We are grateful to anonymous reviewers for comments and suggestions that greatly improved this manuscript. We thank R. Bowes, R. Carreno, and T. Panhuis for assistance in collecting the fish. We also thank M. Lee for assistance with male preference trials. We are grateful to the Ohio Wesleyan University Department of Zoology for helpful advice and suggestions throughout this study and Arizona State University (McGraw Lab) for software advice.

所有实验均与俄亥俄卫斯理大学的机构动物护理和使用委员会的批准进行的。 用分光光度计鱼1.记录反射紫外光<ol><li>校准分光光度计和光源与已知白色标准以上的波长范围，以根据仪器或软件的建议来测定。 </li><li>通过在3-氨基苯甲酸乙酯甲磺酸盐的0.5％溶液放置麻醉鱼（MS-222）的缓冲用碳酸氢钠的等量直到鱼变得无法响应。一旦反应迟钝，立即将鱼放在一个黑色，无反射的背景。注:MS-222是有潜在毒性和手套应始终佩戴。 </li><li>拿在黑暗的房间所有的测量，以尽量减少多余光线超出了光谱仪的光源。 </li><li>使用针对生物体的体保持的光纤探针测量的反射率。使一定要形成光纤和所述主体之间的一致的45°角，如内置于一些探针的设计。注意:对于测量的一致性，可能希望在多个扫描（2 - 5）平均每身体区域。 </li><li>适当的反射分析软件记录反射率数据。注意:此处所使用的软件被设置为:积分时间= 2,000秒，平均扫描= 2，和像素平滑= 5的数据被复制并粘贴到电子表格。 </li><li>重复多个身体部位以确定有无反射紫外光和整个生物表型变异的范围。注意:初步扫描可能是必要的，以确定，如果有的话，身体区域一致显示紫外线标记。 </li><li>返回鱼良好的氧化控股水族馆，直到完全康复，由正常厣节奏和游泳的行为就是证明。注意:商业防护剂恢复粘膜涂覆到主体可以加入在皮肤恢复来帮助。 </li><li>众议员在多个个人吃的，包括如果需要男女双方。 </li><li>上，其中所有的数据都用散点图被复制，绘图数据，与在Y轴和波长上的X轴反射率的电子表格。在UV范围内幅度峰值（300 - 400纳米）表示反射紫外光对鱼的存在。 </li></ol> 2.二分交配试验注:执行控制的观察，其中焦鱼，UV过滤，和不透明滤波器到位。在这些试验中，在不透明过滤除去时，焦鱼将暴露于显示UV光的存在和不存在，但缺乏潜在交配伙伴罐部分。有关的分选择测试在本实验中的有效性详见Palmer和汉克森（2015）10，虽然注意到这些试验也有缺点11。 <ol><li>使用性接受个人作为焦点的有机体。使用女性P. latipinna &lt;48小时产后5次测试，以改善性动机的可能性之前，分离为至少24小时男性。 </li><li>将单焦点个体到相邻两个选择车厢中立地区。对于P. latipinna，使用75.7升试验水族馆（30.5厘米×30.5厘米×75厘米; 图1）由透明有机玻璃分区分为三段，形成了焦点个别由两个大小相等的选择隔间两侧的中心区域。确保没有水或嗅觉线索各个槽部之间共享。重复被观察每个个体。 </li></ol><img alt="图1" src="/files/ftp_upload/54453/54453fig1.jpg" /> 图1.实验水族馆设置一个长方形的水族馆分为三个部分:即持有焦点个人和持有的对象对两端车厢的中心区域（图中显示了与在两端对象男性中间有一个女）。可互换FILTER值，要么阻止或允许的UV反射率可以被放置在分割水族箱的男性和女性的部分中的玻璃隔板。这个数字是从Palmer和汉克森（2015年）10修改。 <a href="http://ecsource.jove.com/files/ftp_upload/54453/54453fig1large.jpg" target="_blank">请点击此处查看本图的放大版本。</a> <ol start="3"><li>在全谱照明条件（照明，其频谱延伸到UV-A范围），隔离在两端车厢（选择车厢）测试水族馆一个潜在的交配伙伴。确保潜在交配伙伴在大小，着色和显示特性相似彼此尽可能接近，使两个分离的配合伙伴之间的主要区别是它们是否出现一个滤波器，其阻止或传输紫外光后面。 注意:这可能是优选的以记录的反​​射率在可见光谱，以使可能的最佳色彩匹配，人虽然这里匹配用眼进行。 </li><li>适应视觉下的隔离（不透明过滤器）的个人15分钟。确保UV +和 - 过滤器在发生在这个时候。继驯化，去除不透明的过滤器。以确保侧偏差不存在（对于水箱的一侧，无论对象的固有偏好），随机地分配在UV +和紫外-槽的两侧的每个每次试验。 </li><li>通过使用秒表记录的时间的聚焦个人花费内的特定偏好区域（邻近的选择隔室区域，在这种情况下，大约两标准体长度）的量。对于女性的鱼，这是男性焦鱼它花费近每两个男性的时间量，而反之亦然。 注:事件记录器软件也可以用于记录。十五分钟试验在这个实验和鱼没有访问至少一次被排除在结果中的每一首区作为是使用反应迟钝。 </li><li>记录和分析鱼花近空UV +和UV的时间-隔室，以确保没有向可能影响交配选择鱼的一个特定的照明环境的固有偏差。使用配对t检验以确定是否个人偏好的特定紫外线条件。 </li></ol>

