我们承认股市布卢明顿中心飞株。这项工作是由BBSRC（BB / G008973 / 1）支持的研究经费。

1，准备飞<ol><li>交配的所有突变体，Gal4的 / UAS和其他线路用野生型菌株，如CSw-，至少六代之前的行为实验，以控制对遗传背景26。 </li><li>在12:12小时明暗周期长的苍蝇在一个标准的玉米粉，酵母糖蜜食品的饮食，在25℃，除非有具体的操作需要不同的温度。 <ol><li>为了确定转基因表达的影响：使用18℃，以防止通过发展Gal4的GAL80 TS（目标系统）转基因的表达，然后在行为实验前1-2天将苍蝇到30℃培养箱中培养。进行实验，在30℃，以确定转基因的表达3,4,6,7,14的影响。 </li><li>对于使用热激活TRPA1通道，以刺激神经元的实验：提高苍蝇在23°C，这是众所周知的温度保持非活动通道，然后转变为行为的房间在30°C，激活神经元表达的TRPA1。 </li><li>对于使用Shibire 温度敏感的阻断突触输出11，14,24实验：后苍蝇在18℃，并进行测试，在30℃。 </li></ol></li><li>收集苍蝇1-2天的实验前，在轻的CO 2麻醉指望他们在大约25组。存储苍蝇至少O / N的食品瓶（不含酵母），在25°C（除非特定的操作需要不同的温度），相对湿度70％的环境控制与空间12:12小时光照：待到黑暗条件实验。 注：该存储允许苍蝇使其恢复到在环境控制的房间，其中有对果蝇学习的最佳条件进行后续的学习和考试，重要的是，删除任何日常ENVI境的​​变化，可能影响到行为的表型。 </li></ol> 2，准备实验前<ol><li>执行在一个特制的有机玻璃T-迷宫实验（ 图1）。 </li><li>定期检查管接头，以保证在实验过程中得到的气密密封。如果需要，可以改变的O型环密封的T-迷宫的内部隔间。 </li><li>将特制的铜网上的培训管道内。定期检查和清理这些网格，并更换，如果氧化。通过运行于开关箱连接到电刺激器鳄鱼夹附的铜网上，以导线。使用电压表，以确保该装置提供所需的冲击。 </li><li>使用G-夹来保存迷宫紧密，以避免漏风。 </li><li>附加的T型迷宫管道运行到一个空气泵，以使气味被苍蝇跨绘制并且随后从T-迷宫除去。保持温和的空气流在〜2升/分钟</li></ol> 3，气味稀释<ol><li>使用两种不同的气味的浓度，使得蝇显示出相等偏好既气味。使用4 -甲基环己醇（1:67）和3 -辛醇（1:100）稀释，在矿物油中7.13。 注意：仔细确定这些浓度，这将通过实验室有所不同。比如别人用1:10既气味24。其他常用的气味包括乙酸乙酯和乙酸异戊酯。 </li><li>吸取30μL稀释成味定做的气味杯放置在顶部有穿孔的，它允许将绘制在气味在杯中空气覆盖了塑料管的气味块，从而暴露了苍蝇的气味羽。 </li></ol> 4，训练协议（图1和图2） <ol><li>对于成年嗅震荡调理，进行下昏暗的红灯所有的实验（ 即 ，红色LED），这使得研究人员看到，但可以防止FLÝ看见，从而使苍蝇集中于嗅觉而非视觉输入。 </li><li>引入果蝇到训练管，然后装到T型迷宫，并让他们适应管和气流为90秒。 </li><li>存在于第一气味（4-甲基环己醇，MCH）与60-V的冲击（包括12 1.25秒的脉冲以3.75秒的脉冲间的间隔）为60秒的总时间。 </li><li>按照有没有异味或冲击30秒的休息时间的震荡。 </li><li>目前，第二气味（3 - 辛醇，华侨城）60秒，而没有震动。 </li><li>按照有没有异味或冲击30秒的休息时间的震荡。 </li><li>将苍蝇从训练室到T-迷宫的中心腔被打开侧面的T型迷宫，轻轻敲打在柔软的表面T型迷宫的底部，如旧的鼠标垫。维持蝇的中央腔室90秒。 </li><li>适合选择管进入微博该装置的TTOM以形成T型迷宫。 </li><li>以测量学习，将苍蝇的选择点在T-迷宫，在那里他们被同时暴露于两者的气味和走向之一。进行试验期为120秒。 </li><li>陷阱中选择管苍蝇滑动中央腔向上从而阻断的选择管子的端部。收集苍蝇在T-迷宫的每个臂和在中央室进入食品小瓶和计数。 </li><li>来测量存储器，训练（4.6）后收集的苍蝇和从T-迷宫无酵母他们转移到食物的小瓶中。商店拥有在黑暗中在25℃和70％湿度下保持必要的剩余时间，以确定所关注的存储器阶段（见介绍）。重新引入果蝇到T型迷宫中阶段4.7。 </li><li>对于长期记忆，使用定制的迷宫，让苍蝇几个批次可以同时培训。辖5个周期的训练，15分钟跨周期的时间间隔（水疗中心CED）或不跨周期的时间间隔（集结）。保持苍蝇在18°C和70％的湿度在黑暗中，直到测试。在测试之前，将苍蝇至25℃，并允许他们适应环境至少1小时。评估长期记忆训练后的24小时。 </li><li>在行为学实验后，干净的气味杯子用热水和无味的洗涤剂。干燥后，涂杯子用10微升sigmacote。干sigmacote加热的微波炉。偶尔清理的T-迷宫有机玻璃管和气味块，用热水和清洁剂无味。 </li></ol> 5，计算性能指数：果蝇的记忆的测量<ol><li>计算出的性能指数（PI）为每个条件苍蝇避免电击配对的气味（CS - ）的数目减去苍蝇选择休克配对的气味（CS +）的果蝇（CS的总数除以数- + CS +）1。 PErformance指数（PI）=（＃CS -飞- ＃CS +飞）/（＃总苍蝇） </li><li>通过平均该实验，其中MCH是休克配对的气味和在其中的OCT是休克配对的气味的PI计算实验的最后的PI。这消除有一个气味更高的优先苍蝇的任何偏差。 </li></ol> 6，感觉运动控制<ol><li>通过引入〜40-50蝇到T-迷宫6,7,17 ​​执行气味视力。 </li><li> 90秒后，将苍蝇的选择点，并且允许他们2分钟纯净的气味和空气之间进行选择。 </li><li>收集和计数的苍蝇。通过将苍蝇选择的气味由那些参加测试的总数计算避免百分比。 </li><li>休克反应6,7,17，介绍苍蝇进入震荡室。 </li><li> 90秒的休息后，管理一个60 V直流触电，从苍蝇能逃脱类似管无震动。 </li><li>等待2分钟的苍蝇选择;收集和计算的苍蝇。通过将苍蝇避免震荡由苍蝇在实验的总数逸出的激波管的数量计算冲击回避百分比。包括保留在中央腔室中的那些逃脱电击的总的苍蝇。 </li></ol>

