This research was funded by Washington College's Provost's Office, Middendorf Fund, Hodson Trust, and Franklin and Marshall's Hackman Fund and College of Grants. We thank E. Counihan, S. Giordano, F. Rauh, and A. Roth for assistance in the field. We thank M. Conner, R. Fleegle, and D. Startt at Chesapeake Farms, and Chino Farms for permission and access. The Washington College GIS Program helped with the preparation of maps.

所有涉及动物对象的程序是由富兰克林和马歇尔和华盛顿学院的机构动物护理和使用委员会批准，并遵循所有当地，州和联邦法规。 1.捕获和处理<ol><li>放置箍陷阱的该已知含有龟水目标体。确定水深确保4 - 陷阱5英寸仍高于水，让被困的海龟浮出水面呼吸。可以肯定，扩大和保护圈陷阱，其最大长度与横梁，防止陷阱崩溃。桩陷阱入水体的床，以防止漂移。 </li><li>诱饵的陷阱与一些死鱼，鸡肝，或鸡脖子，猫粮罐头，和/或蔬菜25即可。 </li><li>检查陷阱，每天两次，并删除海龟。当从陷阱取出乌龟，由边抱动，谨慎使用，以避免爪子或喙伤害。 <ol><li>副渔获物的释放。回覆若继续俘获需要设置陷阱。评估诱饵的状态。如果诱饵已被消耗，增加更多。拉陷阱，如果附加诱捕不希望到岸边。 </li></ol></li><li>如果需要26，27决定龟的性别和年龄。 </li><li>将龟在保持袋，并测量体重用弹簧秤到最近的克</li><li>运输龟在气候受控运输箱在实验室用的少量水。房子单独的动物中，与未处理池塘水水族馆在25℃左右举行，深足只盖住头部。 </li></ol> 2.粘贴无线发射机<ol><li>为了最大限度地提高发射机寿命和输出，选择最大的无线电发射机可能不超过该动物的体重28的5％。 </li><li>大约一半确定发射器放置的位置上背甲背甲中线和横向边缘之间大约升1/3从背甲的后缘ength起来。 </li><li>用干布去除淤泥，杂物和藻类生长准备区。用70％异丙醇拭区。 </li><li>用5分钟环氧树脂少量附加发射机。定向发射机以最大化与甲壳的表面接触。使得它落后平行于主体的长轴动物后面的天线的位置。 </li><li>一旦适当地定位，覆盖整个发射器，并用5分钟环氧约为1厘米甲壳周围的表面。 </li><li>返回龟水族馆，让环氧树脂固化过夜。 </li></ol> 3.药理准备注意:氢溴酸东莨菪碱和东莨菪碱甲基溴是强效的乙酰胆碱受体拮抗剂。当使用这些药物的工作，请查阅材料安全数据表，使用适当的个人防护装备（ 如手套，通风橱），并遵守实验室的安全协议，以避免accidenTAL接触。 <ol><li>使用无菌和antipyrogenic用品，化合物氢溴酸东莨菪碱的储备溶液。称出药物的所需量在分析天平上。混合用注射盐水的东莨菪碱在锥形管的1毫克/毫升的浓度。涡旋溶液直至溶解。确保基的化学品是化学纯度符合或超过美国药典（USP）制剂时可能11。 </li><li>重复步骤3.1与东莨菪碱甲基溴化物。 </li><li>通过0.22微米孔径的尼龙或混合纤维素酯处理溶液注射器过滤器进入无菌密封的血清小瓶中。 </li><li>室温保存。 24小时内使用。 </li></ol> 4.追踪海龟运动使用无线电遥测11，12 <ol><li>移动到目标动物的大致位置，从动物其余至少25米使用定向八木天线和设置在中等接收机增益，扫描卧式上，以确定动物的粗方向和位置。当接收干扰或弱信号，找到了新的位置。增加抬高或持有八木高空时，可以提高信号。 </li><li>一旦一个合适的位置被发现采取轴承，记录与GPS你的立场。 </li><li>使用空/峰值方法28，确定左和右的空值的轴承。 <ol><li>识别最强信号的方向。转动增益向下尽可能同时仍接收可检测信号。在接收器上使用衰减开关，如果这样装备。移动天线向左侧并记录在该信号丢失的罗盘方位。 </li><li>重复正确的轴承上一步。 </li></ol></li><li>重复步骤4.3从同一动物的至少两个附加位置。 注:这些附加分应该采取这样的方式，以包围所述动物。 <ol><li>当一组轴承是从格雷亚取ŧ距离或不可避免的干扰，至少需要两个额外的轴承，以提高准确性。收集套用于尽可能快地估算一个位置，特别是如果动物移动的轴承。 注:此外，多个人员可以协调就可以同时在同一动物的多个独立的轴承。 </li></ol></li><li>使用上述步骤15分钟 - 数字或手工每10动物的记录位置。 </li><li>认知过程的操纵<ol><li>一旦未经处理动物的路径记录（一个主体内的控制），提供了一个既剂量的东莨菪碱或甲基东的。使用在步骤1.5中收集的质量，计算药物的量给予动物以达到6.4毫克/千克东莨菪碱或6.8毫克/千克甲基东19,20的一个特定的爬行动物的最终剂量。 </li><li> 通过使用1.0M的尾腹膜窦直接递送药物进入腹膜升注射器用22号针头。确保输送的总体积不超过1毫升</li><li>在捕获其网站上尽快释放动物。 </li><li>继续监测动物的运动，每10 - 15分钟，直到它到达其预期目的地。 </li></ol></li></ol> 5.空间分析<ol><li>计算动物的位置的估计。 <ol><li>对于每一组的空轴承，平分角（用手或通过软件），以找到所得发射机轴承。重复在给定时间点的所有轴承。 </li><li>根据制造商的协议使用三角测量软件，估计动物的与多个发射机的轴承位置和相关联的错误。转换位置估计为x / y坐标，如果软件提供不同的输出。 </li></ol></li><li>重复步骤5.1轴承所有集合。 </li><li>计算运动的空间精度。 <ol><li>计算累积的几何平均值未经处理的动物（阴性对照），因为他们对自己的目标12移动（ 即平均路径）。 </li><li>用于治疗和阳性对照组，每组个体，计算直到所有的点的100％已列入11,12重叠来自5个米的间隔的几何平均线依次较大幅点的累积数目。 </li><li>计算的平均值和标准误差在每个组中的所有个体的每个分割行的距离。 </li><li>统计分析的数据时，治疗组的时间间隔中任一用于比较在给定的时间间隔，以容纳点或重叠点的100％，这取决于问题。 </li></ol></li></ol>

