Name: using the electroretinogram to assess function in the rodent retina and the protective effects of remote limb ischemic preconditioning
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Description: Watch this Scientific Journal Video about Using the Electroretinogram to Assess Function in the Rodent Retina and the Protective Effects of Remote Limb Ischemic Preconditioning at JoVE.com

作者感谢沙龙Spana夫人的鼠害监测，处理和试验的帮助。博士的资金支持已经由悉尼大学和澳大利亚研究中心卓越视觉提供的。

该协议遵循悉尼大学的动物护理准则。 1.制作电极<ol><li>构造正极（一个其将接触角膜）从铂丝直径1-2毫米的短（5厘米）的长度。时尚它变成一个循环的几个毫米的直径。这个环路连接到一个常规的引线，足够长的时间，以达到你放大器的输入级（参见图4）。 </li><li>构造负极使用银/氯化银沉淀1-2毫米的直径，也连接到一个会议引线（这将在动物的嘴去）（ 见图4）。 </li><li>作为参比电极（将进入动物的臀部），用干净的皮下注射针头（23 G）的，也连接到一个适当长度的导线（ 见图4）。 </li><li>理想情况下，使用由仪器制造商提供的三引脚电缆，连接三个电极（正极û94;角膜，负→口，参考→臀部）的放大器。 </li></ol> 2.连接光刺激和ERG建立和校准<ol><li>创建（或定位）的小记录实验室，可制成暗。装备有一个以上的非长凳光发红色或红色头灯的任一个或两个。 </li><li>用照度计，以确认红色光的照度在安装过程中到达老鼠的眼睛不会超过1勒克斯。 注意:中性密度滤镜可用于降低灯的亮度和灯光的来源必须明确发出红光。如果光源发出低（可见光）波长暗适应将受到损害。 </li><li>封锁进入录音实验室所有杂散光（这往往需要恒心与不透明的胶带），并准备一个中性密度滤镜（可在张购买）足够大，以适应过来，等昏暗，任何计算机屏幕，您将在实验室。 注:杂散光和屏幕的光足以妨害暗适应鼠眼。 </li><li>连接放大器，以数据采集硬件。连接阳性，阴性和参考引线到放大器。确保计算机和LED Ganzfeld电源单元牢固地连接到一地源。 注:一些实验室有专门的接地点，连接到建筑物地面;水管是一种有效的替代。 </li><li>校准与科研质量的辐射计的LED光源。修复仪的传感器在该动物的眼睛将一个实验过程中所处位置。 </li><li>方案Ganzfeld LED的运行与逐步增加闪光能量，闪光持续时间，闪光灯重复和时间闪烁之间的全场ERG协议，称为interstimuls间隔（ISI），设置。举一个例子全场协议见表1。 注:全视野ERG从闪烁朦胧重复闪烁提高到b右闪在步骤明智的方式。双闪存程序遵循从满场的协议，使杆状和锥状响应隔离。 </li></ol> 3.前一天ERG实验<ol><li>录制黑暗中前适应SD大鼠12小时。是很方便的做到这一点，在记录实验室，一旦杂散光已被淘汰。 </li></ol> 4日ERG实验<ol><li>安排动物轻轻加热的同时录制。我们使用内置使动物的头可以放心在正确的点在入口到Ganzfeld轻金属平台。该平台具有内置管，通过它我们泵送水预热到40℃的水浴中。 注意:经验表明，这保持了动物的核心温度在37℃。 </li><li>权衡黑暗条件下的老鼠。记录体重和化妆正确的氯胺酮（60毫克/公斤）和甲苯噻嗪（5毫克/千克）剂量。抑制大鼠根TLY和腹腔注射麻醉剂。 </li><li>注意注射时间。一旦动物是无意识的（通常在5分钟），轻轻捏一只脚垫，看是否有反射反应存在检查麻醉深度。最好是等到这种反射不存在或弱，继续前。 </li><li>应用阿托品一滴proxmethacaine的，另一个角膜。 </li><li>切黑线的有10厘米长。使用一个简单的结的环路和滑环在眼睛的赤道。稍微拧紧;效果是吸引眼球微微前倾，以最小的压力。这样可以使角膜从眼皮清楚。 </li><li>应用卡波姆滴眼液在角膜表面。确保卡波姆保持在角膜表面，不会波及到眼皮或脸部。 </li><li>放置在加热平台的顶吸收床上用品。 </li><li>位置在老鼠的床上用品，用头在Ganzfeld开幕推荐的地方。 </li><li>插入INTernal温度探头插入直肠。用胶带探头线到尾位置的安全温度探头。 </li><li>插入基准电极（23政针）皮下后支腿，并连接到放大器。 </li><li>放置负电极（银/氯化银沉淀）牢固地固定在口中。为了防止这种情况滑出嘴角，加盖连接导线，以稳定的表面。 </li><li>定位正极在角膜的中心。采用显微，确保电极接触角膜平缓。 </li><li>查体温在37.0 - 37.5℃。 </li><li>一旦动物被适当地定位和电极的地方，悬垂整个安装（Ganzfeld和动物）用不透明材料（以保留暗适应）。我们用黑色的软布。 </li><li>在采集软件设置为2 kHz的采样速率为100-1000毫秒，5毫秒预收款采样的集合时间。设置带通滤波器，以1-1,000赫兹并确保采样触发品尝〜250毫秒以下闪光的时期。 </li><li>检查记录基线。它应该是免费的外来噪音，但表现出一定的放大器的噪声和振动的呼吸。 </li><li>如果基线显示多余的噪音，开始排除故障。大多数问题都与在电极位置，或接地滑脱。使用一个法拉第笼，以确保录音都是免费的外来噪音。 </li><li>运行测试闪存，0.4日志税赋cd.sm -2。一个ERG波形类似于图2A应该出现。在我们的0.4日志苏格兰人cd.sm -2闪光灯实验室典型的回答是:（a波:-474±39μV和b波:1,512±160μV，N = 11）。 </li><li>允许动物到暗重新适应10分钟。方便的是使用这些10分钟，以重新检查基线。 </li><li>稳定的信号的确认之后开始录制。 </li><li>在记录会话结束时，检查身体Temperat .. [温度茜保持。取出电极。重新卡波姆聚合物眼角膜。让动物恢复在散热垫，直到其完全的移动和积极的，返回到动物的住房之前。 </li></ol> 5.远程缺血<ol><li>在任何醒着还是anesthetised啮齿类动物进行远程缺血。 </li><li>如果将动物麻醉，把它在加热平台（上）和滑血压计袖带在后肢，明确膝关节的上部。 </li><li>如果动物被用于被处理，就可以不用麻醉执行此过程;这需要两个人。白岩松抑制动物轻轻第二应用血压计袖带和操作血压计。 </li><li>对于清醒的动物，用一块毛巾约15厘米×30〜50厘米轻轻包裹的动物，一个后肢免费。躺在动物在其上（说）左前臂背面，与持有人的手臂和躯干，以及地方之间的藏头袖带刚刚描述。 </li><li>袖带放气，并保证空气压力阀关闭。泵的袖口到160毫米汞柱的anesthetised动物，以及180毫米汞柱清醒的动物。这超出了收缩压（通常140毫米汞柱和160毫米汞柱分别）。 </li><li>保持这些压力的要求，使用手持式泵。 </li><li>之后在计划时间为缺血（我们使用的5分钟再灌注分离5分钟2期），松开气压阀放气袖带压力。 </li><li>确认远程缺血与附连到足垫皮肤温度探测器的作用。皮肤温度通常瀑布从32-30℃，超过5分钟，恢复再灌注。 </li></ol> 6.光损伤<ol><li>确保大鼠在暗适应过夜，之前的光损伤的过程。 </li><li>在肢体缺血（在我们的实验中无延迟）在适当的时候，每个动物被置于单独成有机玻璃盒，无线日水和食物地板为主的容器。 注意:光诱导损伤，只能在白化动物进行。 </li><li>开关上的预校准1000勒克斯的白光以标准时间（通常上午09点），并保持该条件24小时。 </li></ol> 7. ERG数据提取和分析<ol><li>获得ERG的平均波形。如果需要的话，校正非零基线，由减法。 </li><li>测量一个波（在中间呈现给高刺激强度）的振幅，作为基准，并在第一间的电压差（&lt;30毫秒延迟）槽（ 图1）。 </li><li>测量的b波振幅为a波的峰值和正下面波之间的电压差，通常是在80-100毫秒（ 图1）一个延迟发生。 </li><li>用傅立叶隔离振荡电位变换从60-235赫兹筛选数据，以90赫兹的过渡带<sup&gt; 7。如果需要分离的振荡电位的信号然后可从未经滤波波形中减去，以确认一个波槽的身份。 </li><li> a-和b-波峰的隐式时间（等待时间）也可以是一个有用的度量（ 图1）。使用双闪烁以分离杆响应。减去从混合反应（闪光1）隔离杆响应（ 图2）的视锥细胞反应（闪光2）。 </li><li>正常化个别光强度的波和b波振幅（后处理/后处理基线）或平均为治疗组。强度响应曲线绘制组幅度和对错误闪光能量。 </li></ol>