<ol>
	<li>Endler, J. A., Zaret, T. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Natural+and+sexual+selection+on+color+patterns+in+poeciliid+fishes.">Natural and sexual selection on color patterns in poeciliid fishes.</a> Evolutionary ecology of neotropical freshwater fishes. , 95-111 (1984).</li><li>Cummings, M. E., Rosenthal, G. G., Ryan, M. 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+private+ultraviolet+channel+in+visual+communication.">A private ultraviolet channel in visual communication.</a> Proc. R. Soc. Lond. [Biol]. 270 (1518), 897-904 (2003).</li><li>Siebeck, U. E., Parker, A. N., Sprenger, D., M&#228;thger, L. M., Wallis, 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+Species+of+Reef+Fish+that+Uses+Ultraviolet+Patterns+for+Covert+Face+Recognition.">A Species of Reef Fish that Uses Ultraviolet Patterns for Covert Face Recognition.</a> Curr. Biol. 20 (5), 407-410 (2010).</li><li>Hrbek, T., Seckinger, J., Meyer, 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+phylogenetic+and+biogeographic+perspective+on+the+evolution+of+poeciliid+fishes.">A phylogenetic and biogeographic perspective on the evolution of poeciliid fishes.</a> Mol. Phylogenet. Evol. 43 (3), 986-998 (2007).</li><li>K&#246;rner, K. E., Schlupp, I., Plath, M., Loew, E. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spectral+sensitivity+of+mollies:+comparing+surface-+and+cave-dwelling+Atlantic+mollies,+Poecilia+mexicana.">Spectral sensitivity of mollies: comparing surface- and cave-dwelling Atlantic mollies, Poecilia mexicana.</a> J. Fish Biol. 69 (1), 54-65 (2006).</li><li>Farr, J. A., Travis, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fertility+Advertisement+by+Female+Sailfin+Mollies,+Poecilia+latipinna+(Pisces:+Poeciliidae).">Fertility Advertisement by Female Sailfin Mollies, Poecilia latipinna (Pisces: Poeciliidae).</a> Copeia. 1986 (2), 467-472 (1986).</li><li>Ptacek, M. B., Travis, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mate+Choice+in+the+Sailfin+Molly,+Poecilia+latipinna.">Mate Choice in the Sailfin Molly, Poecilia latipinna.</a> Evolution. 51 (4), 1217-1231 (1997).</li><li>Ptacek, M., Grier, H. J., Uribe, M. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mating+signal+divergence,+sexual+selection+and+species+recognition+in+mollies+(Poeciliidae:+Poecilia:+Mollienesia).">Mating signal divergence, sexual selection and species recognition in mollies (Poeciliidae: Poecilia: Mollienesia).</a> Proceedings from the Second International Symposium on Livebearing Fishes. , 71-87 (2005).</li><li>MacLaren, R. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effects+of+male+proximity,+apparent+size,+and+absolute+size+on+female+preference+in+the+sailfin+molly,+Poecilia+latipinna.">The effects of male proximity, apparent size, and absolute size on female preference in the sailfin molly, Poecilia latipinna.</a> Behavior. 143 (12), 1457-1472 (2006).</li><li>Palmer, M. S., Hankison, 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=Use+of+ultraviolet+cues+in+female+mate+preference+in+the+sailfin+molly,+Poecilia+latipinna.">Use of ultraviolet cues in female mate preference in the sailfin molly, Poecilia latipinna.</a> Acta Ethol. 18 (2), 153-160 (2014).</li><li>Wagner, W. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Measuring+female+mating+preferences.">Measuring female mating preferences.</a> Anim. Behav. 55 (4), 1029-1042 (1998).</li><li>R&#233;my, A., Gr&#233;goire, A., Perret, P., Doutrelant, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mediating+male-male+interactions:+the+role+of+the+UV+blue+crest+coloration+in+blue+tits.">Mediating male-male interactions: the role of the UV blue crest coloration in blue tits.</a> Behav. Ecol. Sociobiol. 64 (11), 1839-1847 (2010).</li><li>Guillermo-Ferreira, R., Ther&#233;zio, E. M., Gehlen, M. H., Bispo, P. C., Marletta, 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+Role+of+Wing+Pigmentation,+UV+and+Fluorescence+as+Signals+in+a+Neotropical+Damselfly.">The Role of Wing Pigmentation, UV and Fluorescence as Signals in a Neotropical Damselfly.</a> J. Insect Behav. 27 (1), 67-80 (2013).</li><li>Ord, T. J., Stamps, J. A., Losos, J. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Convergent+evolution+in+the+territorial+communication+of+a+classic+adaptive+radiation:+Caribbean+Anolis+lizards.">Convergent evolution in the territorial communication of a classic adaptive radiation: Caribbean Anolis lizards.</a> Anim. Behav. 85 (6), 1415-1426 (2013).</li><li>Siebeck, U. E., Witzany, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Communication+in+the+Ultraviolet:+Unravelling+the+Secret+Language+of+Fish.">Communication in the Ultraviolet: Unravelling the Secret Language of Fish.</a> Biocommunication of Animals. , 299-320 (2014).</li><li>Modarressie, R., Rick, I. P., Bakker, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=UV+matters+in+shoaling+decisions.">UV matters in shoaling decisions.</a> Proc. Biol. Sci. 273 (1588), 849-854 (2006).</li><li>Lim, M. L. M., Li, J., Li, D. <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+UV-reflecting+markings+on+female+mate-choice+decisions+in+Cosmophasis+umbratica,+a+jumping+spider+from+Singapore.">Effect of UV-reflecting markings on female mate-choice decisions in Cosmophasis umbratica, a jumping spider from Singapore.</a> Behav. Ecol. 19, 61-66 (2008).</li><li>Rick, I. P., Bakker, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Color+signaling+in+conspicuous+red+sticklebacks:+do+ultraviolet+signals+surpass+others?.">Color signaling in conspicuous red sticklebacks: do ultraviolet signals surpass others?.</a> BMC Evol. Biol. 8, 189 (2008).</li><li>Siebeck, U. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Communication+in+coral+reef+fish:+the+role+of+ultraviolet+colour+patterns+in+damselfish+territorial+behaviour.">Communication in coral reef fish: the role of ultraviolet colour patterns in damselfish territorial behaviour.</a> Anim. Behav. 68 (2), 273-282 (2004).</li><li>Cummings, M. E., De Le&#242;n, F. J. G., Mollaghan, D. M., Ryan, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Is+UV+ornamentation+an+amplifier+in+swordtails?.">Is UV ornamentation an amplifier in swordtails?.</a> Zebrafish. 3, 91-100 (2006).</li><li>Mac&#236;as Garcia, c., de Perera, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ultraviolet-based+female+preferences+in+a+viviparous+fish.">Ultraviolet-based female preferences in a viviparous fish.</a> Behav. Ecol. Sociobiol. 52 (1), 1-6 (2002).</li></ol>