<ol>
	<li>Shettleworth, S. 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> Cognition, Evolution and Behavior. , (2010).</li><li>Bingman, V. P., Cheng, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanisms+of+animal+global+navigation:+comparative+perspectives+and+enduring+challenges.">Mechanisms of animal global navigation: comparative perspectives and enduring challenges.</a> Ethol. Ecol. Evol. 17, 295-318 (2005).</li><li>Mueller, T., O'Hara, R. B., Converse, S. J., Urbanek, R. P., Fagan, W. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Social+Learning+of+Migratory+Performance.">Social Learning of Migratory Performance.</a> Science. 341, 999-1002 (2013).</li><li>Putman, N. F., 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=An+inherited+magnetic+map+guides+ocean+navigation+in+juvenile+Pacific+salmon.">An inherited magnetic map guides ocean navigation in juvenile Pacific salmon.</a> Curr. Biol. 24, 446-450 (2014).</li><li>Lohmann, K. J., Putman, N. F., Lohmann, C. M. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Geomagnetic+imprinting:+a+unifying+hypothesis+of+natal+homing+in+salmon+and+sea+turtles.">Geomagnetic imprinting: a unifying hypothesis of natal homing in salmon and sea turtles.</a> Proc. Natl. Acad. Sci., USA. 105, 19096-19101 (2008).</li><li>Fuxjager, M. J., Davidoff, K. R., Mangiamele, L. A., Lohmann, K. 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+geomagnetic+environment+in+which+sea+turtle+eggs+incubate+affects+subsequent+magnetic+navigation+behaviour+of+hatchlings.">The geomagnetic environment in which sea turtle eggs incubate affects subsequent magnetic navigation behaviour of hatchlings.</a> Proc. R. Soc. B. 281, 1218-1225 (2014).</li><li>Shettleworth, S. 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+evolution+of+comparative+cognition:+is+the+snark+still+a+boojum.">The evolution of comparative cognition: is the snark still a boojum.</a> Behav. Processes. 80, 210-217 (2009).</li><li>Fagan, W. F., 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=Spatial+memory+and+animal+movement.">Spatial memory and animal movement.</a> Ecol. Lett. 16, 1316-1329 (2013).</li><li>Collett, T. S., Graham, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Insect+Navigation:+Do+Honeybees+Learn+to+Follow+Highways.">Insect Navigation: Do Honeybees Learn to Follow Highways.</a> Curr. Biol. 25, 240-242 (2015).</li><li>Menzel, R., 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=Honey+bees+navigate+according+to+a+map-like+spatial+memory.">Honey bees navigate according to a map-like spatial memory.</a> Proc. Natl. Acad. Sci., USA. 102, 3040-3045 (2005).</li><li>Roth, T. C., Krochmal, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pharmacological+Evidence+is+Consistent+with+a+Prominent+Role+of+Spatial+Memory+in+Complex+Navigation.">Pharmacological Evidence is Consistent with a Prominent Role of Spatial Memory in Complex Navigation.</a> Proc. R. Soc. B. 283, 20152548 (2016).</li><li>Roth, T. C., Krochmal, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+role+of+age-specific+learning+and+experience+for+turtles+navigating+a+changing+landscape.">The role of age-specific learning and experience for turtles navigating a changing landscape.</a> Curr. Biol. 25, 333-337 (2015).</li><li>Krochmal, A. R., Roth, T. C., Rush, S., Wachter, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Turtles+outsmart+rapid+environmental+change:+the+role+of+cognition+in+navigation.">Turtles outsmart rapid environmental change: the role of cognition in navigation.</a> Comm. Integr. Biol. , (2015).</li><li>Thorup, 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=Evidence+for+a+navigational+map+stretching+across+the+continental+U.S.+in+a+migratory+songbird.">Evidence for a navigational map stretching across the continental U.S. in a migratory songbird.</a> Proc. Natl. Acad. Sci., USA. 104, 18115-18119 (2007).</li><li>Lohmann, K. J., Lohmann, C. M. F., Putman, N. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Magnetic+maps+in+animals:+nature's+GPS.">Magnetic maps in animals: nature's GPS.</a> J. Exp. Biol. 210, 3697-3705 (2007).</li><li>Collett, M., Chittka, L., Collett, T. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spatial+Memory+in+Insect+Navigation.">Spatial Memory in Insect Navigation.</a> Curr. Biol. 23, 789-800 (2013).</li><li>Foden, W., Vi&#233;, J. C., Hilton-Taylor, C., Stuart, S. N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Species+susceptibility+to+climate+change+impacts.">Species susceptibility to climate change impacts.</a> The 2008 Review of The IUCN Red List of Threatened Species. , (2008).