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	<li>Arden, G. B., Heckenlively, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Principles and practice of clinical electrophysiology of vision. , 139-183 (2006).</li><li>Bui, B. V., Fortune, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ganglion+cell+contributions+to+the+rat+full-field+electroretinogram.">Ganglion cell contributions to the rat full-field electroretinogram.</a> Journal of Physiology-London. 555 (1), 153-173 (2004).</li><li>Fortune, B., 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=Selective+ganglion+cell+functional+loss+in+rats+with+experimental+glaucoma.">Selective ganglion cell functional loss in rats with experimental glaucoma.</a> Investigative Ophthalmology &amp; Visual Science. 45 (6), 1854-1862 (2004).</li><li>Alarcon-Martinez, L., 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=Short+and+long+term+axotomy-induced+ERG+changes+in+albino+and+pigmented+rats.">Short and long term axotomy-induced ERG changes in albino and pigmented rats.</a> Molecular Vision. 15 (254-255), 2373-2383 (2009).</li><li>Lyubarsky, A. L., 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=Functionally+rodless+mice:+transgenic+models+for+the+investigation+of+cone+function+in+retinal+disease+and+therapy.">Functionally rodless mice: transgenic models for the investigation of cone function in retinal disease and therapy.</a> Vision Research. 42 (4), 401-415 (2002).</li><li>Bush, R. A., Sieving, P. 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> A PROXIMAL RETINAL COMPONENT IN THE PRIMATE PHOTOPIC ERG A-WAVE. Investigative Ophthalmology &amp; Visual Science. 35 (2), 635-645 (1994).</li><li>Liu, K., 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=Development+of+the+electroretinographic+oscillatory+potentials+in+normal+and+ROP+rats.">Development of the electroretinographic oscillatory potentials in normal and ROP rats.</a> Investigative Ophthalmology &amp; Visual Science. 47 (12), 5447-5452 (2006).</li><li>Casson, R. J., Wood, J. P. M., Melena, J., Chidlow, G., Osborne, N. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+ischemic+preconditioning+on+light-induced+photoreceptor+injury.">The effect of ischemic preconditioning on light-induced photoreceptor injury.</a> Investigative Ophthalmology &amp; Visual Science. 44 (3), 1348-1354 (2003).</li><li>Lawson, E. C., 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=Aerobic+Exercise+Protects+Retinal+Function+and+Structure+from+Light-Induced+Retinal+Degeneration.">Aerobic Exercise Protects Retinal Function and Structure from Light-Induced Retinal Degeneration.</a> Journal of Neuroscience. 34 (7), 2406-2412 (2014).</li><li>Grimm, C., 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=HIF-1-induced+erythropoietin+in+the+hypoxic+retina+protects+against+light-induced+retinal+degeneration.">HIF-1-induced erythropoietin in the hypoxic retina protects against light-induced retinal degeneration.</a> Nature Medicine. 8 (7), 718-724 (2002).</li><li>Weymouth, A. E., Vingrys, A. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rodent+electroretinography:+Methods+for+extraction+and+interpretation+of+rod+and+cone+responses.">Rodent electroretinography: Methods for extraction and interpretation of rod and cone responses.</a> Progress in Retinal and Eye Research. 27 (1), 1-44 (2008).</li><li>Bayer, A. U., Cook, P., Brodie, S. E., Maag, K. P., Mittag, T. <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+different+recording+parameters+to+establish+a+standard+for+flash+electroretinography+in+rodents.">Evaluation of different recording parameters to establish a standard for flash electroretinography in rodents.</a> Vision Research. 41 (17), 2173-2185 (2001).</li><li>Pugh, E. N., Lamb, T. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=AMPLIFICATION+AND+KINETICS+OF+THE+ACTIVATION+STEPS+IN+PHOTOTRANSDUCTION.">AMPLIFICATION AND KINETICS OF THE ACTIVATION STEPS IN PHOTOTRANSDUCTION.</a> Biochimica Et Biophysica Acta. 1141 (2-3), 111-149 (1993).</li><li>Breton, M. E., Schueller, A. W., Lamb, T. D., Pugh, E. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=ANALYSIS+OF+ERG+A-WAVE+AMPLIFICATION+AND+KINETICS+IN+TERMS+OF+THE+G-PROTEIN+CASCADE+OF+PHOTOTRANSDUCTION.">ANALYSIS OF ERG A-WAVE AMPLIFICATION AND KINETICS IN TERMS OF THE G-PROTEIN CASCADE OF PHOTOTRANSDUCTION.</a> Investigative Ophthalmology &amp; Visual Science. 35 (1), 295-309 (1994).</li><li>Mizota, A., Adachi-Usami, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+body+temperature+on+electroretinogram+of+mice.">Effect of body temperature on electroretinogram of mice.</a> Investigative Ophthalmology &amp; Visual Science. 43 (12), 3754-3757 (2002).</li><li>Szabo-Salfay, O., 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=The+electroretinogram+and+visual+evoked+potential+of+freely+moving+rats.">The electroretinogram and visual evoked potential of freely moving rats.</a> Brain Research Bulletin. 56 (1), 7-14 (2001).</li><li>Charng, J., 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=Conscious+Wireless+Electroretinogram+and+Visual+Evoked+Potentials+in+Rats.">Conscious Wireless Electroretinogram and Visual Evoked Potentials in Rats.</a> Plos One. 8 (9), (2013).</li><li>Galambos, R., Juhasz, G., Kekesi, A. K., Nyitrai, G., Szilagyi, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=NATURAL+SLEEP+MODIFIES+THE+RAT+ELECTRORETINOGRAM.">NATURAL SLEEP MODIFIES THE RAT ELECTRORETINOGRAM.