Name: a rat model of ecohiv brain infection
Uploaded: 2021-01-21T14:30:00
Description: Watch this Scientific Journal Video about A Rat Model of EcoHIV Brain Infection at JoVE.com

この研究は、NIH助成金HD043680、MH106392、DA013137、およびNS100624によって資金提供されました。

すべての動物のプロトコルは、サウスカロライナ大学の動物ケアと使用委員会によって見直され、承認されました(連邦保証番号:D16-00028)。6人の成人男性F344/Nラットは、12/12光の下で制御された環境に収容されたペアでした:食物および水へのアドリビタムアクセスを伴う暗いサイクル。すべての動物は、実験動物のケアと使用のためのガイドで国立衛生研究所によって確立されたガイドライン...

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このプロトコルでは、ラットにEcoHIV誘導HIV感染モデルを確立した。具体的には、注射後7日のラット脳における活性HIV感染を誘導することに成功した皮質へのEcoHIVの二国間立体態注入について述べた。Futhermore, 我々はラットのEcoHIV感染が手の重要な側面を研究するための良好な生物学的システムであり得ることを実証する。8週間後- EcoHIV感染, ラットは、重要な神経認知障害を示しました, NAc?...

生物学的システムは、HIV-1関連神経認知障害(HAND)とその基礎となる神経機構2に関する我々の理解を高めている。特定の研究に最も適した生物学的システムを決定することは、しばしば関心の問題に依存する2.宿主動物モデルの範囲の限界は、HIV-1疾患の発症の研究に挑戦する。HIV-1ウイルス複製および病因を調べるため、Potash et3は、HIV表面エンベロープ糖タンパク質のコード領域を置換する活性HIV-1感染のマウスモデルを作成し、gp120を、エコトロピックMLVgp80に置き換え、マウス4におけるウイルス複製に成功した。キメラHIV(EcoHIV)マウスにおける尾静脈注射後、HIV-1血清陽性個体(例えば、感染したリンパ球およびマクロファージ)に似た多くの特徴が認められ、抗ウイルス免疫応答、および炎症3、5、6を標的とした。
マウスとラットはどちらもムリダエのメンバーですが、基本的な種の違いは、特定の実験問題に対するそれらの適合性に影響を与える可能性があります7.したがって、ラットへのEcoHIV感染モデルの拡張(一般的に薬物乱用および神経認知障害の研究で使用される)は、ニューロHIVの研究において有利であろう。例えば、その大きなサイズは、薬物自己投与手順のためのより実用的な8のための頸状カテーテル移植を行う。ラットにおける薬物自己投与技術は、HIV-19における動機づけに利用されている。さらに、多くの神経認知/行動タスクは、最初はラット10のために設計されました。ここでは、ラットにおけるEcoHIVの立体的注射の利用を報告し、EcoHIV感染モデルを拡張し、ニューロHIVおよびHANDに関連する新しい質問に対処する重要な機会を与える。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>293FT cells</td>
			<td>ThermoFisher Scientific</td>
			<td>R70007</td>
			<td></td>
		</tr>
		<tr>
			<td>Antibiotic-Antimycotic solution</td>
			<td>Cellgro</td>
			<td>30004CI</td>
			<td>100X</td>
		</tr>
		<tr>
			<td>Corning BioCoatGelatin 75cm&#178; Rectangular Canted Neck Cell Culture Flask with Vented Cap</td>
			<td>Life Technologies</td>
			<td>354488</td>
			<td></td>
		</tr>
		<tr>
			<td>Corning DMEM with L-Glutamine, 4.5 g/L Glucose and Sodium Pyruvate</td>
			<td>Life Technologies</td>
			<td>10013CV</td>
			<td></td>
		</tr>
		<tr>
			<td>Cover glass</td>
			<td>VWR</td>
			<td>637-137</td>
			<td></td>
		</tr>
		<tr>
			<td>drill</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Dumont #5 Forceps</td>
			<td>World Precision Instruments</td>
			<td>14095</td>
			<td></td>
		</tr>
		<tr>
			<td>Dumont #7 Forceps</td>
			<td>World Precision Instruments</td>
			<td>14097</td>
			<td></td>
		</tr>
		<tr>
			<td>Eppendorf Snap-Cap Microcentrifuge Biopur Safe-Lock Tubes</td>
