Name: rat model of photochemically-induced posterior ischemic optic neuropathy
Uploaded: 2015-11-29T14:00:00.000+00:00
Duration: 14 min 54 s
Description: Watch this Scientific Journal Video about Rat Model of Photochemically-Induced Posterior Ischemic Optic Neuropathy at JoVE.com

我们感谢Eleut埃尔南德斯畜牧业，加布Gaidosh显微镜的专业知识，以及奎Tran和镇扬赵视频编辑。该研究由美国国家眼科研究所授予R01-EY022129到JLG和P30补助EY022589到加州大学圣地亚哥分校和EY014801到UM;美国心脏协会，詹姆斯和Esther国王基金会，复旦大学研究生院（编号2010033）的博士研究生交流计划资金，以及无限制的赠款防盲研究公司

所有动物的程序批准了加州大学圣地亚哥和迈阿密体制动物护理和使用委员会（IACUC）大学等按照ARVO声明对动物的眼科和视觉研究中的应用进行。所有试剂和外科手术使用的仪器是不育的。 1.麻醉和准备鼠外科手术<ol><li>前程序，将大鼠按照体重麻醉腹膜内注射氯胺酮（60毫克/千克）和赛拉嗪（8毫克/千克）的。麻醉足够的深度应到脚趾捏刺激了否定的答复来确定。 </li><li>麻醉后，将舌头向前，以防止窒息和应用润滑软膏双眼，以防止手术中角膜干燥。 </li><li>剃使用理发推子的手术部位和擦拭区域三次，用10％providone碘洗涤剂溶液和70％的乙醇。 </li><li>悬垂吨无菌区域内，他的动物。无菌手套和手术器械的过程中生存手术中使用。重新消毒使用的动物之间的热珠灭菌仪器的提示。 </li></ol> 2.手术方法<ol><li> PION感应 <ol><li> 股静脉插管 <ol><li>制备并清洗手术部位。剃使用理发推子的右大腿内侧和擦拭区域三次，每次用10％providone碘洗涤剂溶液和70％的乙醇。 </li><li>准备管。切聚乙烯管道的40厘米长度（PE 10）灭菌70％的乙醇。冲洗管道用盐水并将其连接到一个1ml注射器含有2％藻红乙染料预先测定溶液（1微升/毫克，得到20毫克/公斤体重的剂量）。装入注射器插入一个脚踏开关控输注泵设定为600微升/分钟的速率。 </li><li>使用15号刀片，做一个小的水平切口处右大腿的基础。剪切和传播里面的膜，清洁用无菌棉签的区域。 </li><li>分离钳肌肉直到股静脉的分支是可见的。鞘周围动脉，静脉和神经。捏，向上拉出此套钳（精尖杜蒙钳），切一小切口（2-4毫米通常就足够了）的三角形楔Vannas弹簧剪刀基地附近。展开切割是必要的。 </li><li>分离静脉和动脉与显微手术钩平行于静脉的方向上的钝。要小心，不要损坏精致膜，静脉分支。然后，轻轻提起静脉和它从底层结缔组织分开。 </li><li>获得无针尼龙缝合线，并把它旁边的股静脉。使用显微外科钩，提升静脉，并通过细尖镊子远侧区域的下方。抓住缝线的一端，并把它下面的静脉。紧紧结扎远端静脉。通过以类似的方式的第二缝合线下的近端静脉和使一个松散结。 </li><li>做一个小切口在靠近远端结扎Vannas弹簧剪刀静脉。如果必要，展开与细尖镊子孔。有些血液可能会泄漏通过晋级。清洁冷，无菌BSS和无菌棉签手术区。 </li><li>在切割的边缘保持静脉壁，导尿容器使用一针固定器制备的盐水冲洗管道。拧紧静脉周围和油管近结。然后，通过将其直接连接到所述远端的缝合锚的管道。 </li><li>检查导管的质量按下脚踏开关注入生理盐水，&lt;1 m1为充分。确保管道通畅，没有泄漏。暂时关闭缝合切口，以保护导管和组织。 </li></ol></li><li> 曝光视神经 <ol><li> PreparE中的手术部位通过擦拭预先刮净区域上面的左眼三次，每次用10％providone碘洗涤剂溶液和70％的乙醇。 </li><li>使沿皮肤2-3毫米的眼睛后面用第15号刀片的切口。捏住并解除与锯齿钳的结缔组织，并做一个小切口，Vannas弹簧剪刀。这个小切口通常为约5毫米的长度，但可以更长，以提供在训练外科医生更大的曝光。通过继续沿眼眶骨的上缘的结缔组织直言不讳地剖析，同时注意避免破坏血管。清洁用棉签手术区。 </li><li>通过结膜向下解剖直到上直肌是可见的。捏并剖析通过肌肉;肌肉会从深被解放的轨道内。现在，周围组织可用于在眼睛的缩回和仰角，以帮助为便于可视化。 </li><li>回覆道皮肤和结缔组织的瓣横向和向下，并保持在适当位置与缝合和止血钳。这将向前和向外转动眼睛，以揭示所述含脂肪鞘围绕视神经。 </li><li>小心地将一对尖锐的镊子，扩大平行于视神经分开周围鞘的结缔组织。请勿触摸视神经与镊子的尖头。 </li><li>视神经和周围鞘的5毫米长现在应该是可见的。微血管上的套表面的网络环绕视神经。这些将激光照射过程中有的放矢。 </li></ol></li><li> 静脉注射赤藓红B和激光照射 <ol><li>同时经营的激光照射装置从绿色激光保护自己戴上橙色护目镜在任何时候。打开激光，打开快门，调整峰值和平均值鲍威激光器的RS作为必要的。关闭快门。 </li><li>定位在激光照射装置的老鼠。为了确保适当的光束的位置，弱对准光束是通过钻在关闭快门叶片的直径为100微米的孔空间滤波的激光产生。再暴露有一对细尖镊子的视神经，和对准光束定位到3之间毫米和视神经头后面4毫米眶内视神经。 </li><li>通过激活输液泵的注入2％赤藓红B中的溶液。它可循环几秒钟，而外科医生增加一小滴的BSS的滋润视神经的表面上。 </li><li>验证瞄准光束的位置，然后单击脚踏开关来启动辐射。橙色安全过滤器，这是添加到显微镜的光路，将立即触发随后快门的一个1秒的延迟后开口。 </li><li>照射为9视神经0秒与135MW的一个峰值功率和18mW功率由光束斩波旋转在250赫兹以15％的占空比产生的平均功率（这可以减少热效应）。黄色荧光，通过安全过滤器可视化为亮橙色，将被从视神经的上表面发射的，并足以确保光束照射神经对称。 </li><li>在辐射后，橙色的安全过滤器将自动打开。微出血可以在某些情况下可以观察到。 </li><li>减轻对额外眼肌牵引和眼睛返回到中立位置。关闭切口间断缝合。然后撤出导管和打结股静脉严密，防止泄漏;关闭间断缝合。应用抗生素软膏，以两个切口。检查眼底验证视网膜中央静脉和动脉的血管完整性。 </li></ol></li></ol></li><li> 逆行标记。视网膜神经节细胞（RGC的）。注:为了评估RGC存活，用荧光金（FG）逆行标记应PION前一星期内完成。该方法中详细朱庇特协议819 13中描述 。 <ol><li>简而言之:麻醉与氯胺酮（60毫克/公斤）和甲苯噻嗪（8毫克/千克）的动物和剃光头。 </li><li>手术擦洗切口部位，使整个头部正中切口，暴露颅骨。 </li><li>钻透颅骨双侧孔（直径2×2毫米）0.5毫米无论从矢状缝。 </li><li>小心吸用真空泵即在于在上丘（SC）的背表面上的脑的内容。然后，将一小块明胶海绵用4％FG浸泡到SC的表面。 </li><li>关闭缝合切口和爱护使用标准的术后护理的动物。 </li></ol></li><li> 术后护理和安乐死 <ol><li>手术后，将动物在一个单独的干净笼子上的再循环水加热垫的顶部，直到动物进行回收。 </li><li>手术后止痛药（盐酸丁丙诺啡，0.01毫克/千克），应每天给予两次，连续三天，以减少不适。 </li><li>鼠应分开存放，并观察，直到他们能够保持胸骨斜卧，恢复足够的意识。 </li><li>回收和身体健康的迹象每天监测至少5天的手术后，或直到拆线和外科手术部位，为准最新的适当愈合。 </li><li>根据调查感兴趣的手术后安乐死的动物通过灌注4％PFA在科学上适当的时间点。 </li></ol></li></ol>

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The authors have nothing to disclose.