<ol>
	<li>Tully, T., Quinn, W. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Classical+conditioning+and+retention+in+normal+and+mutant+Drosophila+melanogaster.">Classical conditioning and retention in normal and mutant Drosophila melanogaster.</a> Journal of Comparative Physiology A. 157, 263-277 (1985).</li><li>Bolduc, F. V., Tully, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fruit+flies+and+intellectual+disability.">Fruit flies and intellectual disability.</a> Fly (Austin). 3, 91-104 (2009).</li><li>McGuire, S. E., Deshazer, M., Davis, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Thirty+years+of+olfactory+learning+and+memory+research+in+Drosophila+melanogaster.">Thirty years of olfactory learning and memory research in Drosophila melanogaster.</a> Prog Neurobiol. 76, 328-347 (2005).</li><li>Keene, A. C., Waddell, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+olfactory+memory:+single+genes+to+complex+neural+circuits.">Drosophila olfactory memory: single genes to complex neural circuits.</a> Nat Rev Neurosci. 8, 341-354 (2007).</li><li>Chiang, H. C., Wang, L., Xie, Z., Yau, A., Zhong, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=PI3+kinase+signaling+is+involved+in+Abeta-induced+memory+loss+in+Drosophila.">PI3 kinase signaling is involved in Abeta-induced memory loss in Drosophila.</a> Proc Natl Acad Sci USA. 107, 7060-7065 (2010).</li><li>Kanellopoulos, A. K., Semelidou, O., Kotini, A. G., Anezaki, M., Skoulakis, E. 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=Learning+and+memory+deficits+consequent+to+reduction+of+the+Fragile+X+mental+retardation+protein+result+from+metabotropic+glutamate-mediated+inhibition+of+cAMP+signalling+in+Drosophila.">Learning and memory deficits consequent to reduction of the Fragile X mental retardation protein result from metabotropic glutamate-mediated inhibition of cAMP signalling in Drosophila.</a> J Neurosci. 32, 13111-13124 (2012).</li><li>Malik, B. R., Gillespie, J. M., Hodge, J. J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=CASK+and+CaMKII+function+in+the+mushroom+body+a&#8217;/&#223;&#8217;+neurons+during+Drosophila+memory+formation.">CASK and CaMKII function in the mushroom body a&#8217;/&#223;&#8217; neurons during Drosophila memory formation.</a> Front Neural Circuits. 7, 52 (2013).</li><li>Gerber, B., Stocker, R. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Drosophila+larva+as+a+model+for+studying+chemosensation+and+chemosensory+learning:+a+review.">The Drosophila larva as a model for studying chemosensation and chemosensory learning: a review.</a> Chem Senses. 32, 65-89 (2007).</li><li>Gillespie, J. M., Hodge, J. J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=CASK+regulates+CaMKII+autophosphorylation+in+control+of+synaptic+growth+and+appetitive+learning.">CASK regulates CaMKII autophosphorylation in control of synaptic growth and appetitive learning.</a> Front Molecular Neuroscience. 6, 27 (2013).</li><li>Apostolopoulou, A. A., Widmann, A., Rohwedder, A., Pfitzenmaier, J. E., Thum, 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=Appetitive+associative+olfactory+learning+in+Drosophila+larvae.">Appetitive associative olfactory learning in Drosophila larvae.</a> J Vis Exp. (72), e4334 (2013).</li><li>Tonoki, A., Davis, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Aging+impairs+intermediate-term+behavioral+memory+by+disrupting+the+dorsal+paired+medial+neuron+memory+trace.">Aging impairs intermediate-term behavioral memory by disrupting the dorsal paired medial neuron memory trace.</a> Proc Natl Acad Sci USA. 109, 6319-6324 (2012).</li><li>Yamazaki, D., Horiuchi, J., Nagano, S., Tamura, T., Saitoe, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Drosophila+DCO+mutation+suppresses+age-related+memory+impairment+without+affecting+lifespan.">The Drosophila DCO mutation suppresses age-related memory impairment without affecting lifespan.</a> Nat Neurosci. 10, 478-484 (2007).</li><li>Cavaliere, S., Malik, B. R., Hodge, J. J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=KCNQ+channels+regulate+age-related+memory+impairment.">KCNQ channels regulate age-related memory impairment.</a> PLoS One. 8, e62445 (2013).</li><li>Venken, K. J., Simpson, J. H., Bellen, H. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+manipulation+of+genes+and+cells+in+the+nervous+system+of+the+fruit+fly.">Genetic manipulation of genes and cells in the nervous system of the fruit fly.