</li><li>Kohler, E. C., Riters, L. V., Chaves, L., Bingman, V. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Muscarinic+Acetylcholine+Antagonist+Scopolamine+Impairs+Short-Distance+Homing+Pigeon+Navigation.">The Muscarinic Acetylcholine Antagonist Scopolamine Impairs Short-Distance Homing Pigeon Navigation.</a> Physiol. Behav. 60, 1057-1061 (1996).</li><li>Powers, A. S., Hogue, P., Lynch, C., Gattuso, B., Lissek, S., Nayal, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Role+of+Acetylcholine+in+negative+patterning+in+turtles+(Chrysemys+picta).">Role of Acetylcholine in negative patterning in turtles (Chrysemys picta).</a> Behav. Neurosci. 123, 804-809 (2009).</li><li>Petrillo, M., Ritter, C. A., Powers, A. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+role+for+Acetylcholine+in+spatial+memory+in+turtles.">A role for Acetylcholine in spatial memory in turtles.</a> Physiol. Behav. 56, 135-141 (1994).</li><li>Klinkenberg, I., Blokland, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+validity+of+scopolamine+as+a+pharmacological+model+for+cognitive+impairment:+A+review+of+animal+behavioral+studies.">The validity of scopolamine as a pharmacological model for cognitive impairment: A review of animal behavioral studies.</a> Neurosci. Biobehav. Rev. 34, 1307-1350 (2010).</li><li>Pradhan, S. N., Roth, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparative+behavioral+effects+of+several+anticholinergic+agents+in+rats.">Comparative behavioral effects of several anticholinergic agents in rats.</a> Psychopharm. (Berlin). 12, 358-366 (1968).</li><li>Harvey, J. A., Gormezano, I., Cool-Hauser, V. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+scopolamine+and+methylscopolamine+on+classical+conditioning+of+the+rabbit+nictitating+membrane+response.">Effects of scopolamine and methylscopolamine on classical conditioning of the rabbit nictitating membrane response.</a> J. Pharmacol. Exp. Therap. 225, 42-49 (1983).</li><li>Evans, H. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Scopolamine+effects+on+visual+discrimination:+modifications+related+to+stimulus+control.">Scopolamine effects on visual discrimination: modifications related to stimulus control.</a> J. Pharmacol. Exp. Therap. 195, 105-113 (1975).</li><li>Dreslik, M. J., Phillips, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Turtle+communities+in+the+upper+midwest,+USA.">Turtle communities in the upper midwest, USA.</a> J. Freshwater Ecol. 20, 149-164 (2005).</li><li>Sexton, O. 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+method+of+estimating+the+age+of+painted+turtles+for+use+in+demographic+studies.">A method of estimating the age of painted turtles for use in demographic studies.</a> Ecology. 40, 716-718 (1959).</li><li>Wilson, D. S., Tracy, C. R., Tracy, C. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Estimating+age+of+turtles+from+growth+rings:+a+critical+evaluation+of+the+technique.">Estimating age of turtles from growth rings: a critical evaluation of the technique.</a> Herpetologica. 59, 178-194 (2003).</li><li>Kenward, R. E. <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 Manual for Wildlife Radio Tagging. , (2000).</li><li>Jones, D. N. C., Higgins, G. A. <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+scopolamine+on+visual+attention+in+rats.">Effect of scopolamine on visual attention in rats.</a> Psychopharm. 120, 142-149 (1995).</li><li>Araujo, J. A., Nobrega, J. N., Raymond, R., Milgram, N. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Aged+dogs+demonstrate+both+increased+sensitivity+to+scopolamine+impairment+and+decreased+muscarinic+receptor+density.">Aged dogs demonstrate both increased sensitivity to scopolamine impairment and decreased muscarinic receptor density.</a> Pharmacol. Biochem. Behav. 98, 203-209 (2011).</li><li>Greggor, A. L., Clayton, N. S., Phalan, B., Thornton, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparative+cognition+for+conservationists.">Comparative cognition for conservationists.</a> Trends Ecol. Evol. 29, 489-495 (2014).</li><li>Roth, T. C., Krochmal, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cognition-centered+conservation+as+a+means+of+advancing+integrative+animal+behavior.">Cognition-centered conservation as a means of advancing integrative animal behavior.</a> Curr. Opinion Behav. Sci. 6, 1-6 (2015).</li><li>LaDage, L. D., Roth, T. C., Cerjanic, A. C., Sinervo, B., Pravosudov, V. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spatial+memory:+Are+lizards+really+deficient.">Spatial memory: Are lizards really deficient.</a> Biol. Lett. 8, 939-941 (2012).</li></ol>