Here we describe in detail a method for inducing PION in a rat model. The most critical part of the protocol is the exposure and irradiation of the optic nerve &#8211; to expose the nerve as long as possible while avoiding damage caused by the sharp fine tip forceps or from stretching. In rats, the ophthalmic artery enters the optic nerve &#8804;1 mm from the optic nerve head. Therefore, irradiation of the optic nerve 3-4 mm away from the optic nerve head should only result in ischemia of the capillaries feeding the nerv...

患者50岁，缺血性视神经病变（ION）是最常见的类型的急性视神经病变2。前（AION）或后（π介子）3:条件可根据具体的受影响的血液供给和临床上介绍的源呈现为一个两种亚型的。虽然AION的发病机理和过程已被广泛研究4-7，PION一直由于其低流行，变呈现，不明确的诊断标准，缺乏动物模型知之甚少。此外，没有任何的治疗已被证明有效地防止或从AION或π介子反向视力丧失。因此，π介子可重复的和可靠的动物模型是很有价值的研究在体内的疾病过程和神经保护和轴突再生测试新的治疗方案。 化学诱发缺血性损伤导致vasog微血管ENIC水肿和血栓形成有效地创建区域组织缺血8-12。注射到血管循环后，光敏染料赤藓红B最终产生在激活活性单线氧分子靶血管激光照射。单重态氧直接peroxidizes血管内皮，刺激血小板粘附/聚集并导致闭塞性血栓的形成。缺血性损伤蔓延到邻近地区和微血管压迫因血管性水肿进一步加剧。此协议的总体目标是光化学诱发缺血的球后视神经镜像造成PION的损害。 据我们所知，这是在后视神经1缺血性损伤的第一模型。由于这种模式产生缺血，同时避免身体创伤，对后部缺血性视神经病变的生理过程是更好的模仿和学习。此外，这种模式提供了一个新的平台，为候选人为治疗视神经病变等中枢神经系统缺血性疾病的筛查。在这里，股静脉置管，视神经接触，静脉注射赤藓红的B和激光照射大鼠模型PION了详细的方案进行了说明。

<table><tbody><tr><td>532-nm Nd:YAG laser&amp;nbsp;</td><td>Laserglow</td><td>LRS-532-KM-200-3</td><td></td></tr><tr><td>Beam chopper</td><td>custom-made</td><td>custom-made</td><td></td></tr><tr><td>Mechanical shutter and corresponding shutter drive timer</td><td>Vincent Associates</td><td>SD-10</td><td></td></tr><tr><td>25-cm focal length spherical lens</td><td>CVI/Melles-Griot</td><td>01 LPX 293</td><td>plano-convexBK7 glass lens with HEBBAR&lt;sup&gt;TM&lt;/sup&gt; antireflection coating</td></tr><tr><td>Erythrosin B&amp;nbsp;</td><td>MP Biomedicals</td><td>190449</td><td></td></tr><tr><td>Fluorogold</td><td>Fluorochrome,LLC</td><td></td><td></td></tr><tr><td>Gelfoam</td><td>Cardinal Health</td><td>CAH1203421</td><td></td></tr><tr><td>Polyethylene tubing (PE10)</td><td>BD Intramedic</td><td>427400</td><td></td></tr><tr><td>No. 10 Blade</td><td>Miltex</td><td>4-110</td><td></td></tr><tr><td>Fine Forceps</td><td>F.S.T.</td><td>91150-20</td><td>Dumont #5 rustless non-magnetic</td></tr><tr><td>Forceps with Teeth</td><td>F.S.T.</td><td>11153-10</td><td>Germany stainless</td></tr><tr><td>Forceps</td><td>F.S.T.</td><td>18025-10&amp;nbsp;</td><td>Germany stainless</td></tr><tr><td>Vannas spring scissors</td><td>F.S.T.</td><td>2-220&amp;nbsp;</td><td>JJECK Stainless</td></tr><tr><td>Polyglactin suture</td><td>Ethicon</td><td>J488G</td><td>7-0 suture</td></tr><tr><td>hemostat</td><td>F.S.T.</td><td>12075-12&amp;nbsp;</td><td>Germany stainless</td></tr></tbody></table>