			<td>Life Technologies</td>
			<td>22600028</td>
			<td></td>
		</tr>
		<tr>
			<td>Ethicon Vicryl Plus Antibacterial, 4-0 Polyglactin 910 Suture, 27in. FS-2</td>
			<td>Med Vet International</td>
			<td>VCP422H</td>
			<td></td>
		</tr>
		<tr>
			<td>Hamilton syringe</td>
			<td>Hamilton</td>
			<td>1701</td>
			<td></td>
		</tr>
		<tr>
			<td>Invitrogen Lipofectamine 3000 Transfection Reagent</td>
			<td>Life Technologies</td>
			<td>L3000015</td>
			<td></td>
		</tr>
		<tr>
			<td>Iris Forceps</td>
			<td>World Precision Instruments</td>
			<td>15914</td>
			<td></td>
		</tr>
		<tr>
			<td>Iris Scissors</td>
			<td>World Precision Instruments</td>
			<td>500216</td>
			<td></td>
		</tr>
		<tr>
			<td>Lentivirus-Associated p24 ELISA Kit</td>
			<td>Cell Biolabs, inc.</td>
			<td>VPK-107-5</td>
			<td></td>
		</tr>
		<tr>
			<td>Lenti-X Concentrator</td>
			<td>Takara</td>
			<td>PT4421-2</td>
			<td></td>
		</tr>
		<tr>
			<td>Opti-MEM I Reduced Serum Medium</td>
			<td>Life Technologies</td>
			<td>11058021</td>
			<td></td>
		</tr>
		<tr>
			<td>Paraformaldehyde</td>
			<td>Sigma-Aldrich</td>
			<td>158127-500G</td>
			<td></td>
		</tr>
		<tr>
			<td>Paraformaldehyde</td>
			<td>Sigma</td>
			<td>P6148</td>
			<td></td>
		</tr>
		<tr>
			<td>ProLong Gold</td>
			<td>Fisher Scientific</td>
			<td>P36930</td>
			<td></td>
		</tr>
		<tr>
			<td>Sevoflurane</td>
			<td>Merritt Veterinary Supply</td>
			<td>347075</td>
			<td></td>
		</tr>
		<tr>
			<td>stereotaxic apparatus</td>
			<td>Kopf Instruments</td>
			<td>Model 900</td>
			<td></td>
		</tr>
		<tr>
			<td>SuperFrost Plus Slides</td>
			<td>Fisher Scientific</td>
			<td>12-550-154%</td>
			<td></td>
		</tr>
		<tr>
			<td>Vannas Scissors</td>
			<td>World Precision Instruments</td>
			<td>500086</td>
			<td></td>
		</tr>
	</tbody>
</table>

a rat model of ecohiv brain infection

EcoHIV感染マウスモデルは、HIV関連の神経学的合併症を調査する上で重要な有用性を有することがよく研究されている。薬物乱用および神経認知障害の研究のためのEcoHIV感染ラットモデルの確立は、神経HIVおよびHIV-1関連神経認知障害(HAND)の研究において有益であろう。本研究では、キメラHIV(EcoHIV)を用いた活動性HIV感染のラットモデルの作成に成功したことを実証する。まず、EcoHIVのレンチウイルス構築物を、培養293 FT細胞に48時間包装した。次いで、条件付き培地を濃縮し、力を入れた。次に、F344/Nラット脳組織へのEcoHIV-EGFPの両側立体態注射を行った。感染の1週間後、EGFP蛍光シグナルが感染した脳組織で検出され、EcoHIVがラットに活性HIV感染を誘導することに成功したことを示している。また、ミクログリア細胞マーカーに対する免疫染色を、Iba1、実施した。結果は、ミクログリアがEcoHIVを収容する主要な細胞型であることを示した。さらに、EcoHIVラットは、感染後8週間に、HANDの潜在的な神経行動メカニズムおよびシナプス機能障害である時間処理に変化を示した。全体として、本研究は、HIV-1感染のEcoHIVモデルをラットに拡張し、脳内のHIV-1ウイルス貯留層ならびにHANDおよび薬物乱用などの関連する併存疾患を研究するための貴重な生物学的システムを提供する。