The authors have nothing to disclose.

这里介绍的协议允许实验者记录和量化在导航认知的作用。在现场操作的认知已经证明是困难的，因为多数的方法发表实验者无法知道哪些动物的行为的具体方面是被操纵。然而，这里提出的协议允许实验者准确操纵，从而评估在导航认知的作用。该技术还允许实验者监测实时动物导航与异常高的空间和时间分辨率，由此赋予研究者清楚地记录在野生动物的认知实验操作的行为后果。 <p class="j...

认知（本文中定义为"涉及采集，存储，并使用从环境信息的所有进程"1）是中央的复杂的导航任务2的阵列。例如，沙丘鹤（ 丹顶鹤加拿大一枝黄花 ），显示精度迁徙对他们的本命的海滩幼体有明显改善与经验; 3，和海龟物种的印记，并返回作为成年人4-6。同样，成功的迁移，扩散和觅食的动物的能力，铰链，以收集有关其空间环境7,8的信息。有些动物出现学习就具体的景观特色9航行路线和筑巢和觅食区10之间移动时，可以使用空间认知。在东部锦龟最近的工作（Chrysemys PICTA）建议在航海，其中成人高地栖息地的成功导航尤文铰链的关键时期一个狭窄的年龄范围（&lt;4岁11-13）内Nile体验。虽然这些一起研究表明，在了解航海4-6，14-16学习的作用所取得的进展，背后这些行为的机制和认知导航的充分作用仍然是谜一般的，尤其是在脊椎动物8，17 18。 实地调查认知导航的作用是罕见的2，8，18，这主要是由于参与监督的方法上的困难，处理和跟踪野生动物。例如，大空间和时间尺度上许多动物进行导航往往妨碍调查两种类型的信息，这些动物可能学习和信息是如何获得的。实验者经常面对的检测和监控行为在如此大面积和时间框架时定位的动物，从而限制了类型的后勤困难的，可以被收集的数据，并且可以得出的结论。虽然使用动物安装全球定位系统（GPS）的记录器可以提高检测广泛范围的动物的概率，通过这些装置收集的空间数据一般都非常粗分辨率的和缺乏详细的行为的组件。因此，能在这种情况下要收集的数据是用于在行为检查不同的组或实验组之间微妙的变化价值有限。同样地，目标行为的直接受控操纵通常是由典型的导航行为，以及由实地研究的固有后勤限制的空间和时间尺度禁止。发现动物的自然栖息地，捕捉和操纵它们，然后收集行为数据，而无需在不经意间产生的寄生行为在外地工作动物的重大挑战。因此，对FR实验设计EE-不等的动物往往是制约和对认知的导航作用进行严格，控制现场实验的能力是有限的。 本研究绕过许多用在野外条件下的新的药理学处理和自由导航动物高分辨率跟踪的组合调查在该领域认知和导航之间的关系的先前困难。东莨菪碱，毒蕈碱乙酰胆碱受体（mAchR的）拮抗剂，已显示在多种脊椎动物类群18-24的脑中阻断胆碱能活性阻断空间记忆形成和召回。东莨菪碱能有效地对自由放养动物的野外条件下11，18下使用并有标记，但暂时的效果（ 例如 ，6 -爬行动物8小时）。甲基东，即不穿过血脑屏障19-21一个mAchR的拮抗剂，可用于控制东莨菪碱和行为11的非认知方面的可能外周作用。药理学允许认知的精确操纵通过直接影响受体，和高精度的无线电遥测允许的对行为所得效果的观察。 通过远程三角同时具有高的空间（±2.5米）和时间（15分钟）的分辨率进行的测量允许精确记录和相对动物行为的量化，以认知的实验操作。 这项研究11切萨皮克农场，在肯特公司，MD一个3300英亩野生动物管理和农业研究领域，美国（39.194°N，76.187°W）五月和2013年8月和2014年间进行的。该协议包括五个主要步骤:（1）捕捉和处理的动物（2）固定用无线电发射机（3）制备的药理学试剂（4）监测和操纵动物的动作，和（5）ANAlyzing空间数据。本文所述集中在东部的研究锦龟（Chrysemys PICTA）。在重点人群海龟参与其中，他们离开自己的家的池塘，然后导航到使用四种非常精确的（±3.5米）的，复杂的，高度可预测的路线11，12替代水生境的年度陆上运动。在动物药理操纵该系统具有高分辨率无线电遥测配对揭示认知的自由航行野生动物的角色光芒。

<table><tbody><tr><td>Scopolamine bromide</td><td>Sigma</td><td>S0929</td><td>USP</td></tr><tr><td>Scopolamine methylbromide</td><td>Sigma</td><td>S8502, 1421009</td><td>USP and non USP versions</td></tr><tr><td>Saline</td><td>Hanna Pharmaceutical Supply Co., Inc.</td><td>409488850</td><td>USP, formulated as an injectable&amp;nbsp;</td></tr><tr><td>Syringe filter</td><td>Fisher</td><td>09-720-004</td><td></td></tr><tr><td>Syringe</td><td>Fisher</td><td>14-823-30</td><td></td></tr><tr><td>Hypodermic needle</td><td>Fisher</td><td>14-823-13</td><td></td></tr><tr><td>Antenna</td><td>Wildlife Materials</td><td>3 Element Folding Yagi</td><td>Antennae with additional elements are available, but can be cumbersome in the field.&amp;nbsp;</td></tr><tr><td>Radio Receiver</td><td>Wildlife Materials</td><td>TRX-2000S</td><td>Water resistant models are also available.</td></tr><tr><td>Compass</td><td>Brunton&amp;nbsp;</td><td>Truarc 15</td><td></td></tr><tr><td>Radio transmitters</td><td>Holohil Inc.</td><td>BD-2, PD-2, RI-2B</td><td>Transmitter models vary in lifespan and signal output as a function of battery size and pulse rate settings, which can be customized based on the study question and organism.</td></tr><tr><td>GPS</td><td>Garmin</td><td>eTrex Venture</td><td></td></tr><tr><td>Coaxial cable</td><td>newegg.com</td><td>C2G 40026</td><td>BNC connections are necessary.</td></tr><tr><td>Hoop net</td><td>Memphis Net and Twine&amp;nbsp;</td><td>TN325</td><td>Net mesh size should be chosen based on the minimum size of the target animal.&amp;nbsp;</td></tr></tbody></table>

using pharmacological manipulation and high-precision radio telemetry to study the spatial cognition in free-ranging animals