rat model of photochemically-induced posterior ischemic optic neuropathy

后部缺血性视神经病变（PION）是一见倾心，破坏性疾病的临床实践。然而，其发病机制与自然史仍然知之甚少。最近，我们开发PION的一个可靠的，可再现的动物模型和测试在这个模型1的一些神经营养因子的治疗效果。该视频的目的是表明我们光化学诱导的后部缺血性视神经病变模型，并评估其与视网膜神经节细胞的逆行标记作用。以下后视神经，光敏染料，赤藓红乙手术暴露，静脉注射和将激光束聚焦到视神经表面。赤藓红B和激光的期间，照射损伤血管内皮，促使微血管闭塞血小板血栓和水肿压缩介导的光化学相互作用。由此产生的缺血性损伤产生一个渐进的，但pronounc编视网膜神经节细胞枯死，由于亏损轴索输入 - 远程，损伤诱发和临床相关的结果。因此，这种模型提供了一种新颖的平台来研究PION的病理生理过程中，而且可用于测试治疗方法为视神经病变以及其它CNS缺血性疾病进一步优化。

诱导这种技术将所得的缺血性损伤的产生视网膜神经节细胞轴突缺血损伤后逐渐但明显的死亡。这类似于在人类疾病观察到临床上相关的结果。 FG逆行标记用于PION后量化RGC存活。相同的方法被用来验证一个模型的创建成功以及评估不同治疗方案的效果。 图1示出 FG阳性细胞的代表性共聚焦图像在视网膜平面坐骑控制（图1A），假治疗（激光只/无赤藓红B， 图1B），</s...

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Rat Model of Photochemically-Induced Posterior Ischemic Optic Neuropathy

该协议的目的是光化学诱导缺血性损伤至大鼠后视神经。这个模型是至关重要的后部缺血性视神经病变的病理生理的研究，以及治疗方法，这和其他视神经病变，以及其它CNS缺血性疾病。

光化学诱导后部缺血性视神经病变的大鼠模型

Posterior Ischemic optic neuropathy (PION) is a sight-devastating disease in clinical practice. However, its pathogenesis and natural history ...

rat-model-photochemically-induced-posterior-ischemic-optic

Research

JoVE Journal

Medicine

8.6K 次观看.  University of Miami Miller School of Medicine. 该协议的目的是光化学诱导缺血性损伤至大鼠后视神经。这个模型是至关重要的后部缺血性视神经病变的病理生理的研究，以及治疗方法，这和其他视神经病变，以及其它CNS缺血性疾病。 

视频: Rat Model of Photochemically-Induced Posterior Ischemic Optic Neuropathy

Photothrombotic Ischemia: A Minimally Invasive and Reproducible Photochemical Cortical Lesion Model for Mouse Stroke Studies