このコンディション培地は、293FT細胞に感染したEcoHIV-EGFPのレンチウイルスから採取した。次に、濃縮し、力を合わせた後、F344/Nラットの脳(皮質領域)に立体的に注入した。注射後7日間、ラットを屠殺し、ブレグマ5.64mmからブレグマ-4.68mmまでのコロナ脳スライスから画像を撮影した。 図1Aでは、脳全体、特に皮質および海馬歯状の歯素回に有意なEcoHIV-EGFPシグナルがある...

Watch this Scientific Journal Video about A Rat Model of EcoHIV Brain Infection at JoVE.com

A Rat Model of EcoHIV Brain Infection

ここでは、キメラHIV(EcoHIV)を用いた活性HIV感染の新しいラットモデルを確立するプロトコルを提示し、脳内のHIV-1ウイルス貯留層の理解を深め、HIV関連神経認知障害および関連する併存疾患(すなわち薬物乱用)を研究するシステムを提供するために重要である。

エコHIV脳感染症のラットモデル

It has been well studied that the EcoHIV infected mouse model is of significant utility in investigating HIV associated neurological complications. ...

ラットを使用して、このプロトコルは、キメラHIVを使用してアクティブHIV感染の新しいモデルを確立するために使用することができます。ラットでは、薬物乱用と神経認知障害の研究のためのEcoHIV感染モデルの確立は、神経HIVおよびHIV-1関連神経認知障害の研究に有益であり得る。デモンストレーションの手順は、私の研究室の研究員であるハイロン・リーです。ウイルスを293個のT細胞に包み込む場合、2つのゼラチンコーティングされた75平方センチメートルのフラスコのそれぞれに10%FBSを加えたDMEMのミリリットル当たり10〜5倍の293個のT細胞を種子化し、培養物が50%合流するまで37度で細胞を培養する。トランスフェクションの日に、1.5ミリリットルのマイクロ遠心チューブに750マイクロフラスコの培地で22.5マイクロリットルのトランスフェクション試薬を希釈し、溶液を3秒間渦巻く。キメラEcoHIVプラスミドDNAの20マイクログラムを250マイクロリットルの培地で20マイクログラム希釈し、フラスコ当たり1.5ミリリットルマイクロ遠心チューブを完全に混合します。希釈したDNAと希釈トランスフェクション試薬を穏やかな混合と組み合わせ、得られた溶液を室温で15分間インキュベートします。インキュベーションの終わりに、各ウイルス懸濁液を10ミリリットルのプリウォームDMEM培地に加え、各ウイルス補充溶液を293個のT細胞の1フラスコに加える。2日間の細胞培養の後、遠心分離のためにフラスコから上澄み液を1本の50ミリリットルの円錐管に溜め、浄化された上清を新しい50ミリリットルチューブに移す。透明化された上清に8ミリリットルのLenti-Xコンセントレータを加え、穏やかな反転と混ぜます。摂氏4度で2日間のインキュベーションを行った後、混合物を遠心分離し、慎重に上清を取り除き、100ミリモルPBSの100マイクロリットルでペレットを静かに再中断し、P24 ELISAキットを使用してウイルス濃度を引き立てる。EcoHIV-EGFP注射の場合、ペダル反射に対する応答の欠如を確認した後、脳領域から髪を剃り、70%エタノールとクロルヘキシジンベースのスクラブで露出した皮膚を2回殺菌する。立体的な装置でラットを起こしやすい位置に固定し、頭皮の正中線に沿って皮膚を通して5〜6センチメートルの切開を行います。1つの掘削位置を0.8ミリメートル横に、1つの1.2ミリメートルのロストラルをブレグマにマークし、各頭蓋骨の位置で直径0.4ミリメートルの穴を開けます。10マイクロリットル注射器に1ミリリットル当たり6ミリのトランスダクションユニットに1.04倍をロードし、ステレオタキシック装置に注射器を固定します。針を1つの穴の表面に近づけ、針を2.5ミリリットルの穴に差し込みます。1分間に0.2マイクロリットルのウイルスの速度で1マイクロリットルのウイルス溶液を注入する。すべてのウイルスが注入されたら、注射領域の中に針を5分間残してから、ゆっくりと針を引っ込めて、ラットの頭蓋骨の外に出るまで引き込みます。4-0シルク糸縫合糸で皮膚切開を閉じ、溶液を70%エタノールで殺菌し、ラットを回復室に入れ、再発まで監視する加熱パッドを付けます。ウイルス注入後1~8週間、ペダル反射に対する応答の欠如を確認した後、ラットをヒュームフード内の上皮の位置に固定し、胸部中線に沿って皮膚を切開する。