动物的感知和了解其环境的能力起着许多行为过程，包括导航，迁移，扩散和觅食了关键作用。然而，认知的导航策略和这些策略背后的机制发展中的作用的认识是通过参与监督的方法上的困难限制，操纵的认知和跟踪野生动物。这项研究描述为解决认知导航，它结合了高精度无线电遥测行为药理操纵角色的协议。该方法使用东莨菪碱，毒蕈碱乙酰胆碱受体拮抗剂，操纵认知空间能力。处理的动物中，然后用高频率和通过远程三角高空间分辨率监测。该协议是一个人口的东锦龟（Chrysemys PICTA）内的应用已经有人居住季节性短暂的水源〜100年间，采用精确的（±3.5米）远源之间移动，复杂的（ 即具有较高的曲折经过多个栖息地非线性），且可预测的路线前4岁学会了。这项研究表明，由这些龟所使用的过程是与空间记忆形成和召回一致。总之，这些结果与空间认知的复杂的导航作用是一致的，突出的生态和药理学技术认知和导航的研究的整合。

采用上述方案，认知导航的作用，东部人口进行评估锦龟（Chrysemys PICTA）已经经历了〜百年的季节性短暂的水源。这一人群栖息的短暂的混合（每年并迅速排出-在几个小时）和永久水生生境（ 图1）。先前的研究表明，它们的池塘排水后，居民龟导航到以高精度（±3.5米）在使用前4岁11-13（ 图1）了解到复杂的，可预测的路线的替代...

Watch this Scientific Journal Video about Using Pharmacological Manipulation and High-precision Radio Telemetry to Study the Spatial Cognition in Free-ranging Animals at JoVE.com

Using Pharmacological Manipulation and High-precision Radio Telemetry to Study the Spatial Cognition in Free-ranging Animals

本文介绍了结合内存和无线电遥测的药理操作记录和量化导航认知的作用新颖的协议。

使用药理操纵和高精度无线电遥测研究空间认知中自由放养的动物

An animal&#39;s ability to perceive and learn about its environment plays a key role in many behavioral processes, including navigation, migration, ...

using-pharmacological-manipulation-high-precision-radio-telemetry-to

Universidad San Sebastian

Research

JoVE Journal

Environment

6.7K 次观看.  Franklin and Marshall College. 本文介绍了结合内存和无线电遥测的药理操作记录和量化导航认知的作用新颖的协议。 

视频: Using Pharmacological Manipulation and High-precision Radio Telemetry to Study the Spatial Cognition in Free-ranging Animals

Flat-floored Air-lifted Platform: A New Method for Combining Behavior with Microscopy or Electrophysiology on Awake Freely Moving Rodents

It is widely acknowledged that the use of general anesthetics can undermine the relevance of electrophysiological or microscopical data obtained from a living animal&#8217;s brain. Moreover, the lengthy recovery from anesthesia limits the frequency of repeated recording/imaging episodes in longitudinal studies. Hence, new methods that would allow stable recordings from non-anesthetized behaving mice are expected to advance the fields of cellular and cognitive neurosciences. Existing solutions range from mere physical restraint to more sophisticated approaches, such as linear and spherical treadmills used in combination with computer-generated virtual reality. Here, a novel method is described where a head-fixed mouse can move around an air-lifted mobile homecage and explore its environment under stress-free conditions. This method allows researchers to perform behavioral tests (e.g., learning, habituation or novel object recognition) simultaneously with two-photon microscopic imaging and/or patch-clamp recordings, all combined in a single experiment. This video-article describes the use of the awake animal head fixation device (mobile homecage), demonstrates the procedures of animal habituation, and exemplifies a number of possible applications of the method.

This method creates a tangible, familiar environment for the mouse to navigate and explore during microscopic imaging or single-cell electrophysiological recordings, which require firm fixation of the animal&#8217;s head.

平地板的空气解禁平台:一种新方法与显微镜或电对清醒结合行为自由移动鼠害

It is widely acknowledged that the use of general anesthetics can undermine the relevance of electrophysiological or microscopical data obtained from a ...

科研

行为学

A Method for Remotely Silencing Neural Activity in Rodents During Discrete Phases of Learning

This protocol describes how to temporarily and remotely silence neuronal activity in discrete brain regions while animals are engaged in learning and memory tasks. The approach combines pharmacogenetics (Designer-Receptors-Exclusively-Activated-by-Designer-Drugs) with a behavioral paradigm (sensory preconditioning) that is designed to distinguish between different forms of learning. Specifically, viral-mediated delivery is used to express a genetically modified inhibitory G-protein coupled receptor (the Designer Receptor) into a discrete brain region in the rodent. Three weeks later, when designer receptor expression levels are high, a pharmacological agent (the Designer Drug) is administered systemically 30 min prior to a specific behavioral session. The drug has affinity for the designer receptor and thus results in inhibition of neurons that express the designer receptor, but is otherwise biologically inert. The brain region remains silenced for 2-5 hr (depending on the dose and route of administration). Upon completion of the behavioral paradigm, brain tissue is assessed for correct placement and receptor expression. This approach is particularly useful for determining the contribution of individual brain regions to specific components of behavior and can be used across any number of behavioral paradigms.