The photothrombotic stroke model aims to induce an ischemic damage within a given cortical area by means of photo-activation of a previously injected light-sensitive dye. Following illumination, the dye is activated and produces singlet oxygen that damages components of endothelial cell membranes, with subsequent platelet aggregation and thrombi formation, which eventually determines the interruption of local blood flow. This approach, initially proposed by Rosenblum and El-Sabban in 1977, was later improved by Watson in 1985 in rat brain and set the basis of the current model. Also, the increased availability of transgenic mouse lines further contributed to raise the interest on the photothrombosis model. Briefly, a photosensitive dye (Rose Bengal) is injected intraperitoneally and enters the blood stream. When illuminated by a cold light source, the dye becomes activated and induces endothelial damage with platelet activation and thrombosis, resulting in local blood flow interruption. The light source can be applied on the intact skull with no need of craniotomy, which allows targeting of any cortical area of interest in a reproducible and non-invasive way. The mouse is then sutured and allowed to wake up. The evaluation of ischemic damage can be quickly accomplished by triphenyl-tetrazolium chloride or cresyl violet staining. This technique produces infarction of small size and well-delimited boundaries, which is highly advantageous for precise cell characterization or functional studies. Furthermore, it is particularly suitable for studying cellular and molecular responses underlying brain plasticity in transgenic mice.

Photothrombosis is a quick, minimally-invasive technique for inducing small and well-delimited infarction in areas of interest in highly reproducible manner. It is particularly suitable for studying cellular and molecular responses underlying brain plasticity in transgenic mice.

光化学缺血：一种微创和可重复性的光化学皮质病变模型小鼠中风研究

The photothrombotic stroke model aims to induce an ischemic damage within a given cortical area by means of photo-activation of a previously injected ...

科研

医学

Photothrombosis-induced Focal Ischemia as a Model of Spinal Cord Injury in Mice

Spinal cord injury (SCI) is a devastating clinical condition causing permanent changes in sensorimotor and autonomic functions of the spinal cord (SC) below the site of injury. The secondary ischemia that develops following the initial mechanical insult is a serious complication of the SCI and severely impairs the function and viability of surviving neuronal and non-neuronal cells in the SC. In addition, ischemia is also responsible for the growth of lesion during chronic phase of injury and interferes with the cellular repair and healing processes. Thus there is a need to develop a spinal cord ischemia model for studying the mechanisms of ischemia-induced pathology. Focal ischemia induced by photothrombosis (PT) is a minimally invasive and very well established procedure used to investigate the pathology of ischemia-induced cell death in the brain. Here, we describe the use of PT to induce an ischemic lesion in the spinal cord of mice. Following retro-orbital sinus injection of Rose Bengal, the posterior spinal vein and other capillaries on the dorsal surface of SC were irradiated with a green light resulting in the formation of a thrombus and thus ischemia in the affected region. Results from histology and immunochemistry studies show that PT-induced ischemia caused spinal cord infarction, loss of neurons and reactive gliosis. Using this technique a highly reproducible and relatively easy model of SCI in mice can be achieved that would serve the purpose of scientific investigations into the mechanisms of ischemia induced cell death as well as the efficacy of neuroprotective drugs. This model will also allow exploration of the pathological changes that occur following SCI in live mice like axonal degeneration and regeneration, neuronal and astrocytic Ca2+ signaling using two-photon microscopy.

Photothrombosis is a minimally invasive and highly reproducible procedure to induce focal ischemia in the spinal cord and serves as a model of spinal cord injury in mice.

Photothrombosis引起的局部脑缺血脊髓损伤的小鼠模型

Spinal cord injury (SCI) is a devastating clinical condition causing permanent changes in sensorimotor and autonomic functions of the spinal cord (SC) ...

A Mouse Model of Retinal Ischemia-Reperfusion Injury Through Elevation of Intraocular Pressure

Retinal ischemia-reperfusion (I/R) is a pathophysiological process contributing to cellular damage in multiple ocular conditions, including glaucoma, diabetic retinopathy, and retinal vascular occlusions. Rodent models of I/R injury are providing significant insights into mechanisms and treatment strategies for human I/R injury, especially with regard to neurodegenerative damage in the retinal neurovascular unit. Presented here is a protocol for inducing retinal I/R injury in mice through elevation of intraocular pressure (IOP). In this protocol, the ocular anterior chamber is cannulated with a needle, through which flows the drip of an elevated saline reservoir. Using this drip to raise IOP above systolic arterial blood pressure, a practitioner temporarily halts inner retinal blood flow (ischemia). When circulation is reinstated (reperfusion) by removal of the cannula, severe cellular damage ensues, resulting ultimately in retinal neurodegeneration. Recent studies demonstrate inflammation, vascular permeability, and capillary degeneration as additional elements of this model. Compared to alternative retinal I/R methodologies, such as retinal arterial ligation, retinal I/R injury by elevated IOP offers advantages in its&#160;anatomical specificity, experimental tractability, and technical accessibility, presenting itself as a valuable tool for examining neuronal pathogenesis and therapy in the retinal neurovascular unit.