</a> Neuron. 72, 202-230 (2011).</li><li>Isabel, G., Pascual, A., Preat, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exclusive+consolidated+memory+phases+in+Drosophila.">Exclusive consolidated memory phases in Drosophila.</a> Science. 304, 1024-1027 (2004).</li><li>Perisse, E., Burke, C., Huetteroth, W., Waddell, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Shocking+revelations+and+saccharin+sweetness+in+the+study+of+Drosophila+olfactory+memory.">Shocking revelations and saccharin sweetness in the study of Drosophila olfactory memory.</a> Curr Biol. 23, R752-R763 (2013).</li><li>Tully, T., Preat, T., Bonyton, S. C., Del Vecchio, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+dissection+of+consolidated+memory+in+Drosophila.">Genetic dissection of consolidated memory in Drosophila.</a> Cell. 79, 35-47 (1994).</li><li>Lyons, L. C., Roman, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Circadian+modulation+of+short-term+memory+in+Drosophila.">Circadian modulation of short-term memory in Drosophila.</a> Learning and memory. 16, 19-27 (2009).</li><li>Le Glou, E., Seugnet, L., Shaw, P. J., Preat, T., Gouguel, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Circadian+modulation+of+consolidated+memory+retrieval+following+sleep+deprivation+in+Drosophila.">Circadian modulation of consolidated memory retrieval following sleep deprivation in Drosophila.</a> Sleep. 35 (10), 1377-1384 (2012).</li><li>Placais, P. Y., Preat, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=To+favour+survival+under+food+shortage,+the+brain+disables+costly+memory.">To favour survival under food shortage, the brain disables costly memory.</a> Science. 339, 440-442 (2012).</li><li>Hirano, Y., 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=Fasting+launches+CRTC+to+faciltate+long-term+memory+formation+in+Drosophila.">Fasting launches CRTC to faciltate long-term memory formation in Drosophila.</a> Science. 339, 443-446 (2012).</li><li>Schroll, 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=Light-induced+activation+of+distinct+modulatory+neurons+triggers+appetitive+or+aversive+learning+in+Drosophila+larvae.">Light-induced activation of distinct modulatory neurons triggers appetitive or aversive learning in Drosophila larvae.</a> Curr Biol. 16, 1741-1747 (2006).</li><li>Claridge-Chang, A., 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=Writing+memories+with+light-addressable+reinforcement+circuitry.">Writing memories with light-addressable reinforcement circuitry.</a> Cell. 139, 405-415 (2009).</li><li>Aso, Y., 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=Three+dopamine+pathways+induce+aversive+odor+memories+with+different+stability.">Three dopamine pathways induce aversive odor memories with different stability.</a> PLoS Genetics. 8, e1002768 (2012).</li><li>Connolly, J. B., Tully, T., Roberts, D. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Drosophila: a Practical Approach. , 265-319 (1998).</li><li>Connolly, J. 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=Associative+learning+disrupted+by+impaired+Gs+signaling+in+Drosophila+mushroom+bodies.">Associative learning disrupted by impaired Gs signaling in Drosophila mushroom bodies.</a> Science. 274, 2104-2107 (1996).</li><li>Davis, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Traces+of+Drosophila+memory.">Traces of Drosophila memory.</a> Neuron. 70, 8-19 (2011).</li><li>Gu, H., O'Dowd, D. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cholinergic+synaptic+transmission+in+adult+Drosophila+kenyon+cells+in+situ.">Cholinergic synaptic transmission in adult Drosophila kenyon cells in situ.</a> J Neurosci. 26, 265-272 (2006).</li><li>Young, J. M., Wessnitzer, J., Armstrong, J. D., Webb, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Elemental+and+non-elemental+olfactory+learning+in+Drosophila.">Elemental and non-elemental olfactory learning in Drosophila.</a> Neurobiol Learn Mem. 96, 339-353 (2011).</li><li>Kaun, K. R., Azanchi, R., Maung, Z., Hirsh, J., Heberlein, U. .. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+model+for+alcohol+reward.">Drosophila model for alcohol reward.</a> Nat Neurosci. 14, 612-619 (2011).</li><li>Waddell, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dopamine+reveals+neural+circuit+mechanisms+of+fly+memory.">Dopamine reveals neural circuit mechanisms of fly memory.</a> Trends Neurosci. 33, 457-464 (2010).</li></ol>