This protocol describes how to temporarily and remotely silence neuronal activity in discrete brain regions while rats are engaged in learning and memory tasks. The approach combines pharmacogenetics (Designer-Receptors-Exclusively-Activated-by-Designer-Drugs) with a behavioral paradigm (sensory preconditioning) that is designed to distinguish between different components of learning.

为远程沉默的神经活动在鼠害的方法在学习独立的阶段

This protocol describes how to temporarily and remotely silence neuronal activity in discrete brain regions while animals are engaged in learning and ...

Implantation of Radiotelemetry Transmitters Yielding Data on ECG, Heart Rate, Core Body Temperature and Activity in Free-moving Laboratory Mice

The laboratory mouse is the animal species of choice for most biomedical research, in both the academic sphere and the pharmaceutical industry. Mice are a manageable size and relatively easy to house. These factors, together with the availability of a wealth of spontaneous and experimentally induced mutants, make laboratory mice ideally suited to a wide variety of research areas.
In cardiovascular, pharmacological and toxicological research, accurate measurement of parameters relating to the circulatory system of laboratory animals is often required. Determination of heart rate, heart rate variability, and duration of PQ and QT intervals are based on electrocardiogram (ECG) recordings. However, obtaining reliable ECG curves as well as physiological data such as core body temperature in mice can be difficult using conventional measurement techniques, which require connecting sensors and lead wires to a restrained, tethered, or even anaesthetized animal. Data obtained in this fashion must be interpreted with caution, as it is well known that restraining and anesthesia can have a major artifactual influence on physiological parameters1, 2.
Radiotelemetry enables data to be collected from conscious and untethered animals. Measurements can be conducted even in freely moving animals, and without requiring the investigator to be in the proximity of the animal. Thus, known sources of artifacts are avoided, and accurate and reliable measurements are assured. This methodology also reduces interanimal variability, thus reducing the number of animals used, rendering this technology the most humane method of monitoring physiological parameters in laboratory animals3, 4. Constant advancements in data acquisition technology and implant miniaturization mean that it is now possible to record physiological parameters and locomotor activity continuously and in realtime over longer periods such as hours, days or even weeks3, 5.
Here, we describe a surgical technique for implantation of a commercially available telemetry transmitter used for continuous measurements of core body temperature, locomotor activity and biopotential (i.e. onelead ECG), from which heart rate, heart rate variability, and PQ and QT intervals can be established in freeroaming, untethered mice. We also present pre-operative procedures and protocols for post-operative intensive care and pain treatment that improve recovery, well-being and survival rates in implanted mice5, 6.

A surgical technique for implantation of commercially available telemetry transmitters used for continuous measurement of biopotential (one-lead ECG), heart rate, core body temperature and locomotor activity in freely moving mice is shown. Recommendations and protocols for post-operative care and pain relief, improving recovery, well being and survival rate are also presented.

植入的无线电遥测发射器产生心电图，心率，身体核心温度和自由移动的实验室小鼠的活动上的数据

The laboratory mouse is the animal species of choice for most biomedical research, in both the academic sphere and the pharmaceutical industry. Mice are a ...

医学

Survivable Stereotaxic Surgery in Rodents 

The ability to measure extracellular basal levels of neurotransmitters in the brain of awake animals allows for the determination of effects of different systemic challenges (pharmacological or physiological) to the CNS. For example, one can directly measure how the animal's midbrain dopamine projections respond to dopamine-releasing drugs like d-amphetamine or natural stimuli like food. In this video, we show you how to implant guide cannulas targeting specific sites in the rat brain, how to insert and implant a microdialysis probe and how to use high performance liquid chromatography coupled with electrochemical detection (HPLC-EC) to measure extracellular levels of oxidizable neurotransmitters and metabolites. Local precise introduction of drugs through the microdialysis probe allows for refined work on site specificity in a compound s mechanism of action. This technique has excellent anatomical and chemical resolution but only modest time resolution as microdialysis samples are usually processed every 20-30 minutes to ensure detectable neurotransmitter levels. Complementary ex vivo tools (i.e., slice and cell culture electrophysiology) can assist with monitoring real-time neurotransmission.

Survivable Stereotaxic Surgery in Rodents

The monitoring of extracellular neurotransmitter levels in distinct brain regions of freely moving animals offers insights on the link between neurotransmitter release and behavior. In vivo microdialysis coupled with electrochemical detection provides excellent anatomical and chemical resolution; and information on how basal neurotransmission is altered by pharmacological or physiological manipulations.

在啮齿类动物的存活立体定向手术

The ability to measure extracellular basal levels of neurotransmitters in the brain of awake animals allows for the determination of effects of different ...