The authors have nothing to disclose.

分光光度法成功地确定对P. UV标志latipinna。 P的两性latipinna具有沿其两侧UV标记。此外，一些男性对他们背鳍紫外标记，先前发现的性状是在雌性交配偏好7重要。 我们建议使用紫外分光光度法作为一种机制来检测紫外线标志的存在。进一步的测试可能确定其在社会互动，包括交配偏好（如这里详细介绍）的作用。可替代地，紫外线特性可能?...

了解线索，并在动物社会交往使用的信号可以让我们内部和物种之间理解的表型变异。此变化起着进化过程如人口发散，性选择，和形态的重要作用。通常，然而，研究人员正在限于探索最明显的人类感觉系统，特别是那些在视觉或听觉领域内的提示。光度法的使用，然而，使我们能够扩展超出人类可见光谱并进入波长可能在其他物种的社会互动重要我们的调查。 特别是，通过紫外线带来的短距离通信（UV; 300 - 400纳米）的灵敏度必须是择偶1中非常有利的可能性。鸟类和鱼类的许多视觉捕猎的食肉动物，比如，无法检测紫外线辐射。在男性精心显示系统到女性，这些男性会减少他们的捕食的风险，同时保持其通过利用UV光谱，而不是开发线索可检测在可见光谱2,3到吸引异性能力。如果一个发生故障考虑到生物体与每个通信的可能性其他使用这些"私人沟通渠道"，行为和进化显著驱动程序可能会被错过。 该协议概述了调查的茉莉花鳉， 孔雀鱼latipinna，已经没有先前已知的检测UV或利用UV标记能力的一夫多妻鱼使用UV线索的择偶。该鱼种有着密切的亲缘接近其它紫外线敏感的活轴承鱼4和有显微光谱证据表明P. latipinna，与其它茉莉物种如铜绿沿墨西哥和P.台塑 ，拥有一流的视锥细胞（负责统筹感光细胞LOR视觉）是最敏感的UV波长5。在这种性别二态性的物种，女性选择了在男性&#39;颜色鲜艳和扩大散热片6-9的进化中发挥了强有力的作用。这种方法使我们能够探索紫外线是由女性评估男性的质量的附加网上平台。 检测和P.紫外标记测量latipinna使用分光光度计用光纤探头这里详细描述。此外，接受女性mollies是否差异通过光学滤镜观察男性发射全光谱光源包括UV-A关联（[UV +]; 320 - 700纳米），并通过UV阻隔滤镜观察男性（[紫外线]; 400 - 700纳米）进行了讨论。该方法具有用于发现在鱼类和其他生物的UV敏感性和彩色图案，允许研究多种方法包括紫外线和其行为的作用的问题广泛的应用。

<table border="0" cellpadding="0" cellspacing="0" width="1044">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:357px;">Spectrophotometer, P1000</td>
			<td style="width:217px;">Ocean Optics</td>
			<td style="width:119px;"></td>
			<td style="width:351px;">newer models are availabe</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:357px;">DT 1000 xenon UV light source</td>
			<td style="width:217px;">Ocean Optics</td>
			<td style="width:119px;"></td>
			<td>newer models are availabe</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Ocean Optics Overture Software</td>
			<td style="width:217px;">Ocean Optics</td>
			<td style="width:119px;"></td>
			<td>newer software is available</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">R200-Angle-UV bifurcated fiber-optic probe (Guided Wave)</td>
			<td style="width:217px;">Ocean Optics</td>
			<td style="width:119px;"></td>
			<td>newer models are availabe</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Certified reflectance standard, white</td>
			<td style="width:217px;">labsphere</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:357px;">75.7 L Aquarium, divided</td>
			<td style="width:217px;">Experimental Builder</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:357px;">Full Spectrum Bulb</td>
			<td style="width:217px;">Nature&#39;s Sunlight</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:357px;">UV blocking sheet</td>
			<td style="width:217px;">GAM UV Sheet</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
	</tbody>
</table>

assays to detect uv-reflecting structures and determine their importance in mate preference using the sailfin molly poecilia latipinna

许多生物过程中的社会交往使用暗示和信号超出人类的敏感性。试图了解他们的行为和生态环境时，要考虑到生物体是如何看待他们的世界是很重要的。灵敏度的紫外光谱（UV; 300 - 400 nm）的跨鸟类，鱼类，爬行类，两栖类，甚至哺乳动物的多个属找到。这个协议描述用于检查生物体紫外线反射结构的存在和用于测试这些线索是否被用作在择偶的上下文社会信号的方法的技术。分光光度计被用来检测反射紫外光和变异在个体和两性之间的反射强度的存在。提出这种技术的一个例子，其中一个二分法的择偶测试暴露性接受个人异性个体，其外观可以通过过滤器，无论是传输全谱或阻挡紫外线的波长进行操作。这个系统允许的女性，而不是男性，sailfin mollies（孔雀鱼latipinna）采用UV的标记，作为其交配的决定的一部分的决心。这些类型的研究，有助于扩大我们的，利用UV生物体的各种知识和洞察UV如何发挥他们生活中的作用。

<html> 图1和2示出了交配偏好水族馆建立和UV测量部位为我们的实验。 测量允许的决心，紫外线反射P. latipinna做具有紫外线的特性，特别是沿其身体（ 图3）的侧面，除了在这些性状的个体差异。一旦UV性状发现，测试显示，女性，而不是男性，P. latipinna在它们的交配决定使用?...