作者什么都没有透露。

这里介绍的果蝇成虫嗅觉休克学习法允许的存储器不同阶段，包括长期存储器15-17的分子机制的分析。以及判定的昼夜节律18，睡眠19，饮食20,21，衰老11-13，神经变性疾病5和药物治疗5,6,19对存储器的影响。 许多强大的方法对神经回路的调解嗅觉记忆中的苍蝇3,4,7,11,16,27的功能成像最近研制。这些光遗传学技术使用在果蝇 14,16可在不同启动子的丰富剧目。这些启动子用于表达基因编码的钙和cAMP记者在存储器神经元16,27来研究记忆痕迹的特定基因突变的影响。 日成人E使用条件的推 ​​动者和突变使一个基因产物在内存3,4,6,7,13,14后发展作用的研究。成像和行为的方法可以与光与热激活通道来刺激或抑制不同的神经元中的存储器电路11,14,16,22-24进一步阐明其功能进行组合。此外，蘑菇体存储器的神经元可访问的全细胞膜片钳记录28和数学和计算技术被用来模拟果蝇嗅觉记忆29。 这些实验的进展，结合不同形式的介绍到这里联想记忆的协议，让果蝇被用于模拟发生在响应奖励，惩罚，激励，成瘾联想记忆的molecular-和电路级的变化，老龄化和疾病5,6,11-13,16,30-31。

这里介绍的方法是描述塔利和奎因的一些小的修改1。该实验是在两个阶段进行：苍蝇被训练在第一阶段中，和经过培训的蝇在第二阶段进行测试。在培训过程中，一组苍蝇都同时暴露异味1（CS +）和电击（美国）在训练管。苍蝇再收到异味2（CS - ）无触电。一个特定的气味与休克的这个单个的配对被称为1个周期的训练，这是最常用的气味为4-甲基环己醇（MCH）和3 - 辛醇（OCT）。 一个周期的训练导致的存储器不稳定相，可以长达7小时来检测的形成;然而，存储器通常立即进行测试，以确定什么是所谓学习，获得或2分钟的存储器。在30分钟或1小时测得的存储器被称为短期记忆，而3小时存储器被称为中期记忆。果蝇暴露于重复训练周期的训练周期（间隔训练）之间的差距导致了长期记忆的综合形式，CREB的转录依赖性，持续了一个星期。训练无间隙（集结训练）导致麻醉性存储器（ARM），这类似于长期记忆，通常测定24小时后的5个周期的训练7,13,15-17,20,21的形成。 用这种方法，可确定对存储器这些不同阶段的各种基因的突变的影响。中轻或温度敏感的转基因启动子驱动的表达来激活或阻断特定神经元的神经活动使人们得以观察其神经元所需要的存储器采集，整理和检索3,4,11,15,16,20， 22-24。在1小时记忆学习与年龄有关的记忆障碍，因为该F时，通常是衡量的存储器ORM出现特别容易老化11-13的影响。与存储器实验中进行全范围的行为和遗传控制的，例如，以确定性能缺陷是否是因为中央存储器缺陷或末梢感觉缺陷，可防止苍蝇从感测震动或嗅觉提示5 -7， 1 7，25,26。