生物学

A Wireless, Bidirectional Interface for In Vivo Recording and Stimulation of Neural Activity in Freely Behaving Rats

In vivo electrophysiology is a powerful technique to investigate the relationship between brain activity and behavior at a millisecond and micrometer scale. However, current methods mostly rely on tethered cable recordings or only use unidirectional systems, allowing either recording or stimulation of neural activity, but not at the same time or same target. Here, a new wireless, bidirectional device for simultaneous multichannel recording and stimulation of neural activity in freely behaving rats is described. The system operates through a single portable head stage that both transmits recorded activity and can be targeted in real-time for brain stimulation using a telemetry-based multichannel software. The head stage is equipped with a preamplifier and a rechargeable battery, allowing stable long-term recordings or stimulation for up to 1 h. Importantly, the head stage is compact, weighs 12 g (including battery) and thus has minimal impact on the animal&#180;s behavioral repertoire, making the method applicable to a broad set of behavioral tasks. Moreover, the method has the major advantage that the effect of brain stimulation on neural activity and behavior can be measured simultaneously, providing a tool to assess the causal relationships between specific brain activation patterns and behavior. This feature makes the method particularly valuable for the field of deep brain stimulation, allowing precise assessment, monitoring, and adjustment of stimulation parameters during long-term behavioral experiments. The applicability of the system has been validated using the inferior colliculus as a model structure.

A Wireless, Bidirectional Interface for In Vivo Recording and Stimulation of Neural Activity in Freely Behaving Rats

A wireless, bidirectional system for multi-channel neural recordings and stimulation in freely behaving rats is introduced. The system is light and compact, thus having minimal impact on the animal&#180;s behavioral repertoire. Moreover, this bidirectional system provides a sophisticated tool to assess causal relationships between brain activation patterns and behavior.

用于体内的无线双向接口记录和刺激自由行为大鼠的神经活动

In vivo electrophysiology is a powerful technique to investigate the relationship between brain activity and behavior at a millisecond and micrometer ...

Automated, Long-term Behavioral Assay for Cognitive Functions in Multiple Genetic Models of Alzheimer's Disease, Using IntelliCage

Multiple factors&#8212;such as aging and genes&#8212;are frequently associated with cognitive decline. Genetically modified mouse models of cognitive decline, such as Alzheimer&#39;s disease (AD), have become a promising tool to elucidate the underlying mechanisms and promote the therapeutic advances. An important step is the validation and characterization of expected behavioral abnormality in the models, in the case of AD, cognitive decline. The long-term behavioral investigations of laboratory animals to study the effect of aging demand substantial efforts from researchers. The IntelliCage system is a high-throughput and cost-effective test battery for mice that eliminates the need for daily human handling. Here, we describe how the system is utilized in the long-term phenotyping of a genetic Alzheimer&#39;s disease model, specifically focusing on the cognitive functions. The experiment employs repeated battery of tests that assess spatial learning and executive functions. This cost-effective age-dependent phenotyping allows us to identify the transient and/or permanent effects of genes on various cognitive aspects.

This paper describes a protocol for cognitive assessments for genetic models of the Alzheimer&#39;s disease using the IntelliCage system, which is a high throughput automated behavioral monitoring system with operant conditioning.

基于 IntelliCage 的阿尔茨海默病多基因模型认知功能的自动、长期行为检测

Multiple factors&#8212;such as aging and genes&#8212;are frequently associated with cognitive decline. Genetically modified mouse models of cognitive ...

Optogenetic Manipulation of Neuronal Activity to Modulate Behavior in Freely Moving Mice

Optogenetic modulation of neuronal circuits in freely moving mice affects acute and long-term behavior. This method is able to perform manipulations of single neurons and region-specific transmitter release, up to whole neuronal circuitries in the central nervous system, and allows the direct measurement of behavioral outcomes. Neurons express optogenetic tools via an injection of viral vectors carrying the DNA of choice, such as Channelrhodopsin2 (ChR2). Light is brought into specific brain regions via chronic optical implants that terminate directly above the target region. After two weeks of recovery and proper tool-expression, mice can be repeatedly used for behavioral tests with optogenetic stimulation of the neurons of interest.
Optogenetic modulation has a high temporal and spatial resolution that can be accomplished with high cell specificity, compared to the commonly used methods such as chemical or electrical stimulation. The light does not harm neuronal tissue and can therefore be used for long-term experiments as well as for multiple behavioral experiments in one mouse. The possibilities of optogenetic tools are nearly unlimited and enable the activation or silencing of whole neurons, or even the manipulation of a specific receptor type by light.
The results of such behavioral experiments with integrated optogenetic stimulation directly visualizes changes in behavior caused by the manipulation. The behavior of the same animal without light stimulation as a baseline is a good control for induced changes. This allows a detailed overview of neuronal types or neurotransmitter systems involved in specific behaviors, such as anxiety. The plasticity of neuronal networks can also be investigated in great detail through long-term stimulation or behavioral observations after optical stimulation. Optogenetics will help to enlighten neuronal signaling in several kinds of neurological diseases.

With optogenetic manipulation of specific neuronal populations or brain regions, behavior can be modified with high temporal and spatial resolution in freely moving animals. By using different optogenetic tools in combination with chronically implanted optical fibers, a variety of neuronal modulations and behavioral testing can be performed.

神经活性的光遗传学操作，调节自由移动小鼠的行为

Optogenetic modulation of neuronal circuits in freely moving mice affects acute and long-term behavior. This method is able to perform manipulations of ...