Watch this Scientific Journal Video about Assays to Detect UV-reflecting Structures and Determine their Importance in Mate Preference using the Sailfin Molly Poecilia latipinna at JoVE.com

Assays to Detect UV-reflecting Structures and Determine their Importance in Mate Preference using the Sailfin Molly Poecilia latipinna

这个协议概括了使用分光光度法来检测生物体紫外线反射结构（在本例中，茉莉花鳉孔雀latipinna），并描述了对分选择测试鱼类允许要在紫外线线索的伴侣选择期间的作用作出推论。

试验以检测紫外线反射结构，并使用茉莉花鳉确定其重要性伴侣偏好<em&gt;孔雀鱼latipinna</em

Many organisms use cues and signals beyond human sensitivity during social interactions. It is important to take into account how organisms perceive their ...

assays-to-detect-uv-reflecting-structures-determine-their-importance

Research

JoVE Journal

Neuroscience

Assays to Detect UV-reflecting Structures and Determine their Importance in Mate Preference using the Sailfin Molly Poecilia latipinna

6.5K 次观看.  Ohio Wesleyan University. 这个协议概括了使用分光光度法来检测生物体紫外线反射结构（在本例中，茉莉花鳉孔雀latipinna），并描述了对分选择测试鱼类允许要在紫外线线索的伴侣选择期间的作用作出推论。 

视频: Assays to Detect UV-reflecting Structures and Determine their Importance in Mate Preference using the Sailfin Molly Poecilia latipinna

Light Preference Assay to Study Innate and Circadian Regulated Photobehavior in Drosophila Larvae

Light acts as environmental signal to control animal behavior at various levels. The Drosophila larval nervous system is used as a unique model to answer basic questions on how light information is processed and shared between rapid and circadian behaviors. Drosophila larvae display a stereotypical avoidance behavior when exposed to light. To investigate light dependent behaviors comparably simple light-dark preference tests can be applied. In vertebrates and arthropods the neural pathways involved in sensing and processing visual inputs partially overlap with those processing photic circadian information. The fascinating question of how the light sensing system and the circadian system interact to keep behavioral outputs coordinated remains largely unexplored. Drosophila is an impacting biological model to approach these questions, due to a small number of neurons in the brain and the availability of genetic tools for neuronal manipulation. The presented light-dark preference assay allows the investigation of a range of visual behaviors including circadian control of phototaxis.

Light Preference Assay to Study Innate and Circadian Regulated Photobehavior in Drosophila Larvae

Here we describe a light-dark preference test for Drosophila larva. This assay provides information about innate and circadian regulation of light sensing and processing photobehavior.

光偏好的检测，以研究的先天和昼夜调节感光特性<em&gt;果蝇</em&gt;幼虫

Light acts as environmental signal to control animal behavior at various levels. The Drosophila larval nervous system is used as a unique model to answer ...

科研

神经科学

Fast Micro-iontophoresis of Glutamate and GABA: A Useful Tool to Investigate Synaptic Integration

One of the fundamental interests in neuroscience is to understand the integration of excitatory and inhibitory inputs along the very complex structure of the dendritic tree, which eventually leads to neuronal output of action potentials at the axon. The influence of diverse spatial and temporal parameters of specific synaptic input on neuronal output is currently under investigation, e.g. the distance-dependent attenuation of dendritic inputs, the location-dependent interaction of spatially segregated inputs, the influence of GABAergig inhibition on excitatory integration, linear and non-linear integration modes, and many more.
With fast micro-iontophoresis of glutamate and GABA it is possible to precisely investigate the spatial and temporal integration of glutamatergic excitation and GABAergic inhibition. Critical technical requirements are either a triggered fluorescent lamp, light-emitting diode (LED), or a two-photon scanning microscope to visualize dendritic branches without introducing significant photo-damage of the tissue. Furthermore, it is very important to have a micro-iontophoresis amplifier that allows for fast capacitance compensation of high resistance pipettes. Another crucial point is that no transmitter is involuntarily released by the pipette during the experiment.
Once established, this technique will give reliable and reproducible signals with a high neurotransmitter and location specificity. Compared to glutamate and GABA uncaging, fast iontophoresis allows using both transmitters at the same time but at very distant locations without limitation to the field of view. There are also advantages compared to focal electrical stimulation of axons: with micro-iontophoresis the location of the input site is definitely known and it is sure that only the neurotransmitter of interest is released. However it has to be considered that with micro-iontophoresis only the postsynapse is activated and presynaptic aspects of neurotransmitter release are not resolved. In this article we demonstrate how to set up micro-iontophoresis in brain slice experiments.