横隔膜を切って胸腔を開き、20ゲージを25ミリメートル針ずつ左心室に挿入します。すぐにはさみで右心房を開き、毎分5ミリリットルの流量で50ミリリットルの予冷100ミリモルPBSを浸透させます。すべてのPBSが配信されたら、100ミリリットルの冷たい4%パラホルムアルデヒドをパーフューズします。すべての固定剤が送達されたら、頭蓋骨から脳を取り除き、新鮮な4%パラホルムアルデヒドで一晩組織を固定する。翌朝、脳がチューブの底に落ち着くまで、脳がチューブの底に落ち着くまで、30ミリモルPBSの30%スクロースの40ミリリットルにサンプルを入れ、マイナス80°Cで2分間メチルブタノールで脳組織を凍結する。凍結後、マイナス20度のクライオスタットと細かいペイントブラシを使用して、厚さ50ミクロンのコロナセクションを取得します。すべてのセクションが得られたら、300マイクロリットルのアンチフェード媒体でサンプルを覆い、22の50ミリリットルのカバーリップで取り付けます。培地が乾燥したら、共焦点顕微鏡で断面を画像化する。レンチウイルス注射の7日後、ラットコロナ脳スライス画像は、脳組織全体、特に皮質および海馬デンテート回内のEcoHIV-EGFPシグナルの有意な存在を明らかにする。Iba1およびEcoHIV-EGFPプローブとの二重標識は、ミクログリアが脳内のEcoHIV発現を有する主要な細胞型であるという強力な証拠を提供する。感染後8週間、EcoHIV注射動物は生理学コントロールと比較して比較的平坦な間隔関数によって証明されるように、刺激間隔の操作に対して相対的な鈍感を示す。さらに、EcoHIV注射ラットは、制御動物に対する頭頸部径が増加した短い樹状脊椎の相対的頻度の増加によって証明されるように、樹状脊椎形態に深い変化を示す。立体性注入前の条件培地の濃度と濃度は、実験全体で一貫した複製可能な結果を確保するために重要です。EcoHIVのこの特定の地域の立体態注入は、脳内の効率的なHIV感染を提供し、ラットの時間的処理の欠陥を生成する。ラットにおける立体性HIV注射を利用してEcoHIV感染モデルを拡張することは、目の前の神経HIVに関連する新しい質問に取り組む重要な機会を与える。

Research

JoVE Journal

Neuroscience

Gene Delivery to Postnatal Rat Brain by Non-ventricular Plasmid Injection and Electroporation

Creation of transgenic animals is a standard approach in studying functions of a gene of interest in vivo. However, many knockout or transgenic animals are not viable in those cases where the modified gene is expressed or deleted in the whole organism. Moreover, a variety of compensatory mechanisms often make it difficult to interpret the results. The compensatory effects can be alleviated by either timing the gene expression or limiting the amount of transfected cells.
The method of postnatal non-ventricular microinjection and in vivo electroporation allows targeted delivery of genes, siRNA or dye molecules directly to a small region of interest in the newborn rodent brain. In contrast to conventional ventricular injection technique, this method allows transfection of non-migratory cell types. Animals transfected by means of the method described here can be used, for example, for two-photon in vivo imaging or in electrophysiological experiments on acute brain slices.