<table border="1" cellpadding="0" cellspacing="0">
	<tbody>
		<tr>
			<td style="width:189px;height:28px;">Materials</td>
			<td style="width:139px;height:28px;">Source</td>
			<td style="width:116px;height:28px;">Cat. No.</td>
			<td></td>
		</tr>
		<tr>
			<td style="width:189px;height:20px;">3-Octanol</td>
			<td style="width:139px;height:20px;">Sigma</td>
			<td style="width:116px;height:20px;">218405</td>
			<td></td>
		</tr>
		<tr>
			<td style="width:189px;height:20px;">4-Methyl cyclohexanol</td>
			<td style="width:139px;height:20px;">Sigma</td>
			<td style="width:116px;height:20px;">15,309-5</td>
			<td></td>
		</tr>
		<tr>
			<td style="width:189px;height:20px;">Benzaldehyde</td>
			<td style="width:139px;height:20px;">Sigma</td>
			<td style="width:116px;height:20px;">418099</td>
			<td></td>
		</tr>
		<tr>
			<td style="width:189px;height:20px;">Mineral Oil</td>
			<td style="width:139px;height:20px;">Fluka</td>
			<td style="width:116px;height:20px;">BP2629-1</td>
			<td></td>
		</tr>
		<tr>
			<td style="width:189px;height:20px;">Hexyl acetate</td>
			<td style="width:139px;height:20px;">Sigma</td>
			<td style="width:116px;height:20px;">108154</td>
			<td></td>
		</tr>
		<tr>
			<td style="width:189px;height:20px;">Fructose</td>
			<td style="width:139px;height:20px;">Sigma</td>
			<td style="width:116px;height:20px;">F0127</td>
			<td></td>
		</tr>
		<tr>
			<td style="width:189px;height:22px;">Agarose</td>
			<td style="width:139px;height:22px;">Bioline</td>
			<td style="width:116px;height:22px;">BIO-41025</td>
			<td></td>
		</tr>
	</tbody>
</table>

drosophila adult olfactory shock learning

果蝇已经被用在古典制约实验了40多年，从而大大方便我们的记忆理解，包括参与认知1-7疾病的分子机制的阐明。学习和记忆所用幼虫来研究神经发育的基因8-10的效果与在果蝇进行测定，以测定的成年可塑性基因1-7的贡献。此外， 果蝇的寿命短有利于基因介导与年龄有关的记忆障碍5,11-13分析。该细分果蝇神经系统的许多诱导型启动子的可用性，能够确定何时以及在哪里所需的正常存储器以及所述加固信号3,4,14,16的不同方面的中继感兴趣的基因。 在成年果蝇学习记忆允许的详细分析行为和所涉及的电路和长期存储器15 -17的测定。成年阶段的长度容纳的存储器更长期的遗传，行为，食物和药物的操作，除了确定在存储器3-6,11-13,15-21衰老和神经变性疾病的效果。 经典条件是诱发的同时呈现中性气味线索（条件刺激，CS +）和增强的刺激， 如 ，触电或蔗糖，（无条件刺激，美国），即成为彼此的动物相关的1,16。第二个条件刺激（CS - ）随后提出没有美国。在测试阶段， 果蝇则同时出现在CS +和CS-异味。后果蝇设置时间的气味之间作出选择，这些动物的分布被记录下来。此过程人低点联想到不受先天偏爱无论是条件刺激的推出了一款偏向可靠计量的厌恶或食欲调节。各种控制实验还进行测试是否正常气味和加固单独所有基因型回应。