</a> Proceedings of the National Academy of Sciences of the United States of America. 91 (11), 5153-5157 (1994).</li><li>Galambos, R., Szabo-Salfay, O., Szatmar, E., Szilagyi, N., Juhasz, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sleep+modifies+retinal+ganglion+cell+responses+in+the+normal+rat.">Sleep modifies retinal ganglion cell responses in the normal rat.</a> Proceedings of the National Academy of Sciences of the United States of America. 98 (4), 2083-2088 (2001).</li><li>Guarino, I., Loizzo, S., Lopez, L., Fadda, A., Loizzo, 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+chronic+implant+to+record+electroretinogram,+visual+evoked+potentials+and+oscillatory+potentials+in+awake,+freely+moving+rats+for+pharmacological+studies.">A chronic implant to record electroretinogram, visual evoked potentials and oscillatory potentials in awake, freely moving rats for pharmacological studies.</a> Neural Plasticity. 11 (3-4), 241-250 (2004).</li><li>Huang, J. C., Salt, T. E., Voaden, M. J., Marshall, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=NON-COMPETITIVE+NMDA-RECEPTOR+ANTAGONISTS+AND+ANOXIC+DEGENERATION+OF+THE+ERG+B-WAVE+IN-VITRO.">NON-COMPETITIVE NMDA-RECEPTOR ANTAGONISTS AND ANOXIC DEGENERATION OF THE ERG B-WAVE IN-VITRO.</a> Eye (London). 5 (4), 476-480 (1991).</li><li>Sasovetz, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> KETAMINE HYDROCHLORIDE - EFFECTIVE GENERAL ANESTHETIC FOR USE IN ELECTRORETINOGRAPHY. Annals of Ophthalmology. 10 (11), 1510-1514 (1978).</li><li>Mojumder, D. K., Wensel, T. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Topical+Mydriatics+Affect+Light-Evoked+Retinal+Responses+in+Anesthetized+Mice).">Topical Mydriatics Affect Light-Evoked Retinal Responses in Anesthetized Mice).</a> Investigative Ophthalmology &amp; Visual Science. 51 (1), 567-576 (2010).</li><li>Fraunfel, F. t., Burns, R. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=ACUTE+REVERSIBLE+LENS+OPACITY+-+CAUSED+BY+DRUGS,+COLD,+ANOXIA,+ASPHYXIA,+STRESS,+DEATH+AND+DEHYDRATION.">ACUTE REVERSIBLE LENS OPACITY - CAUSED BY DRUGS, COLD, ANOXIA, ASPHYXIA, STRESS, DEATH AND DEHYDRATION.</a> Experimental Eye Research. 10 (1), 19 (1970).</li><li>Calderone, L., Grimes, P., Shalev, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=ACUTE+REVERSIBLE+CATARACT+INDUCED+BY+XYLAZINE+AND+BY+KETAMINE-XYLAZINE+ANESTHESIA+IN+RATS+AND+MICE.">ACUTE REVERSIBLE CATARACT INDUCED BY XYLAZINE AND BY KETAMINE-XYLAZINE ANESTHESIA IN RATS AND MICE.</a> Experimental Eye Research. 42 (4), 331-337 (1986).</li><li>Behn, D., 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=Dark+adaptation+is+faster+in+pigmented+than+albino+rats.">Dark adaptation is faster in pigmented than albino rats.</a> Documenta Ophthalmologica. 106 (2), 153-159 (2003).</li><li>Sugawara, T., Sieving, P. A., Bush, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantitative+relationship+of+the+scotopic+and+photopic+ERG+to+photoreceptor+cell+loss+in+light+damaged+rats.">Quantitative relationship of the scotopic and photopic ERG to photoreceptor cell loss in light damaged rats.</a> Experimental Eye Research. 70 (5), 693-705 (2000).</li><li>Machida, S., 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=P23H+rhodopsin+transgenic+rat:+Correlation+of+retinal+function+with+histopathology.">P23H rhodopsin transgenic rat: Correlation of retinal function with histopathology.</a> Investigative Ophthalmology &amp; Visual Science. 41 (10), 3200-3209 (2000).</li><li>Brandli, A., Stone, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Remote+Ischemia+Influences+the+Responsiveness+of+the+Retina.">Remote Ischemia Influences the Responsiveness of the Retina.</a> Observations in the Rat. Investigative Ophthalmology &amp; Visual Science. 55 (4), 2088-2096 (2014).</li><li>Maccarone, R., Di Marco, S., Bisti, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Saffron+supplement+maintains+morphology+and+function+after+exposure+to+damaging+light+in+mammalian+retina.">Saffron supplement maintains morphology and function after exposure to damaging light in mammalian retina.</a> Investigative Ophthalmology &amp; Visual Science. 49 (3), 1254-1261 (2008).</li><li>Hood, D. C., Birch, D. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assessing+abnormal+rod+photoreceptor+activity+with+the+a-wave+of+the+electroretinogram:+Applications+and+methods.">Assessing abnormal rod photoreceptor activity with the a-wave of the electroretinogram: Applications and methods.</a> Documenta Ophthalmologica. 92 (4), 253-267 (1996).</li><li>Robson, J. G., Frishman, L. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+rod-driven+a-wave+of+the+dark-adapted+mammalian+electroretinogram.">The rod-driven a-wave of the dark-adapted mammalian electroretinogram.</a> Progress in Retinal and Eye Research. 39, 1-22 (2014).</li><li>Hood, D. C., Birch, D. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+COMPUTATIONAL+MODEL+OF+THE+AMPLITUDE+AND+IMPLICIT+TIME+OF+THE+B-WAVE+OF+THE+HUMAN+ERG.">A COMPUTATIONAL MODEL OF THE AMPLITUDE AND IMPLICIT TIME OF THE B-WAVE OF THE HUMAN ERG.</a> Visual Neuroscience. 8 (2), 107-126 (1992).</li><li>Wachtmeister, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oscillatory+potentials+in+the+retina:+what+do+they+reveal.">Oscillatory potentials in the retina: what do they reveal.</a> Progress in Retinal and Eye Research. 17 (4), 485-521 (1998).</li></ol>