This protocol describes a non-viral method of delivery of genetic constructs to a certain area of living rodent brain. The method consists of plasmid preparation, micropipette fabrication, neonatal rat pup surgery, microinjection of the construct, and in vivo electroporation.

非心室プラスミド注入し、エレクトロポレーションにより産後のラット脳への遺伝子デリバリー

Creation of transgenic animals is a standard approach in studying functions of a gene of interest in vivo. However, many knockout or transgenic animals ...

神経科学

Stereotaxic Surgery for Excitotoxic Lesion of Specific Brain Areas in the Adult Rat

Many behavioral functions in mammals, including rodents and humans, are mediated principally by discrete brain regions. A common method for discerning the function of various brain regions for behavior or other experimental outcomes is to implement a localized ablation of function. In humans, patient populations with localized brain lesions are often studied for deficits, in hopes of revealing the underlying function of the damaged area. In rodents, one can experimentally induce lesions of specific brain regions.
Lesion can be accomplished in several ways. Electrolytic lesions can cause localized damage but will damage a variety of cell types as well as traversing fibers from other brain regions that happen to be near the lesion site. Inducible genetic techniques using cell-type specific promoters may also enable site-specific targeting. These techniques are complex and not always practical depending on the target brain area. Excitotoxic lesion using stereotaxic surgery, by contrast, is one of the most reliable and practical methods of lesioning excitatory neurons without damaging local glial cells or traversing fibers.
Here, we present a protocol for stereotaxic infusion of the excitotoxin, N-methyl-D-aspartate (NMDA), into the basolateral amygdala complex. Using anatomical indications, we apply stereotaxic coordinates to determine the location of our target brain region and lower an injection needle in place just above the target. We then infuse our excitotoxin into the brain, resulting in excitotoxic death of nearby neurons. While our experimental subject of choice is a rat, the same methods can be applied to other mammals, with the appropriate adjustments in equipment and coordinates.
This method can be used on a variety of brain regions, including the basolateral amygdala1-6, other amygdala nuclei6, 7, hippocampus8, entorhinal cortex9 and prefrontal cortex10. It can also be used to infuse biological compounds such as viral vectors1, 11. The basic stereotaxic technique could also be adapted for implantation of more permanent osmotic pumps, allowing more prolonged exposure to a compound of interest.

Targeted ablation of specific brain region(s) by infusion of an excitotoxin using stereotaxic coordinates is described. This technique could also be adapted for infusion of other chemicals into the rat brain.

大人のラットの特定の脳領域の興奮毒性病変に対する定位手術

Many behavioral functions in mammals, including rodents and humans, are mediated principally by discrete brain regions. A common method for discerning the ...

A Rat Model of Mild Intrauterine Hypoperfusion with Microcoil Stenosis

Intrauterine hypoperfusion/ischemia is one of the major causes of intrauterine/fetal growth restriction, preterm birth, and low birth weight. Most studies of this phenomenon have been performed in either models with severe intrauterine ischemia or models with gradient degree of intrauterine hypoperfusion. No study has been performed in a model on uniform mild intrauterine hypoperfusion (MIUH). Two models have been used for studies of MIUH: a model based on suture ligation of either side of the arterial arcade formed with the uterine and ovarian arteries, and a transient model based on clipping the bilateral ovarian arteries and aorta having patency. Those two rodent models of MIUH have some limitations, e.g., not all fetuses are subjected to MIUH, depending on their position in the uterine horn. In our MIUH model, all fetuses are subjected to a comparable level of intrauterine hypoperfusion. MIUH was achieved by mild stenosis of all four arteries feeding the uterus, i.e., the bilateral uterine and ovarian arteries.
Arterial stenosis was induced by metal microcoils wrapped around the feeding arteries. Producing arterial stenosis with microcoils allowed us to control, optimize, and reproduce decreased blood flow with very little inter-animal variability and a low mortality rate, thus enabling accurate evaluation. When microcoils with an inner diameter of 0.24 mm were used, the blood flow in both the placenta and fetus was mildly decreased (approximately 30% from the pre-stenosis level in the placenta). The offspring of our MIUH model clearly demonstrates long-lasting alterations in neurological, neuroanatomical and behavioral test results.