In this article we introduce fast micro-iontophoresis of neurotransmitters as a technique to investigate integration of postsynaptic signals with high spatial and temporal precision.

谷氨酸和GABA快速微离子导入:一个有用​​的工具，调查突触整合

One of the fundamental interests in neuroscience is to understand the integration of excitatory and inhibitory inputs along the very complex structure of ...

Isolation and Quantification of Botulinum Neurotoxin From Complex Matrices Using the BoTest Matrix Assays

Accurate detection and quantification of botulinum neurotoxin (BoNT) in complex matrices is required for pharmaceutical, environmental, and food sample testing. Rapid BoNT testing of foodstuffs is needed during outbreak forensics, patient diagnosis, and food safety testing while accurate potency testing is required for BoNT-based drug product manufacturing and patient safety. The widely used mouse bioassay for BoNT testing is highly sensitive but lacks the precision and throughput needed for rapid and routine BoNT testing. Furthermore, the bioassay&#39;s use of animals has resulted in calls by drug product regulatory authorities and animal-rights proponents in the US and abroad to replace the mouse bioassay for BoNT testing. Several in vitro replacement assays have been developed that work well with purified BoNT in simple buffers, but most have not been shown to be applicable to testing in highly complex matrices. Here, a protocol for the detection of BoNT in complex matrices using the BoTest Matrix assays is presented. The assay consists of three parts: The first part involves preparation of the samples for testing, the second part is an immunoprecipitation step using anti-BoNT antibody-coated paramagnetic beads to purify BoNT from the matrix, and the third part quantifies the isolated BoNT&#39;s proteolytic activity using a fluorogenic reporter. The protocol is written for high throughput testing in 96-well plates using both liquid and solid matrices and requires about 2 hr of manual preparation with total assay times of 4-26 hr depending on the sample type, toxin load, and desired sensitivity. Data are presented for BoNT/A testing with phosphate-buffered saline, a drug product, culture supernatant, 2% milk, and fresh tomatoes and includes discussion of critical parameters for assay success.

The BoTest Matrix botulinum neurotoxin (BoNT) detection assays rapidly purify and quantify BoNT from a range of sample matrices.&#160;Here, we present a protocol for the detection and quantification of BoNT from both solid and liquid matrices and demonstrate the assay with BOTOX, tomatoes, and milk.

隔离和肉毒杆菌神经毒素定量从复杂的基质使用BoTest矩阵测定

Accurate detection and quantification of botulinum neurotoxin (BoNT) in complex matrices is required for pharmaceutical, environmental, and food sample ...

Use of a Caspase Multiplexing Assay to Determine Apoptosis in a Hypothalamic Cell Model

The ability to multiplex assays in studies of complex cellular mechanisms eliminates the need for repetitive experiments, provides internal controls, and decreases waste in costs and reagents. Here we describe optimization of a multiplex assay to assess apoptosis following a palmitic acid (PA) challenge in an in vitro hypothalamic model, using both fluorescent and luminescent based assays to measure viable cell counts and caspase-3/7 activity in a 96-well microtiter plate format. Following PA challenge, viable cells were determined by a resazurin-based fluorescent assay. Caspase-3/7 activity was then determined using a luminogenic substrate, DEVD, and normalized to cell number. This multiplexing assay is a useful technique for determining change in caspase activity following an apoptotic stimulus, such as saturated fatty acid challenge. The saturated fatty acid PA can increase hypothalamic oxidative stress and apoptosis, indicating the potential importance of assays such as that described here in studying the relationship between saturated fatty acids and neuronal function.

Multiplex assays can provide beneficial information for basic cellular mechanisms&#160;and eliminate waste of reagents and unnecessary repetitive experiments. We describe here a multiplex caspase-3/7 activity assay, using fluorescent- and luminescent-based methods, to determine cell viability in an in vitro hypothalamic model following oxidative challenge with palmitic acid.

使用胱天蛋白酶复分析，以确定细胞凋亡的下丘脑细胞模型

The ability to multiplex assays in studies of complex cellular mechanisms eliminates the need for repetitive experiments, provides internal controls, and ...