乔纳森·斯通是CSCM私人有限公司的董事

上述的暗适应闪光ERG方法是评估在大鼠视网膜功能的可靠方法。两者的a波和b波被光伤害减少。远程缺血预处理减轻光损伤诱导的减少在一个波和B波。这种防护视网膜功能的建议，远程缺血性预处理已引起神经保护作用，类似其他形式的防护预处理，如缺氧，缺血和运动8-10。记录设置中，光刺激的参数，和动物的状态 - 记录ERG的信号是由三组因素来决定。 录像设置 ERG的下降幅度，当电极放置不正确或准备不完全接地11。邻近的电气设备正确接地是重要的，以减少在记录噪声;如果显著的噪音仍然存在一个法拉唉笼应使用。正极应牢固地定位在角膜与确认起始ERG的全视野协议之前和在完成检查的位置的中心。重要的是，这种电极仅接触角膜;与眼睑接触，甚至晶须可降低信号幅度。松动棉线已经在这个协议被用来防止眼睑接触到正极。一些研究者已经开发了隐形眼镜与正极嵌入，以确保可靠的接触和防止眼睑接触12。 光刺激设置我们已经使用了刺激器提供了广谱的白光，从LED光源。其它光源适合作为光刺激，如氙气闪光灯的灯光和卤素灯照明，看到韦茅斯Vingrys光刺激11之间的比较。 LED灯的优点，但是，我s表示每个闪存和其能量的持续时间是易于编程和快速地复位在宽范围内的光强度。我们已经制定了一套分级的能量闪烁，这在暗适应灭鼠范围阈值（生产只是检测的响应）饱和（产生最大响应）的。 通过反复试验，我们已经建立了刺激间的间隔（峰峰），其确保响应于闪光的幅度是独立的先前相同强度的闪光灯。该闪光灯亮时，不再需要为这个独立的ISI。 还通过反复试验，我们已经建立了在每一个能量需要提供一个干净的信号响应的最小数目。平均多个响应总是会提供一个更清晰的信号。我们使用的最小值，以使能量系列可迅速完成（在我们的协议11分钟）;快速完成减少的变化，由于变化的麻醉状态和异体ws的时间为其它变量进行研究，如果需要的话。 该动物的状态动物的生理机能的几个参数是很重要的优化和规范获得的ERG录音。 温度从光致活化的外段的G蛋白偶联型光转导级联的产生的一个波信号;这个级联的动态是，像所有的酶促反应，温度依赖性13,14。在麻醉下鼠害容易发生低温和需要外部加热以保持37.5℃，中心温度在整个记录。如果体温下降超过1-2℃时，a波和b波振幅减小，并且其延迟增加15。 麻醉稳定ERG录音要求动物是一动不动。神经肌肉阻滞剂和anaesthetIC剂用于ERG实验来达到一种无意识的和静止状态。只是有清醒ERG录音大鼠16-20五份报告。在这些研究中，将电极进行手术预植入颅骨和两个这些研究测试对ERG 17,20麻醉的效果。 用于ERG记录的最常见的麻醉药一直氯胺酮和甲苯噻嗪的组合（在我们的实验中60毫克/千克氯胺酮和5mg / kg的甲苯噻嗪的使用）。这会影响比气态麻醉如异氟烷和氟烷ERG的更小，并且已经证明相对无毒，具有高回收率17,21,22。这种方法保持不动的动物为〜40分钟;半剂量可用于记录条件延长为相同的时间。由常研究直接比较ERG的有和没有麻醉并表明氯胺酮 - 赛拉嗪没有可测量扰动α-和B-的幅度和延迟波17。大多数研究者规范麻醉条件下，然后测试实验参数;麻醉药有些效果不能完全忽视。 眼部环境眼睛的生理需要维护，优化和标准化ERG记录。学生们应该是一个标准尺寸;这是与一个散瞳实现，施加如眼药水，以达到最大的扩张。在啮齿类动物，阿托品或福林使用23。角膜的水化通过施加之前的录音卡波姆聚合物的维持;这也稳定了正极和角膜之间的电传导。如果角膜变得脱水，角膜瘢痕及白内障的形成，可能会出现24。白内障的形成是在小鼠25更常见，并维持角膜水合的各种方法已被用于小鼠的ERG的录音，包括水性流体或以恒定流定做接触式的电极陷阱水化在角膜表面12。 视网膜自适应状态这是一个重大的变量。上面提供的协议被设计为确保视网膜是暗适应，其最敏感的状态。理想的是，着色大鼠需要3小时黑暗壳体为完全暗适应而不含颜料的动物，如Sprague Dawley大鼠，需要最少5小时26的。这是标准的做法暗ERG录音隔夜适应动物12小时。部分或全部适应光线可以通过打开的Ganzfeld刺激标准强度的背景光可轻松快速地实现。经过光适应，但是，全暗适应需要时间来实现;因此非常谨慎的建议，以确保眼睛不小心暴露在光线录制前。 ERG的记录技术是由限制上述决定因素（ 即 ERG和刺激的建立）和研究员ERG测试的能力。没有经验的研究人员可能有变量ERG录音。方差可以通过创建足够大的样本大小比较的结果，如在视觉功能减少或增益被减小。可替代地，ERG记录可以基线记录和治疗后的记录之间进行标准化。归一化的数据然后可以被分组和分析。当呈现ERG的数据，这是标准的做法，以显示组数据和有代表性的波形。 当所有上述的仔细控制，所述的ERG的振幅的视网膜的功能状态的量度。 ERG的是在振幅一致减少由光引起的损坏或基因诱导的变性27,28在感光层的枯竭。相反，可以在AMPL检测到干预如RIP的保护作用itude的ERG 29。 ERG的也被用于在证明在视网膜上8-10,30缺血预处理，低氧预处理，锻炼和饮食藏红花的保护作用。 虽然动力学建模基于光转导的光感受器已知生理事件生长的视紫红质的光转导级联的动力学的知识，以及视网膜的突触连接，已鼓励ERG产生的模型的开发，以及复杂的ERG波形分析是可能和我们的视网膜内层电路31的理解。例如，一个波动力学模型是基于发生光转期间和拟合模型使模型参数进行比较，如峰值响应，定时延迟和灵敏度14生化步骤。 建模的缺点是，它依赖于关于视网膜circuitr假设Y，也只能是作为信息的假设允许。鉴于这一缺点，在A波动力学模型最近已经批评一个过分简单化波动力学32。在光感受器变性的研究，一般不进行ERG波形分析是出于不同的原因。光感受器变性是常严重，导致视觉功能，因此，不保证没有一个波和b波参数的进一步分析8,9,27,30戏剧性的损失。无论如何，一个波和B波ERG模型已经通过在许多啮齿类动物的研究和ERG建模的详细信息标准的做法，对于一个波，b波和有机磷可以在研究由胡德，并审查发现文章韦茅斯和Vingrys，渔人和沃什梅斯特11,32-34。 总之，暗适应ERG方法提出可以记录有和没有神经保护干预s视网膜变性之间可测量的差异UCH远程缺血性预处理。到可靠的ERG记录的元素必需进行了描述。感光体和内视网膜功能的ERG测量是研究人员在研究视网膜变性，以及各种遗传性，生物制药和药理干预对视觉功能的影响是有用的。