Mild intrauterine hypoperfusion was produced by artery stenosis with metal microcoils wrapped around the uterine and ovarian arteries in rats at embryonic day 17. This procedure produced prenatal hypoperfusion and intrauterine growth restriction.

栓塞狭窄軽度子宮内血流の低下のモデルラット

Intrauterine hypoperfusion/ischemia is one of the major causes of intrauterine/fetal growth restriction, preterm birth, and low birth weight. Most studies ...

An In Vivo Assessment of Blood-Brain Barrier Disruption in a Rat Model of Ischemic Stroke

Ischemic stroke leads to vasogenic cerebral edema and subsequent primary brain injury, which is mediated through destruction of the blood-brain barrier (BBB). Rats with induced ischemic stroke were established and used as in vivo models to investigate the functional integrity of the BBB. Spectrophotometric detection of Evans blue (EB) in the brain samples with ischemic injury could provide reliable justification for the research and development of novel therapeutic modalities. This method generates reproducible results, and is applicable in any laboratory without a need for special equipment. Here, we present a visualized and technical guideline on the detection of the extravasation of EB following induction of ischemic stroke in rats.

An In Vivo Assessment of Blood-Brain Barrier Disruption in a Rat Model of Ischemic Stroke

The overall goal of this procedure is to provide a highly reproducible technique for in vivo assessment of the blood-brain barrier disruption in rat models of ischemic stroke.

虚血性脳卒中のモデルラットの血液脳関門の破壊のIn Vivo評価

Ischemic stroke leads to vasogenic cerebral edema and subsequent primary brain injury, which is mediated through destruction of the blood-brain barrier ...

A Rat Model of Central Fatigue Using a Modified Multiple Platform Method

In this article, we introduced a rat model of central fatigue using the modified multiple platform method (MMPM). The Multiple Platform box was designed as a water tank with narrow platforms on the bottom. The model rats were put into the tank and stood on the platforms for 14 h (18:00 - 8:00) per day for a consecutive 21 days, with a blank control group set for contrast. At the end of modeling, rats in the model group showed an obvious fatigued appearance. To assess the model, we performed several behavioral tests, including the open field test (OFT), the elevated plus maze (EPM) test, and the exhaustive swimming (ES) test. The results showed that anxiety, spatial cognition impairment, poor muscle performance, and declined voluntary activity presented in model rats confirm the diagnosis of&#160;central fatigue. Changes of the central neurotransmitters also verified the result. In conclusion, the model successfully simulated central fatigue, and future study with the model may help reveal the pathological mechanism of the disease.

Here, we present a protocol to introduce a rat model of central fatigue using the modified multiple platform method (MMPM).

ラット モデル改良を使用して中枢性疲労の複数プラットフォーム メソッド

In this article, we introduced a rat model of central fatigue using the modified multiple platform method (MMPM). The Multiple Platform box was designed ...

An Alternative Approach to Study Primary Events in Neurodegeneration Using Ex Vivo Rat Brain Slices

