrounding the optic chiasma and ...

A Simple One-step Dissection Protocol for Whole-mount Preparation of Adult Drosophila Brains

There is an increasing interest in using Drosophila to model human brain degenerative diseases, map neuronal circuitries in adult brains, and study the molecular and cellular basis of higher brain functions. A whole-mount preparation of adult brains with well-preserved morphology is critical for such whole brain-based studies, but can be technically challenging and time-consuming. This protocol describes an easy-to-learn, one-step dissection approach of an adult fly head in less than 10 s, while keeping the intact brain attached to the rest of the body to facilitate subsequent processing steps. The procedure helps remove most of the eye and tracheal tissues normally associated with the brain that can interfere with the later imaging step, and also places less demand on the quality of the dissecting forceps. Additionally, we describe a simple method that allows convenient flipping of the mounted brain samples on a coverslip, which is important for imaging both sides of the brains with similar signal intensity and quality. As an example of the protocol, we present an analysis of dopaminergic (DA) neurons in adult brains of WT (w1118) flies. The high efficacy of the dissection method makes it particularly useful for large-scale adult brain-based studies in Drosophila.

A Simple One-step Dissection Protocol for Whole-mount Preparation of Adult Drosophila Brains

The adult Drosophila brain is a valuable system for studying neuronal circuitry, higher brain functions, and complex disorders. An efficient method to dissect whole brain tissue from the small fly head will facilitate brain-based studies. Here we describe a simple, one-step dissection protocol of adult brains with well-preserved morphology.

大人のホールマウントの準備のためのシンプルなワンステップ解剖プロトコル<em&gt;ショウジョウバエ</em&gt;ブレインズ

There is an increasing interest in using Drosophila to model human brain degenerative diseases, map neuronal circuitries in adult brains, and study the ...

A Protocol for the Administration of Real-Time fMRI Neurofeedback Training

Neurologic disorders are characterized by abnormal cellular-, molecular-, and circuit-level functions in the brain. New methods to induce and control neuroplastic processes and correct abnormal function, or even shift functions from damaged tissue to physiologically healthy brain regions, hold the potential to dramatically improve overall health. Of the current neuroplastic interventions in development, neurofeedback training (NFT) from functional Magnetic Resonance Imaging (fMRI) has the advantages of being completely non-invasive, non-pharmacologic, and spatially localized to target brain regions, as well as having no known side effects. Furthermore, NFT techniques, initially developed using fMRI, can often be translated to exercises that can be performed outside of the scanner without the aid of medical professionals or sophisticated medical equipment. In fMRI NFT, the fMRI signal is measured from specific regions of the brain, processed, and presented to the participant in real-time. Through training, self-directed mental processing techniques, that regulate this signal and its underlying neurophysiologic correlates, are developed. FMRI NFT has been used to train volitional control over a wide range of brain regions with implications for several different cognitive, behavioral, and motor systems. Additionally, fMRI NFT has shown promise in a broad range of applications such as the treatment of neurologic disorders and the augmentation of baseline human performance. In this article, we present an fMRI NFT protocol developed at our institution for modulation of both healthy and abnormal brain function, as well as examples of using the method to target both cognitive and auditory regions of the brain.

The ability to induce and/or control neural plasticity may be critical in future treatments for neurologic disorders and the recovery from brain injury. In this paper, we present a protocol on the use of neurofeedback training with functional magnetic resonance imaging to modulate human brain function.

管理のリアルタイム fMRI ニューロフィード バック トレーニングのプロトコル

Neurologic disorders are characterized by abnormal cellular-, molecular-, and circuit-level functions in the brain. New methods to induce and control ...

Using a Microfluidics Device for Mechanical Stimulation and High Resolution Imaging of C. elegans

One central goal of mechanobiology is to understand the reciprocal effect of mechanical stress on proteins and cells. Despite its importance, the influence of mechanical stress on cellular function is still poorly understood. In part, this knowledge gap exists because few tools enable simultaneous deformation of tissue and cells, imaging of cellular activity in live animals, and efficient restriction of motility in otherwise highly mobile model organisms, such as the nematode Caenorhabditis elegans. The small size of C. elegans makes them an excellent match to microfluidics-based research devices, and solutions for immobilization have been presented using microfluidic devices. Although these devices allow for high-resolution imaging, the animal is fully encased in polydimethylsiloxane (PDMS) and glass, limiting physical access for delivery of mechanical force or electrophysiological recordings. Recently, we created a device that integrates pneumatic actuators with a trapping design that is compatible with high-resolution fluorescence microscopy. The actuation channel is separated from the worm-trapping channel by a thin PDMS diaphragm. This diaphragm is deflected into the side of a worm by applying pressure from an external source. The device can target individual mechanosensitive neurons. The activation of these neurons is imaged at high-resolution with genetically-encoded calcium indicators. This article presents the general method using C. elegans strains expressing calcium-sensitive activity indicator (GCaMP6s) in their touch receptor neurons (TRNs). The method, however, is not limited to TRNs nor to calcium sensors as a probe, but can be expanded to other mechanically-sensitive cells or sensors.

Using a Microfluidics Device for Mechanical Stimulation and High Resolution Imaging of C. elegans

New tools for mechanobiology research are needed to understand how mechanical stress activates biochemical pathways and elicits biological responses. Here, we showcase a new method for selective mechanical stimulation of immobilized animals with a microfluidic trap allowing high-resolution imaging of cellular responses.