ERG的是由视网膜响应于光产生的，并且从眼睛的角膜表面记录的电势。当记录条件精心管理，ERG的可以以各种不同的方式来评估视网膜功能被使用。在这里，我们介绍了如何记录"闪光ERG"，时产生的视网膜暴露在一个简短的，明亮的闪光在Ganzfeld背景呈现的潜力。该Ganzfeld分散的均匀光与光的闪光灯到达视网膜整体大致均匀。如果视网膜记录前暗适应和暗适应被保持为动物准备用于记录，ERG的获得是通过两个杆和视锥感光细胞生成的。 暗适应闪光ERG的具有特征的波形，这已在两个方面进行了分析。首先，ERG的波形的早期和晚期成分被区分开来，并与神经元的序人活化在视网膜上。最早的组件是一个短潜伏期负向电位的一个波（ 图1）。这之后是一个正向电位，称为b波。 b波的上升阶段表示振荡，这被认为是一个单独的部件（振荡电位或有机磷）。在一个波被认为是由光感受器产生，b波由内核层细胞和有机磷由无长突细胞1。 根据刺激强度，反应非常昏暗闪烁称为暗门槛响应是可能的。暗视阈响应被理解为从视网膜神经节细胞2-4生成的。第二，闪光ERG可以通过光适应来分离，或通过两个闪光协议如下所述，成棒状和锥驱动的组件。明视下的条件下，一个波是在大鼠未检测到，因为锥体人口低，但有机磷和b波是明确5。在灵长类动物，其视网膜具有较高的人口锥，既杆状和锥途径产生可探测的一个波6。 常从闪速ERG提取两个有用措施是a-和b-波的振幅，测量为在图1中，与图2中所示的典型闪光的反应。当感光体人口被减少，例如通过暴露于破坏力亮光，ERG的所有成分减少。神经保护措施，如远程缺血预处理（RIP），可以由a-和b-波（ 图3）的振幅的保存进行验证。综上所述，ERG的分析使得健康，光损伤和neuroprotected视网膜之间的比较。

<table><tbody><tr><td>PC computer</td><td></td><td></td><td></td></tr><tr><td>Powerlab, 4 channel acquistion hardware</td><td>AD Instruments</td><td>PL 35044</td><td>Acquistion of ERG</td></tr><tr><td>Animal Bio Amp</td><td>AD Instruments</td><td>FE 136</td><td>Amplifier for ERG</td></tr><tr><td>Lab chart</td><td>AD Instruments</td><td></td><td>Signal collection software</td></tr><tr><td>Ganzfield</td><td>Photometric solutions</td><td>FS-250A</td><td>Light stimulus</td></tr><tr><td>Ganzfield operating system</td><td>Photometric solutions</td><td></td><td></td></tr><tr><td>Research Radiometer</td><td>International light technologies</td><td>ILT-1700</td><td>calibrate light series</td></tr><tr><td>Lux meter</td><td></td><td>LX-1010B</td><td>check red light illumanation</td></tr><tr><td>Excel</td><td>Microsoft</td><td></td><td></td></tr><tr><td>Lead wires</td><td>AD Instruments</td><td></td><td>Connect postive, negative ground electrodes to amplifier</td></tr><tr><td>Lead wires - alligator</td><td>AD Instruments</td><td></td><td>ground ganzfield and acquistion hardware to computer</td></tr><tr><td>Platinum wire 95%</td><td>A&amp;amp;E metals</td><td></td><td>postive electrode</td></tr><tr><td>Mouth electrode Ag/AgCl Pellet</td><td>SDR</td><td>E205</td><td>negative electode</td></tr><tr><td>26 G needle</td><td>BD</td><td></td><td>ground electode</td></tr><tr><td>Water pump</td><td></td><td></td><td></td></tr><tr><td>Water bath</td><td></td><td></td><td></td></tr><tr><td>Tubing</td><td></td><td></td><td></td></tr><tr><td>Homeothermic blanket system with flexible probe</td><td>Harvard Appartus</td><td>507222F</td><td></td></tr><tr><td>Atropine 1% w/v</td><td>Bausch &amp;amp; Lomb</td><td></td><td>topical mydriasis</td></tr><tr><td>Proxmethycaine 0.5% w/v</td><td>Bausch &amp;amp; Lomb</td><td></td><td>topical anaesthetic</td></tr><tr><td>Visco tears eye drops</td><td>Novartis</td><td></td><td>carbomer polymer</td></tr><tr><td>Thread</td><td></td><td></td><td>retract eye lid</td></tr><tr><td>Tweezers</td><td></td><td></td><td></td></tr><tr><td>Reusable adhesive</td><td>Blu tac</td><td></td><td>Dim red headlamp. Affix electrodes</td></tr><tr><td>Absorbent bedding</td><td></td><td></td><td></td></tr><tr><td>Ketamil - ketamine 100 mg/ml - 50 ml</td><td>Troy Laboratories Pty Ltd</td><td></td><td>dissociative</td></tr><tr><td>Xylium - Xylazine 100 mg/ml - 50 ml</td><td>Troy Laboratories Pty Ltd</td><td></td><td>muscle relaxant</td></tr><tr><td>Scale</td><td></td><td></td><td></td></tr></tbody></table>

using the electroretinogram to assess function in the rodent retina and the protective effects of remote limb ischemic preconditioning