Investigating the Function of Deep Cortical and Subcortical Structures Using Stereotactic Electroencephalography: Lessons from the Anterior Cingulate Cortex

Stereotactic Electroencephalography (SEEG) is a technique used to localize seizure foci in patients with medically intractable epilepsy. This procedure involves the chronic placement of multiple depth electrodes into regions of the brain typically inaccessible via subdural grid electrode placement. SEEG thus provides a unique opportunity to investigate brain function. In this paper we demonstrate how SEEG can be used to investigate the role of the dorsal anterior cingulate cortex (dACC) in cognitive control. We include a description of the SEEG procedure, demonstrating the surgical placement of the electrodes. We describe the components and process required to record local field potential (LFP) data from consenting subjects while they are engaged in a behavioral task. In the example provided, subjects play a cognitive interference task, and we demonstrate how signals are recorded and analyzed from electrodes in the dorsal anterior cingulate cortex, an area intimately involved in decision-making. We conclude with further suggestions of ways in which this method can be used for investigating human cognitive processes.

Stereotactic Electroencephalography (SEEG) is an operative technique used in epilepsy surgery to help localize seizure foci. It also affords a unique opportunity to investigate brain function. Here we describe how SEEG can be used to investigate cognitive processes in human subjects.

调查深皮层的功​​能和皮层下结构采用立体定向脑电图:由前扣带皮层的经验教训

Stereotactic Electroencephalography (SEEG) is a technique used to localize seizure foci in patients with medically intractable epilepsy. This procedure ...

Using Single Sensillum Recording to Detect Olfactory Neuron Responses of Bed Bugs to Semiochemicals

The insect olfactory system plays an important role in detecting semiochemicals in the environment. In particular, the antennal sensilla which house single or multiple neurons inside, are considered to make the major contribution in responding to the chemical stimuli. By directly recording action potential in the olfactory sensillum after exposure to stimuli, single sensillum recording (SSR) technique provides a powerful approach for investigating the neural responses of insects to chemical stimuli. For the bed bug, which is a notorious human parasite, multiple types of olfactory sensillum have been characterized. In this study, we demonstrated neural responses of bed bug olfactory sensilla to two chemical stimuli and the dose-dependent responses to one of them using the SSR method. This approach enables researchers to conduct early screening for individual chemical stimuli on the bed bug olfactory sensilla, which would provide valuable information for the development of new bed bug attractants or repellents and benefits the bed bug control efforts.

Bed bugs rely on olfactory receptor neurons housed in their antennal olfactory sensilla to detect semiochemicals in the environment. Utilizing single sensillum recording, we demonstrate a method to evaluate bed bug response to semiochemicals and explore the coding process involved.

使用单一感器录音来检测臭虫的嗅觉神经元的反应，以信息化学物质

The insect olfactory system plays an important role in detecting semiochemicals in the environment. In particular, the antennal sensilla which house ...

Taste Preference Assay for Adult Drosophila

Olfactory and gustatory perception of the environment is vital for animal survival. The most obvious application of these chemosenses is to be able to distinguish good food sources from potentially dangerous food sources. Gustation requires physical contact with a chemical compound which is able to signal through taste receptors that are expressed on the surface of neurons. In insects, these gustatory neurons can be located across the animal's body allowing taste to play an important role in many different behaviors. Insects typically prefer compounds containing sugars, while compounds that are considered bitter tasting are avoided. Given the basic biological importance of taste, there is intense interest in understanding the molecular mechanisms underlying this sensory modality. We describe an adult Drosophila taste assay which reflects the preference of the animals for a given tastant compound. This assay may be applied to animals of any genetic background to examine the taste preference for a desired soluble compound.

Taste Preference Assay for Adult Drosophila

Taste is an important sensory process which facilitates attraction to beneficial substances and avoidance of toxic substances. This protocol describes a simple ingestion assay for determining Drosophila gustatory preference for a given chemical compound.

口味偏好测定成人<em&gt;果蝇</em

Olfactory and gustatory perception of the environment is vital for animal survival.  The most obvious application of these chemosenses is to be able to ...

Enumeration of Neural Stem Cells Using Clonal Assays

Neural stem cells (NSCs) have the ability to self-renew and generate the three major neural lineages &#8212; astrocytes, neurons and oligodendrocytes. NSCs and neural progenitors (NPs) are commonly cultured in vitro as neurospheres. This protocol describes in detail how to determine the NSC frequency in a given cell population under clonal conditions. The protocol begins with the seeding of the cells at a density that allows for the generation of clonal neurospheres. The neurospheres are then transferred to chambered coverslips and differentiated under clonal conditions in conditioned medium, which maximizes the differentiation potential of the neurospheres. Finally, the NSC frequency is calculated based on neurosphere formation and multipotency capabilities. Utilities of this protocol include the evaluation of candidate NSC markers, purification of NSCs, and the ability to distinguish NSCs from NPs. This method takes 13 days to perform, which is much shorter than current methods to enumerate NSC frequency.