This article describes a procedure for inducing retinal ischemia-reperfusion injury by elevated intraocular pressure in mice. Retinal ischemia-reperfusion injury by elevated intraocular pressure serves to model human pathologies characterized by compromised oxygen and nutrient delivery in the retina, enabling researchers to examine potential cellular mechanisms and treatments for human diseases of the retinal neurovascular unit.

视网膜缺血再灌注损伤的小鼠模型通过眼压升高

Retinal ischemia-reperfusion (I/R) is a pathophysiological process contributing to cellular damage in multiple ocular conditions, including glaucoma, ...

The Rodent Model of Nonarteritic Anterior Ischemic Optic Neuropathy (rNAION)

Nonarteritic anterior ischemic optic neuropathy (NAION) is a focal ischemic lesion of the optic nerve that affects 1/700 individuals throughout their lifetime. NAION results in optic nerve edema, selective loss of the retinal ganglion cell neurons (RGCs) and atrophy of the optic nerve. A rodent model of NAION that expresses most NAION features and sequelae has been developed, which is applicable to both rats and mice. This model utilizes a focal laser application of 532 nm wavelength to illuminate a photoactive dye, Rose Bengal (RB), to cause capillary damage and leakage at the targeted anterior optic nerve (the laminar region). After rNAION induction, there is an early optic nerve ischemia, optic nerve edema, and intraneural inflammation, followed by selective RGC and axonal loss. Since the optic nerve is a CNS white matter tract, the rNAION model is applicable to mechanistic studies of selective white matter ischemia, as well as neuroprotective analyses and short and long-term mechanisms of glial and neuronal response to ischemia.

The Rodent Model of Nonarteritic Anterior Ischemic Optic Neuropathy rNAION

The following report describes how to replicate the rodent model of nonarteritic anterior ischemic optic neuropathy (rNAION), using the appropriate dye, contact lens and laser parameters. We also reveal the appropriate steps for evaluating the rNAION lesion in vivo.

非动脉炎性前部缺血性视神经病变的啮齿动物模型（rNAION）

Nonarteritic anterior ischemic optic neuropathy (NAION) is a focal ischemic lesion of the optic nerve that affects 1/700 individuals throughout their ...

Partial Optic Nerve Transection in Rats: A Model Established with a New Operative Approach to Assess Secondary Degeneration of Retinal Ganglion Cells

Previous studies have shown that the secondary degeneration of retinal ganglion cells (RGCs) occurs commonly in glaucoma. Partial optic nerve transection is considered a useful and reproducible model. Compared with other optic nerve injury models used commonly for assessing secondary degeneration, e.g. complete optic nerve transection and optic nerve crush models, the partial optic nerve transection model is superior as it distinguishes primary from secondary degeneration in situ. Therefore, it serves as an excellent tool for evaluating secondary degeneration. This study describes a novel operative approach of partial optic nerve transection by directly accessing the area of the retrobulbar optic nerve through the orbital lateral wall of the eyeball. Moreover, we present a newly designed, low cost surgical instrument to assist with transection. As demonstrated by the representative results in distinguishing the boundary of primary and secondary injury areas, the new approach and instrument ensures high efficiency and stability of the model by providing adequate space for surgical operation. This in turn makes it easy to separate the meningeal sheath and ophthalmic vessels from the optic nerve before transection. An additional benefit is that this space-saving operative approach improves the investigators&#39; ability to administer drugs, carriers, or selective RGC tracers to the stump of the partially transected optic nerve, allowing the exploration of mechanisms behind secondary injury in RGCs, in a new way.