Despite&#160;numerous studies that attempt to develop reliable animal models&#160;which reflecting the primary processes underlying neurodegeneration, very few have been widely accepted. Here, we propose&#160;a new procedure&#160;adapted from the well-known ex vivo brain slice technique, which offers a closer in vivo-like scenario than in vitro preparations, for investigating the early events triggering cell degeneration, as observed in Alzheimer&#39;s disease (AD). This variation consists of simple and easily reproducible steps, which enable preservation of the anatomical cytoarchitecture of the selected brain region and its local functionality in a physiological milieu. Different anatomical areas can be obtained from the same brain, providing the opportunity to perform multiple experiments with the treatments in question in a site-, dose-, and time-dependent manner. Potential limitations&#160;which could affect the outcomes related to this methodology are related to the conservation of the tissue, i.e., the maintenance of its anatomical integrity during the slicing and incubation steps and the section thickness, which can influence the biochemical and immunohistochemical analysis. This approach can be employed for different purposes, such as exploring molecular mechanisms involved in physiological or pathological conditions, drug screening, or dose-response assays. Finally, this protocol could also reduce the number of animals employed in behavioral studies. The application reported here has been recently described and tested for the first time on ex vivo rat brain slices containing the basal forebrain (BF), which is one of the cerebral regions primarily affected in AD. Specifically, it has been demonstrated that the administration of a toxic peptide derived from the C-terminus of acetylcholinesterase (AChE) could prompt an AD-like profile, triggering, along the antero-posterior axis of the BF, a differential expression of proteins altered in AD, such as the alpha7 nicotinic receptor (&#945;7-nAChR), phosphorylated Tau (p-Tau), and amyloid beta (A&#946;).

An Alternative Approach to Study Primary Events in Neurodegeneration Using Ex Vivo Rat Brain Slices

We present a method which can&#160;provide further insights into the early events underlying neurodegeneration and based on the established ex-vivo brain technique, combining the advantages of in vivo and in vitro experiments. Moreover, it represents a unique opportunity for direct comparison of treated and untreated group in the same anatomical plane.

Ex Vivoラット脳切片を用いた神経変性疾患の主要なイベントを勉強する方法

Despite&#160;numerous studies that attempt to develop reliable animal models&#160;which reflecting the primary processes underlying neurodegeneration, ...

A Benchtop Approach to the Location Specific Blood Brain Barrier Opening using Focused Ultrasound in a Rat Model

Stereotaxic surgery is the gold standard for localized drug and gene delivery to the rodent brain. This technique has many advantages over systemic delivery including precise localization to a target brain region and reduction of off target side effects. However, stereotaxic surgery is highly invasive which limits its translational efficacy, requires long recovery times, and provides challenges when targeting multiple brain regions. Focused ultrasound (FUS) can be used in combination with circulating microbubbles to transiently open the blood brain barrier (BBB) in millimeter sized regions. This allows intracranial localization of systemically delivered agents that cannot normally cross the BBB. This technique provides a noninvasive alternative to stereotaxic surgery. However, to date this technique has yet to be widely adopted in neuroscience laboratories due to the limited access to equipment and standardized methods. The overall goal of this protocol is to provide a benchtop approach to FUS BBB opening (BBBO) that is affordable and reproducible and can therefore be easily adopted by any laboratory.

Focused ultrasound with microbubble agents can open the blood brain barrier focally and transiently. This technique has been used to deliver a wide range of agents across the blood brain barrier. This article provides a detailed protocol for the localized delivery to the rodent brain with or without MRI guidance.

ラットモデルにおける集中超音波を用いた位置特異的血液脳関門開口へのベンチトップアプローチ

Stereotaxic surgery is the gold standard for localized drug and gene delivery to the rodent brain. This technique has many advantages over systemic ...

A Hydrophobic Tissue Clearing Method for Rat Brain Tissue

Hydrophobic tissue clearing methods are easily adjustable, fast, and low-cost procedures that allows for the study of a molecule of interest in unaltered tissue samples. Traditional immunolabeling procedures require cutting the sample into thin sections, which restricts the ability to label and examine intact structures. However, if brain tissue can remain intact during processing, structures and circuits can remain intact for the analysis. Previously established clearing methods take significant time to completely clear the tissue, and the harsh chemicals can often damage sensitive antibodies. The iDISCO method quickly and completely clears tissue, is compatible with many antibodies, and requires no special lab equipment. This technique was initially validated for the use in mice tissue, but the current protocol adapts this method to image hemispheres of control and transgenic rat brains. In addition to this, the present protocol also makes several adjustments to preexisting protocol to provide clearer images with less background staining. Antibodies for Iba-1 and tyrosine hydroxylase were validated in the HIV-1 transgenic rat and in F344/N control rats using the present hydrophobic tissue clearing method. The brain is an interwoven network, where structures work together more often than separately of one another. Analyzing the brain as a whole system as opposed to a combination of individual pieces is the greatest benefit of this whole brain clearing method.