The ERG is the sum of all retinal activity. The ERG is usually recorded from the cornea, which acts as an antenna that collects and sums signals from the retina. The ERG is a sensitive measure of changes in retinal function that are pan-retinal, but is less effective for detecting damage confined to a small area of retina. In the present work we describe how to record the &#8216;flash&#8217; ERG, which is the potential generated when the retina is exposed to a brief light flash. We describe methods of anaesthesia, mydriasis and corneal management during recording; how to keep the retina dark adapted; electrode materials and placement; the range and calibration of stimulus energy; recording parameters and the extraction of data. We also describe a method of inducing ischemia in one limb, and how to use the ERG to assess the effects of this remote-from-the-retina ischemia on retinal function after light damage. A two-flash protocol is described which allows isolation of the cone-driven component of the dark-adapted ERG, and thereby the separation of the rod and cone components. Because it can be recorded with techniques that are minimally invasive, the ERG has been widely used in studies of the physiology, pharmacology and toxicology of the retina. We describe one example of this usefulness, in which the ERG is used to assess the function of the light-damaged retina, with and without a neuroprotective intervention; preconditioning by remote ischemia.

该协议可以用于测量啮齿动物视网膜的体内视觉功能。在一个波，光感受器功能的量度，和b波，内视网膜功能的量度，被注释在图1中。 随光刺激的棒为主的ERG的信号增加时， 如图2A所示 。在〜0.4日志苏格兰人cd.sm -2和的一个波增加，直到饱和为2.5日志苏格兰人cd.sm振幅-2（未示出）的一个波变得很明显。双子闪光范式已经用于分离混合的ERG信号成锥体和杆分离物反应，如在图2B中 。 此ERG记录技术可以用来检查神经保护的干预措施。基线记录 ​​前一周光损伤是出现在图3A中完成。光损伤降低两者波和b波振幅，在图展示了URE 3B。远程缺血预处理能够减少ERG振幅的损失，如在图3C中看出。远程缺血技术取决于止血带上面的"拐点"的正确应用。不正确应用止血带并不妨碍光视网膜损伤， 如图3D所示。 <img alt="图1" src="/files/ftp_upload/52658/52658fig1highres.jpg" width="400" /> 图1:在一个波和b波从显示由暗适应眼睛的角膜记录到的光的明亮的闪光在所示时刻t0给出一个暗适应ERG跟踪的测量 。在一个波的幅度是从基线到第一槽（红色箭头）进行测定。 b波的振幅是从一个波到以下正峰值（蓝色箭头）的槽测量。隐式时间（等待时间），从刺激测定加工过的食品（T0），以对跟踪兴趣点，如A波（方括号）的低谷。 <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/52658/52658fig1highres.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图2" src="/files/ftp_upload/52658/52658fig2highres.jpg" width="700" /> 图2:暗适应ERG的发展而增加闪光灯强度和杆和视锥的响应分离示出了从一个暗适应眼睛角膜被记录到增加光的闪烁的痕迹。在一个波出现在明亮的强度。 （A）相比1.4至0.4日志逍遥法外cd.sm -2，高峰b波已经饱和，但一个波持续增长。在（B），双闪重叠。两个2.0日志苏格兰人cd.sm -2闪烁由500毫秒ISI分开。第一次闪光生成一个混合响应（黑色），而第二个闪蒸产生锥形只响应（虚线）。减去视锥细胞反应产生的隔离杆响应（灰色）。 <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/52658/52658fig2highres.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图3" src="/files/ftp_upload/52658/52658fig3highres.jpg" width="700" /> 图3:ERG提供视网膜的功能的量度代表波形，（C）的视网膜条件由RIP前暴露于有害的光示此处（A） 的正常视网膜，（B）中由光的视网膜损伤，和。 （四）视网膜抿着嘴由RIP调节，然后暴露于有害光线。相同的闪光能量被用来为每个记录（2.0日志cd.sm -2）。对于在 D记录的袖带压力在后肢被错误地放置和缺血不成立的。光损伤降低ERG（B）的幅度和RIP减轻了减少。 <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/52658/52658fig3highres.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图4" src="/files/ftp_upload/52658/52658fig4highres.jpg" width="400" /> 图4:近拍ERG电极的电极要建被示出，从左到右;正极接触角膜，负电极被放置在口中并且其由一个鳄鱼夹连接到针，然后将其插入到臀部皮下的参考电极。 <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/52658/52658fig4highres.jpg" target="_blank">请按此查看大版这个数字。</a>

Watch this Scientific Journal Video about Using the Electroretinogram to Assess Function in the Rodent Retina and the Protective Effects of Remote Limb Ischemic Preconditioning at JoVE.com

Using the Electroretinogram to Assess Function in the Rodent Retina and the Protective Effects of Remote Limb Ischemic Preconditioning

The electroretinogram (ERG) is an electrical potential generated by the retina in response to light. This paper describes how to use the ERG to assess retinal function, in dark-adapted rats, and how it can be can be used to assess a neuroprotective intervention, in the present case remote ischemic preconditioning.

使用电生理评估功能，在啮齿动物的视网膜和远程肢体缺血预适应的保护作用

The ERG is the sum of all retinal activity. The ERG is usually recorded from the cornea, which acts as an antenna that collects and sums signals from the ...

using-electroretinogram-to-assess-function-rodent-retina-protective

Universidad San Sebastian

Research

JoVE Journal

Neuroscience

16.1K 次观看.  University of Sydney. The electroretinogram (ERG) is an electrical potential generated by the retina in response to light. This paper describes how to use the ERG to assess retinal function, in dark-adapted rats, and how it can be can be used to assess a neuroprotective intervention, in the present case remote ischemic preconditioning.

视频: Using the Electroretinogram to Assess Function in the Rodent Retina and the Protective Effects of Remote Limb Ischemic Preconditioning

Implantation and Recording of Wireless Electroretinogram and Visual Evoked Potential in Conscious Rats

The full-field electroretinogram (ERG) and visual evoked potential (VEP) are useful tools to assess retinal and visual pathway integrity in both laboratory and clinical settings. Currently, preclinical ERG and VEP measurements are performed with anesthesia to ensure stable electrode placements. However, the very presence of anesthesia has been shown to contaminate normal physiological responses. To overcome these anesthesia confounds, we develop a novel platform to assay ERG and VEP in conscious rats. Electrodes are surgically implanted sub-conjunctivally on the eye to assay the ERG and epidurally over the visual cortex to measure the VEP. A range of amplitude and sensitivity/timing parameters are assayed for both the ERG and VEP at increasing luminous energies. The ERG and VEP signals are shown to be stable and repeatable for at least 4 weeks post surgical implantation. This ability to record ERG and VEP signals without anesthesia confounds in the preclinical setting should provide superior translation to clinical data.