Secondary degeneration of retinal ganglion cells (RGCs) occurs commonly in glaucoma. This study describes an innovative operative approach for partial optic nerve transection. The use of this space-saving operative approach extends the model's application range, and allows exploration of secondary injury mechanisms in RGCs in a new way.

大鼠部分视神经切断术: 建立一种新的手术方法评价视网膜神经节细胞二次变性的模型

Previous studies have shown that the secondary degeneration of retinal ganglion cells (RGCs) occurs commonly in glaucoma. Partial optic nerve transection ...

神经科学

A Fibrin-Enriched and tPA-Sensitive Photothrombotic Stroke Model

An ideal thromboembolic stroke model requires certain properties, including relatively simple surgical procedures with low mortality, a consistent infarction size and location, precipitation of platelet:fibrin intermixed blood clots similar to those in patients, and an adequate sensitivity to fibrinolytic treatment. The rose bengal (RB) dye-based photothrombotic stroke model meets the first two requirements but is highly refractory to tPA-mediated lytic treatment, presumably due to its platelet-rich, but fibrin-poor clot composition. We reason that combination of RB dye (50 mg/kg) and a sub-thrombotic dose of thrombin (80 U/kg) for photoactivation aimed at the proximal branch of middle cerebral artery (MCA) may produce fibrin-enriched and tPA-sensitive clots. Indeed, the thrombin and RB (T+RB)-combined photothrombosis model triggered mixed platelet:fibrin blood clots, as shown by immunostaining and immunoblots, and maintained consistent infarct sizes and locations plus low mortality. Moreover, intravenous injection of tPA (Alteplase, 10 mg/kg) within 2 h post-photoactivation significantly decreased the infarct size in T+RB photothrombosis. Thus, the thrombin-enhanced photothrombotic stroke model may be a useful experimental model to test novel thrombolytic therapies.

Traditional photothrombotic stroke (PTS) models mainly induce dense platelet aggregates of a high resistance to tissue plasminogen activator (tPA)-lytic treatment. Here a modified murine PTS model is introduced by co-injecting thrombin and photosensitive dye for photoactivation. The thrombin-enhanced PTS model produces mixed platelet:fibrin clots and is highly sensitive to tPA-thrombolysis.

纤维蛋白富集和 tPA 敏感的光血栓形成卒中模型

An ideal thromboembolic stroke model requires certain properties, including relatively simple surgical procedures with low mortality, a consistent ...

Optimization of the Retinal Vein Occlusion Mouse Model to Limit Variability

Mouse models of retinal vein occlusion (RVO) are often used in ophthalmology to study hypoxic-ischemic injury in the neural retina. In this report, a detailed method pointing out critical steps is provided with recommendations for optimization to achieve consistently successful occlusion rates across different genetically modified mouse strains. The RVO mouse model consists primarily of the intravenous administration of a photosensitizer dye followed by laser photocoagulation using a retinal imaging microscope attached to an ophthalmic guided laser. Three variables were identified as determinants of occlusion consistency. By adjusting the wait time after rose bengal administration and balancing the baseline and experimental laser output, the variability across experiments can be limited and a higher success rate of occlusions achieved. This method can be used to study retinal diseases that are characterized by retinal edema and hypoxic-ischemic injury. Additionally, as this model induces vascular injury, it can also be applied to study the neurovasculature, neuronal death, and inflammation.

Here, we describe an optimized protocol for retinal vein occlusion using rose bengal and a laser-guided retinal imaging microscope system with recommendations to maximize its reproducibility in genetically modified strains.

优化视网膜静脉遮挡小鼠模型以限制变异性

Mouse models of retinal vein occlusion (RVO) are often used in ophthalmology to study hypoxic-ischemic injury in the neural retina. In this report, a ...

Oxygen-Induced Retinopathy Model for Ischemic Retinal Diseases in Rodents

One of the commonly used models for ischemic retinopathies is the oxygen-induced retinopathy (OIR) model. Here we describe detailed protocols for the OIR model induction and its readouts in both mice and rats. Retinal neovascularization is induced in OIR by exposing rodent pups either to hyperoxia (mice) or alternating levels of hyperoxia and hypoxia (rats). The primary readouts of these models are the size of neovascular (NV) and avascular (AVA) areas in the retina. This preclinical in vivo model can be used to evaluate the efficacy of potential anti-angiogenic drugs or to address the role of specific genes in the retinal angiogenesis by using genetically manipulated animals. The model has some strain and vendor specific variation in the OIR induction which should be taken into consideration when designing the experiments.