Here we present a hydrophobic tissue clearing method that allows for the viewing of target molecules as part of intact brain structures. This technique has now been validated for F344/N control and HIV-1 transgenic rats of both sexes.

ラット脳組織の疎水性組織除去法

Hydrophobic tissue clearing methods are easily adjustable, fast, and low-cost procedures that allows for the study of a molecule of interest in unaltered ...

Induction and Assessment of Levodopa-induced Dyskinesias in a Rat Model of Parkinson's Disease

Levodopa (L-DOPA) remains the gold-standard therapy used to treat Parkinson&#39;s disease (PD) motor symptoms. However, unwanted involuntary movements known as L-DOPA-induced dyskinesias (LIDs) develop with prolonged use of this dopamine precursor.&#160;It is estimated that the incidence of LIDs escalates to approximately 90% of individuals with PD within 10&#8211;15 years of treatment. Understanding the mechanisms of this malady and developing both novel and effective anti-dyskinesia treatments requires consistent and accurate modeling for pre-clinical testing of therapeutic interventions. A detailed method for reliable induction and comprehensive rating of LIDs following 6-OHDA-induced nigral lesioning in a rat model of PD is presented here. Dependable LID assessment in rats provides a powerful tool that can be readily utilized across laboratories to test emerging therapies focused on reducing or eliminating this common treatment-induced burden for individuals with PD.

Induction and Assessment of Levodopa-induced Dyskinesias in a Rat Model of Parkinson&#039;s Disease

This article describes methods to induce and evaluate levodopa-induced dyskinesias in a rat model of Parkinson&#39;s disease. The protocol offers detailed information regarding the intensity and frequency of a range of dyskinetic behaviors, both dystonic and hyperkinetic, providing a reliable tool to test treatments targeting this unmet medical need.

パーキンソン病のラットモデルにおけるレボドパ誘発性ジスキネジアの誘導と評価

Levodopa (L-DOPA) remains the gold-standard therapy used to treat Parkinson&#39;s disease (PD) motor symptoms. However, unwanted involuntary movements ...

mTBIラットモデルにおける傷害反応と行動評価の検証

Rat Model of Closed-Head Mild Traumatic Injury and its Validation

Animal models are crucial for advancing our understanding of mild traumatic brain injury (mTBI) and guiding clinical research. To achieve meaningful insights, developing a stable and reproducible animal model is essential. In this study, we report a detailed description of a closed-head mTBI model and a representative validation method using Sprague-Dawley rats to verify the modeling effect. The model involves dropping a 550 g mass weight from a height of 100 cm directly onto the head of a rat on a destructible surface, followed by a 180-degree turn. To assess the injury, rats underwent a series of neurobehavioral assessments 10 min post-injury, including time of loss of consciousness, first seeking-behavior time, escape ability, and beam balance ability test. During the acute and subacute stages following the injury, behavioral tests were conducted to assess motor coordination ability (Beam task), anxiety (Open Field test), and learning and memory abilities (Morris Water Maze test). The closed-head mTBI model produced a consistent injury response with minimal mortality and replicated real-life situations. The validation method effectively verified the model development and ensured the stability and consistency of the model.

著者スポットライト:外傷性脳損傷研究の進歩 - 正確な複製と迅速な評価のためのクローズドヘッドモデル 

Here, we present a closed-head mild traumatic brain injury (mTBI) rat model and its validation exhibiting remarkable similarity to human mTBI concerning behavioral manifestations during the acute and subacute stages.

閉鎖性軽度外傷のラットモデルとその検証

Animal models are crucial for advancing our understanding of mild traumatic brain injury (mTBI) and guiding clinical research. To achieve meaningful ...