Simultaneous Monitoring of Wireless Electrophysiology and Memory Behavioral Test as a Tool to Study Hippocampal Neurogenesis

Brainwaves amplitude obtained from electroencephalography (EEG) has been well-recognized as a basis for cognitive capacity, memory, and learning on animals and humans. Adult neurogenesis mechanism is also linked to memory and learning improvement. Traditionally, researchers used to assess learning and memory parameters in rodent models by behavioral tasks. Therefore, the simultaneous monitoring of behavioral changes and EEG is particularly interesting in correlating data between brain activity and task-related behaviors. However, most of the equipment required to perform both studies are either complex, expensive, or uses a wired setup network that hinders the natural animals&#8217; movement. In this study, EEG was recorded with a wireless electrophysiology device along with the execution of a novel object recognition task (NORT). The animal&#8217;s behavior was monitored simultaneously by a video tracking system. Both recordings were analyzed offline by their timestamps which were synched to link EEG signals with the animal&#8217;s actions. Subjects consist of adult Wistar rats after medium-term environmental enrichment treatment. Six skull screw electrodes were fixed in pairs on both hemispheres over frontal, central, and parietal regions and were referenced to an electrode located posterior of the nasal bone. NORT protocol consists of exposing the animal to two identical objects for 10 min. After 2 h and 24 h, one of the objects was replaced with a novel one. Exploration time for each object was monitored by a behavioral tracking software (BTS) and EEG data recording. The analysis of the EEG synced with behavioral data consists of estimations of alpha and beta relative band power and comparisons between novel object recognition versus familiar object exploration, between three experimental stages. In this manuscript, we have discussed electrodes manufacturing process, epidural electrodes implantation surgery, environmental enrichment protocol, NORT protocol, BTS setup, EEG &#8211; BTS coupling for simultaneous monitoring in real-time, and EEG data analysis based on automatic events detection.

The protocol presented here provides information on the simultaneous electroencephalography (EEG) and behavioral assessment in real-time. We have discussed all steps involved in this protocol as an attractive solution for researchers in many fields of neuroscience, particularly in learning and memory areas.

同时监测无线电生理学和记忆行为测试作为研究海马神经发生的工具

Brainwaves amplitude obtained from electroencephalography (EEG) has been well-recognized as a basis for cognitive capacity, memory, and learning on ...

Non-restraining EEG Radiotelemetry: Epidural and Deep Intracerebral Stereotaxic EEG Electrode Placement

Implantable EEG radiotelemetry is of central relevance in the neurological characterization of transgenic mouse models of neuropsychiatric and neurodegenerative diseases as well as epilepsies. This powerful technique does not only provide valuable insights into the underlying pathophysiological mechanisms, i.e., the etiopathogenesis of CNS related diseases, it also facilitates the development of new translational, i.e., therapeutic approaches. Whereas competing techniques that make use of recorder systems used in jackets or tethered systems suffer from their unphysiological restraining to semi-restraining character, radiotelemetric EEG recordings overcome these disadvantages. Technically, implantable EEG radiotelemetry allows for precise and highly sensitive measurement of epidural and deep, intracerebral EEGs under various physiological and pathophysiological conditions. First, we present a detailed protocol of a straight forward, successful, quick and efficient technique for epidural (surface) EEG recordings resulting in high-quality electrocorticograms. Second, we demonstrate how to implant deep, intracerebral EEG electrodes, e.g., in the hippocampus (electrohippocampogram). For both approaches, a computerized 3D stereotaxic electrode implantation system is used. The radiofrequency transmitter itself is implanted into a subcutaneous pouch in both mice and rats. Special attention also has to be paid to pre-, peri- and postoperative treatment of the experimental animals. Preoperative preparation of mice and rats, suitable anesthesia as well as postoperative treatment and pain management are described in detail.

Non-restraining EEG radiotelemetry is a valuable methodological approach to record in vivo long-term electroencephalograms from freely moving rodents. This detailed protocol describes stereotaxic epidural and deep intracerebral electrode placement in different brain regions in order to obtain reliable recordings of CNS rhythmicity and CNS related behavioral stages.

非约束脑电遥测:硬膜外和脑深部脑电立体定位放置电极

Implantable EEG radiotelemetry is of central relevance in the neurological characterization of transgenic mouse models of neuropsychiatric and ...

神经科学

Stereotaxic Surgery for Implantation of Microelectrode Arrays in the Common Marmoset (Callithrix jacchus)

Marmosets (Callithrix jacchus) are small non-human primates that are gaining popularity in biomedical and preclinical research, including the neurosciences. Phylogenetically, these animals are much closer to humans than rodents. They also display complex behaviors, including a wide range of vocalizations and social interactions. Here, an effective stereotaxic neurosurgical procedure for implantation of recording electrode arrays in the common marmoset is described. This protocol also details the pre- and postoperative steps of animal care that are required to successfully perform such a surgery. Finally, this protocol shows an example of local field potential and spike activity recordings in a freely behaving marmoset 1 week after the surgical procedure. Overall, this method provides an opportunity to study the brain function in awake and freely behaving marmosets. The same protocol can be readily used by researchers working with other small primates. In addition, it can be easily modified to allow other studies requiring implants, such as stimulating electrodes, microinjections, implantation of optrodes or guide cannulas, or ablation of discrete tissue regions.

Stereotaxic Surgery for Implantation of Microelectrode Arrays in the Common Marmoset Callithrix jacchus

This work presents a protocol to perform a stereotaxic, neurosurgical implantation of&#160;microelectrode arrays in the common marmoset. This method specifically enables electrophysiological recordings in freely behaving animals but can be easily adapted to any other similar neurosurgical intervention in this species (e.g., cannula for drug administration or electrodes for brain stimulation).