We show surgical implantation and Recording procedures to measure visual electrophysiological signals from the eye (electroretinogram) and brain (visual evoked potential) in conscious rats, which is more analogous to the human condition where recordings are conducted without anesthesia confounds.

植入和无线电图记录和视觉诱发电位清醒大鼠

The full-field electroretinogram (ERG) and visual evoked potential (VEP) are useful tools to assess retinal and visual pathway integrity in both ...

科研

行为学

Direct-Coupled Electroretinogram (DC-ERG) for Recording the Light-Evoked Electrical Responses of the Mouse Retinal Pigment Epithelium

The retinal pigment epithelium (RPE) is a specialized monolayer of cells strategically located between the retina and the choriocapillaris that maintain the overall health and structural integrity of the photoreceptors. The RPE is polarized, exhibiting apically and basally located receptors or channels, and performs vectoral transport of water, ions, metabolites, and secretes several cytokines.
In vivo noninvasive measurements of RPE function can be made using direct-coupled ERGs (DC-ERGs). The methodology behind the DC-ERG was pioneered by Marmorstein, Peachey, and colleagues using a custom-built stimulation recording system and later demonstrated using a commercially available system. The DC-ERG technique uses glass capillaries filled with Hank&#8217;s buffered salt solution (HBSS) to measure the slower electrical responses of the RPE elicited from light-evoked concentration changes in the subretinal space due to photoreceptor activity. The prolonged light stimulus and length of the DC-ERG recording make it vulnerable to drift and noise resulting in a low yield of useable recordings. Here, we present a fast, reliable method for improving the stability of the recordings while reducing noise by using vacuum pressure to reduce/eliminate bubbles that result from outgassing of the HBSS and electrode holder. Additionally, power line artifacts are attenuated using a voltage regulator/power conditioner. We include the necessary light stimulation protocols for a commercially available ERG system as well as scripts for analysis of the DC-ERG components: c-wave, fast oscillation, light peak, and off response. Due to the improved ease of recordings and rapid analysis workflow, this simplified protocol is particularly useful in measuring age-related changes in RPE function, disease progression, and in the assessment of pharmacological intervention.

Direct-Coupled Electroretinogram DC-ERG for Recording the Light-Evoked Electrical Responses of the Mouse Retinal Pigment Epithelium

Here, we present a method for recording light-evoked electrical responses of the retinal pigment epithelium (RPE) in mice using a technique known as DC-ERGs first described by Marmorstein, Peachey, and colleagues in the early 2000s.

直接耦合电雷电图（DC-ERG），用于记录小鼠视网膜颜料皮皮的光唤起电响应

The retinal pigment epithelium (RPE) is a specialized monolayer of cells strategically located between the retina and the choriocapillaris that maintain ...

化学

Remote Limb Ischemic Preconditioning:  A Neuroprotective Technique in Rodents

Sublethal ischemia protects tissues against subsequent, more severe ischemia through the upregulation of endogenous mechanisms in the affected tissue. Sublethal ischemia has also been shown to upregulate protective mechanisms in remote tissues. A brief period of ischemia (5-10 min) in the hind limb of mammals induces self-protective responses in the brain, lung, heart and retina. The effect is known as remote ischemic preconditioning (RIP). It is a therapeutically promising way of protecting vital organs, and is already under clinical trials for heart and brain injuries. This publication demonstrates a controlled, minimally invasive method of making a limb &#8211; specifically the hind limb of a rat &#8211; ischemic. A blood pressure cuff developed for use in human neonates is connected to a manual sphygmomanometer and used to apply 160 mmHg pressure around the upper part of the hind limb. A probe designed to detect skin temperature is used to verify the ischemia, by recording the drop in skin temperature caused by pressure-induced occlusion of the leg arteries, and the rise in temperature which follows release of the cuff. This method of RIP affords protection to the rat retina against bright light-induced damage and degeneration.

Remote ischemic preconditioning (RIP) is a method of conditioning tissues against damaging stress. We have established a method of remote ischemia at the hind limb, by inflating a sphygmomanometer cuff for 5-10 min. The neuroprotective capabilities of RIP have been demonstrated in a model of retinal degeneration in rodents.

远程肢体缺血预适应:在啮齿动物的保护技术

Sublethal ischemia protects tissues against subsequent, more severe ischemia through the upregulation of endogenous mechanisms in the affected tissue. ...

医学

Glutamate and Hypoxia as a Stress Model for the Isolated Perfused Vertebrate Retina

Neuroprotection has been a strong field of investigation in ophthalmological research in the past decades and affects diseases such as glaucoma, retinal vascular occlusion, retinal detachment, and diabetic retinopathy. It was the object of this study to introduce a standardized stress model for future preclinical therapeutic testing.
Bovine retinas were prepared and perfused with an oxygen saturated standard solution, and the ERG was recorded. After recording stable b-waves, hypoxia (pure N2) or glutamate stress (250 &#181;m glutamate) was exerted for 45 min. To investigate the effects on photoreceptor function alone, 1 mM aspartate was added to obtain a-waves. ERG-recovery was monitored for 75 min.
For hypoxia, a decrease in a-wave amplitude of 87.0% was noted (p &#60;0.01) after an exposition time of 45 min (decrease of 36.5% after the end of the washout p = 0.03). Additionally, an initial decrease in b-wave amplitudes of 87.23% was recorded, that reached statistical significance (p &#60;0.01, decrease of 25.5% at the end of the washout, p = 0.03).
For 250 &#181;m glutamate, an initial 7.8% reduction of a-wave amplitudes (p &#62;0.05) followed by a reduction of 1.9% (p &#62;0.05). A reduction of 83.7% of b-wave amplitudes (p &#60;0.01) was noted; after a washout of 75 min the reduction was 2.3% (p = 0.62). In this study, a standardized stress model is presented that may be useful to identify possible neuroprotective effects in the future.

With this study, we introduce a standardized stress model for the isolated superfused bovine retina for future preclinical therapeutic testing. The effect of either hypoxia (pure N2) or glutamate stress (250 &#181;M glutamate) on retinal function represented by a- and b-wave amplitudes was evaluated.

谷氨酸和缺氧作为离体脊椎动物视网膜应力模型

Neuroprotection has been a strong field of investigation in ophthalmological research in the past decades and affects diseases such as glaucoma, retinal ...