Oxygen-induced retinopathy (OIR) can be used to model ischemic retinal diseases such as retinopathy of prematurity and proliferative diabetic retinopathy and to serve as a model for proof-of-concept studies in evaluating antiangiogenic drugs for neovascular diseases. OIR induces robust and reproducible neovascularization in the retina that can be quantified.

罗登斯缺血性视网膜疾病的氧诱导视网膜病变模型

One of the commonly used models for ischemic retinopathies is the oxygen-induced retinopathy (OIR) model. Here we describe detailed protocols for the OIR ...

A Murine Model of Ischemic Retinal Injury Induced by Transient Bilateral Common Carotid Artery Occlusion

Diverse vascular diseases such as diabetic retinopathy, occlusion of retinal veins or arteries and ocular ischemic syndrome can lead to retinal ischemia. To investigate pathological mechanisms of retinal ischemia, relevant experimental models need to be developed. Anatomically, a main retinal blood supplying vessel is the ophthalmic artery (OpA) and OpA originates from the internal carotid artery of the common carotid artery (CCA). Thus, disruption of CCA could effectively cause retinal ischemia. Here, we established a mouse model of retinal ischemia by transient bilateral common carotid artery occlusion (tBCCAO) to tie the right CCA with 6-0 silk sutures and to occlude the left CCA transiently for 2 seconds via a clamp, and showed that tBCCAO could induce acute retinal ischemia leading to retinal dysfunction. The current method reduces reliance on surgical instruments by only using surgical needles and a clamp, shortens occlusion time to minimize unexpected animal death, which is often seen in mouse models of middle cerebral artery occlusion, and maintains reproducibility of common retinal ischemic findings. The model can be utilized to investigate the pathophysiology of ischemic retinopathies in mice and further can be used for in vivo drug screening.

Here, we describe a mouse model of retinal ischemia by transient bilateral common carotid artery occlusion using simple sutures and a clamp. This model can be useful for understanding the pathological mechanisms of retinal ischemia caused by cardiovascular abnormalities.

由瞬态双边常见胡萝卜动脉闭塞引起的缺血性视网膜损伤的 Murine 模型

Diverse vascular diseases such as diabetic retinopathy, occlusion of retinal veins or arteries and ocular ischemic syndrome can lead to retinal ischemia. ...

Monitoring Dynamic Growth of Retinal Vessels in Oxygen-Induced Retinopathy Mouse Model

Oxygen-induced retinopathy (OIR) is widely used to study abnormal vessel growth in ischemic retinal diseases, including retinopathy of prematurity (ROP), proliferative diabetic retinopathy (PDR), and retinal vein occlusion (RVO). Most OIR studies observe retinal neovascularization at specific time points; however, the dynamic vessel growth in live mice along a time course, which is essential for understanding the OIR-related vessel diseases, has been understudied. Here, we describe a step-by-step protocol for the induction of the OIR mouse model, highlighting the potential pitfalls, and providing an improved method to quickly quantify areas of vaso-obliteration (VO) and neovascularization (NV) using immunofluorescence staining. More importantly, we monitored vessel regrowth in live mice from P15 to P25 by performing fluorescein fundus angiography (FFA) in the OIR mouse model. The application of FFA to the OIR mouse model allows us to observe the remodeling process during vessel regrowth.

This protocol describes a detailed method for the preparation and immunofluorescence staining of mice retinal flat mounts and analysis. The use of fluorescein fundus angiography (FFA) for mice pups and image processing are described in detail as well.

监测氧诱导视网膜病变小鼠模型中视网膜血管的动态生长

Oxygen-induced retinopathy (OIR) is widely used to study abnormal vessel growth in ischemic retinal diseases, including retinopathy of prematurity (ROP), ...