Neural stem cells (NSCs) refer to cells&#160;which can self-renew and differentiate into the three neural lineages. Here, we describe a protocol to determine NSC frequency in a given cell population using neurosphere formation and differentiation under clonal conditions.

使用克隆测定神经干细胞的计数。

Neural stem cells (NSCs) have the ability to self-renew and generate the three major neural lineages &#8212; astrocytes, neurons and oligodendrocytes. ...

Microglia as a Surrogate Biosensor to Determine Nanoparticle Neurotoxicity

Nanoparticles found in air pollutants can alter neurotransmitter profiles, increase neuroinflammation, and alter brain function. Therefore, the assay described here will aid in elucidating the role of microglia in neuroinflammation and neurodegenerative diseases. The use of microglia, resident immune cells of the brain, as a surrogate biosensor provides novel insight into how inflammatory responses mediate neuronal insults. Here, we utilize an immortalized murine microglial cell line, designated BV2, and describe a method for nanoparticle exposure using silver nanoparticles (AgNPs) as a standard. We describe how to expose microglia to nanoparticles, how to remove nanoparticles from supernatant, and how to use supernatant from activated microglia to determine toxicity, using hypothalamic cell survival as a measure. Following AgNP exposure, BV2 microglial activation was validated using a tumor necrosis factor alpha (TNF-&#945;) enzyme linked immunosorbent assay (ELISA). The supernatant was filtered to remove the AgNP and to allow cytokines and other secreted factors to remain in the conditioned media. Hypothalamic cells were then exposed to supernatant from AgNP activated microglia and survival of neurons was determined using a resazurin-based fluorescent assay. This technique is useful for utilizing microglia as a surrogate biomarker of neuroinflammation and determining the effect of neuroinflammation on other cell types.

Microglia (immune cells of the brain), are used as a surrogate biosensor to determine how nanoparticles influence neurotoxicity. We describe a series of experiments designed to assay microglial response to nanoparticles and exposure of hypothalamic neurons to supernatant from activated microglia to determine neurotoxicity.

小胶质细胞作为替代生物传感器确定纳米颗粒神经毒性

Nanoparticles found in air pollutants can alter neurotransmitter profiles, increase neuroinflammation, and alter brain function. Therefore, the assay ...

Exploring Deep Space - Uncovering the Anatomy of Periventricular Structures to Reveal the Lateral Ventricles of the Human Brain

Anatomy students are typically provided with two-dimensional (2D) sections and images when studying cerebral ventricular anatomy and students find this challenging. Because the ventricles are negative spaces located deep within the brain, the only way to understand their anatomy is by appreciating their boundaries formed by related structures. Looking at a 2D representation of these spaces, in any of the cardinal planes, will not enable visualisation of all of the structures that form the boundaries of the ventricles. Thus, using 2D sections alone requires students to compute their own mental image of the 3D ventricular spaces. The aim of this study was to develop a reproducible method for dissecting the human brain to create an educational resource to enhance student understanding of the intricate relationships between the ventricles and periventricular structures. To achieve this, we created a video resource that features a step-by-step guide using a fiber dissection method to reveal the lateral and third ventricles together with the closely related limbic system and basal ganglia structures. One of the advantages of this method is that it enables delineation of the white matter tracts that are difficult to distinguish using other dissection techniques. This video is accompanied by a written protocol that provides a systematic description of the process to aid in the reproduction of the brain dissection. This package offers a valuable anatomy teaching resource for educators and students alike. By following these instructions educators can create teaching resources and students can be guided to produce their own brain dissection as a hands-on practical activity. We recommend that this video guide be incorporated into neuroanatomy teaching to enhance student understanding of the morphology and clinical relevance of the ventricles.

This paper demonstrates the effective use of a fiber dissection method to reveal the superficial white matter tracts and periventricular structures of the human brain, in three-dimensional space, to aid student comprehension of ventricular morphology.

探索深空-揭开人脑侧脑室的脑室结构解剖

Anatomy students are typically provided with two-dimensional (2D) sections and images when studying cerebral ventricular anatomy and students find this ...