Simultaneous Recording of Electroretinography and Visual Evoked Potentials in Anesthetized Rats

The electroretinogram (ERG) and visual evoked potential (VEP) are commonly used to assess the integrity of the visual pathway. The ERG measures the electrical responses of the retina to light stimulation, while the VEP measures the corresponding functional integrity of the visual pathways from the retina to the primary visual cortex following the same light event. The ERG waveform can be broken down into components that reflect responses from different retinal neuronal and glial cell classes. The early components of the VEP waveform represent the integrity of the optic nerve and higher cortical centers. These recordings can be conducted in isolation or together, depending on the application. The methodology described in this paper allows simultaneous assessment of retinal and cortical visual evoked electrophysiology from both eyes and both hemispheres. This is a useful way to more comprehensively assess retinal function and the upstream effects that changes in retinal function can have on visual evoked cortical function.

This protocol describes simultaneous measurement of electroretinogram and visual evoked potentials in anesthetized rats.

电图同时记录和麻醉大鼠视觉诱发电位

The electroretinogram (ERG) and visual evoked potential (VEP) are commonly used to assess the integrity of the visual pathway. The ERG measures the ...

神经科学

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, ...

Optimizing the Setup and Conditions for Ex Vivo Electroretinogram to Study Retina Function in Small and Large Eyes

Measurements of retinal neuronal light responses are critical to investigating the physiology of the healthy retina, determining pathological changes in retinal diseases, and testing therapeutic interventions. The ex vivo electroretinogram (ERG) allows the quantification of contributions from individual cell types in the isolated retina by addition of specific pharmacological agents and evaluation of tissue-intrinsic changes independently of systemic influences. Retinal light responses can be measured using a specialized ex vivo ERG specimen holder and recording setup, modified from existing patch clamp or microelectrode array equipment. Particularly, the study of ON-bipolar cells, but also of photoreceptors, has been hampered by the slow but progressive decline of light responses in the ex vivo ERG over time. Increased perfusion speed and adjustment of the perfusate temperature improve ex vivo retinal function and maximize response amplitude and stability. The ex vivo ERG uniquely allows the study of individual retinal neuronal cell types. In addition, improvements to maximize response amplitudes and stability allow the investigation of light responses in retina samples from large animals, as well as human donor eyes, making the ex vivo ERG a valuable addition to the repertoire of techniques used to investigate retinal function.

Optimizing the Setup and Conditions for Ex Vivo Electroretinogram to Study Retina Function in Small and Large Eyes

Modification of existing multielectrode array or patch clamp equipment makes the ex vivo electroretinogram more widely accessible. Improved methods to record and maintain ex vivo light responses facilitate the study of photoreceptor and ON-bipolar cell function in the healthy retina, animal models of eye diseases, and human donor retinas.

优化离 体 视网膜电图的设置和条件，以研究小眼睛和大眼睛的视网膜功能

Measurements of retinal neuronal light responses are critical to investigating the physiology of the healthy retina, determining pathological changes in ...

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. ...

<meta charset="utf8"></head><body>用于研究视网膜病变的视网膜缺血再灌注损伤小鼠模型

Induction of Retinal Ischemia-Reperfusion Injury in a Mouse Eye Model

Ischemia-reperfusion injuries are known to cause a range of retinal pathologies, including diabetic retinopathy, glaucoma, retinal vascular occlusions, and other vaso-occlusive conditions. This manuscript presents a method for inducing ischemia-reperfusion injury in a mouse model. The method utilized anterior chamber cannulation attached to a saline reservoir, generating hydrostatic pressure to raise the intraocular pressure to 90-100 mmHg. This method effectively caused constriction of retinal capillaries to induce retinal ischemia. At the end of the ischemic period (60 min), the intraocular pressure was normalized (&#8804;20 mmHg) before removing the cannula from the anterior chamber to initiate reperfusion. Days after the ischemia/reperfusion procedure, the eyes were collected and sectioned for histological staining. The histopathology of the retinal sections was scored by evaluating eight parameters of retinal injury: folds, hemorrhage, deformation, cell loss in the ganglion cell, inner nuclear, outer nuclear, and photoreceptor layers, and damage to retinal pigment epithelial cells. This method provided a reproducible model to study the mechanisms and pathology of retinal ischemia/reperfusion injury. In addition, this model can facilitate the discovery of potential therapeutic targets to treat retinal ischemia/reperfusion injury, advancing the study of retinal pathologies and improving patient outcomes.

<meta charset="utf8"></head><body>小鼠眼模型中视网膜缺血再灌注损伤的诱导

This protocol models retinal ischemia-reperfusion injury in a mouse eye by inducing retinal ischemia via anterior chamber cannulation and intraocular pressure elevation, followed by intraocular pressure normalization to initiate the reperfusion.

小鼠眼模型中视网膜缺血再灌注损伤的诱导

Ischemia-reperfusion injuries are known to cause a range of retinal pathologies, including diabetic retinopathy, glaucoma, retinal vascular occlusions, ...

使用电生理评估功能，在啮齿动物的视网膜和远程肢体缺血预适应的保护作用

摘要

探索更多视频

此视频中的章节

植入和无线电图记录和视觉诱发电位清醒大鼠

直接耦合电雷电图（DC-ERG），用于记录小鼠视网膜颜料皮皮的光唤起电响应

远程肢体缺血预适应:在啮齿动物的保护技术

谷氨酸和缺氧作为离体脊椎动物视网膜应力模型

电图同时记录和麻醉大鼠视觉诱发电位

视网膜缺血再灌注损伤的小鼠模型通过眼压升高

优化离体视网膜电图的设置和条件，以研究小眼睛和大眼睛的视网膜功能

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

小鼠眼模型中视网膜缺血再灌注损伤的诱导

小鼠眼模型中视网膜缺血再灌注损伤的诱导

使用电生理评估功能，在啮齿动物的视网膜和远程肢体缺血预适应的保护作用

摘要

探索更多视频

此视频中的章节

植入和无线电图记录和视觉诱发电位清醒大鼠

直接耦合电雷电图（DC-ERG），用于记录小鼠视网膜颜料皮皮的光唤起电响应

远程肢体缺血预适应:在啮齿动物的保护技术

谷氨酸和缺氧作为离体脊椎动物视网膜应力模型

电图同时记录和麻醉大鼠视觉诱发电位

视网膜缺血再灌注损伤的小鼠模型通过眼压升高

优化离 体 视网膜电图的设置和条件，以研究小眼睛和大眼睛的视网膜功能

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

小鼠眼模型中视网膜缺血再灌注损伤的诱导

小鼠眼模型中视网膜缺血再灌注损伤的诱导

优化离体视网膜电图的设置和条件，以研究小眼睛和大眼睛的视网膜功能