性能指数（PI）作为内存的措施。表1说明了一个有代表性的计算圆周率。 <table border="1" cellpadding="0" cellspacing="0"><tbody><tr><td> 妇幼保健搭配震撼 </td><td> 10月3日搭配震撼 </td></tr><tr><td>苍蝇避免妇幼保健院（华侨城管）= 80 苍蝇喜欢妇幼保健院（妇幼保健管）= 20 PI 1 - （80-20）/ 80 + 20） = 0.6 </td><td>苍蝇避免华侨城（妇幼保健管）= 75 苍蝇喜欢华侨城（华侨城管）= 25 PI 2 =（75-25）/（75 + 25） = 0.5 </td></tr><tr><td> PI实验=（0.6 +0.5）的/2=0.55 </td><td></td></tr></tbody></table> 表1代表性的计算用数据说明性能指标，性能指标在不同的实验可以比作澄清记忆效应。一旦这样的比较示于图3，其中载有一系列的实验以广š野生型成年果蝇（WT）和傻瓜学习突变成蝇1执行的结果。 10蛋白酶抑制剂的平均值被设置，与代表平均值的标准误差（SEM）的误差棒。这些结果表明， 傻瓜蝇显示在学习相比，野生型的降低。 <img alt="图1" fo:content-width="6in" fo:src="/files/ftp_upload/50107/50107fig1highres.jpg" src="/files/ftp_upload/50107/50107fig1.jpg" /> 图1：成人实验装置的苍蝇被训练和一件T迷宫测试。培训包括提出的气味与电击后无触电第二气味乙。后在中间腔室中的休息时间段的苍蝇都带有两个同时的气味。苍蝇被困两管中，并收集和计算，获得学习/记忆分数。 <img alt="图2" fo:content-width="6in" fo:src="/files/ftp_upload/50107/50107fig2highres.jpg" src="/files/ftp_upload/50107/50107fig2.jpg" /> 图2成人训练协议的苍蝇培养在两个步骤。其中蝇60秒接受的气味（CS +）配对的电击（US）的第一步。在接下来的步骤苍蝇收到第二气味（CS-）无触电。苍蝇，然后静置90秒，之后将被测试的CS +和CS-之间的选择。 <img alt="图3" fo:content-width="4in" src="/files/ftp_upload/50107/50107fig3highres.jpg" width="400" /> 图3示出了成年果蝇傻瓜和野生型学习的代表图。 </strong&gt; WT和下面的训练一次会议傻瓜苍蝇进行了测试。 傻瓜苍蝇显示在学习相比，WT组（n = 10）的减少。

Watch this Scientific Journal Video about Drosophila Adult Olfactory Shock Learning at JoVE.com

Drosophila Adult Olfactory Shock Learning

衡量成年果蝇相联存储器的方法进行说明。该测定是基于在飞行中对带有负的加强件（电击）的气味相关联，然后回想起这些信息在以后的时间，使被测量存储器的能力。

果蝇嗅觉成人学习的冲击

Drosophila have been used in classical conditioning experiments for over 40 years, thus greatly facilitating our understanding of memory, including the ...

drosophila-adult-olfactory-shock-learning

Research

JoVE Journal

Neuroscience

Drosophila Adult Olfactory Shock Learning

27.7K 次观看.  University of Bristol. 衡量成年果蝇相联存储器的方法进行说明。该测定是基于在飞行中对带有负的加强件（电击）的气味相关联，然后回想起这些信息在以后的时间，使被测量存储器的能力。 

视频: Drosophila Adult Olfactory Shock Learning

Assaying Locomotor, Learning, and Memory Deficits in Drosophila Models of Neurodegeneration

Advances in genetic methods have enabled the study of genes involved in human neurodegenerative diseases using Drosophila as a model system1. Most of these diseases, including Alzheimer's, Parkinson's and Huntington's disease are characterized by age-dependent deterioration in learning and memory functions and movement coordination2. Here we use behavioral assays, including the negative geotaxis assay3 and the aversive phototaxic suppression assay (APS assay)4,5, to show that some of the behavior characteristics associated with human neurodegeneration can be recapitulated in flies. In the negative geotaxis assay, the natural tendency of flies to move against gravity when agitated is utilized to study genes or conditions that may hinder locomotor capacities. In the APS assay, the learning and memory functions are tested in positively-phototactic flies trained to associate light with aversive bitter taste and hence avoid this otherwise natural tendency to move toward light. Testing these trained flies 6 hours post-training is used to assess memory functions. Using these assays, the contribution of any genetic or environmental factors toward developing neurodegeneration can be easily studied in flies.

Assaying Locomotor, Learning, and Memory Deficits in Drosophila Models of Neurodegeneration

Behavior assays for measuring locomotor functions, learning, and memory abilities in Drosophila.

含量的运动能力，学习和记忆的赤字果蝇</em神经退行性疾病>模型

Advances in genetic methods have enabled the study of genes involved in human neurodegenerative diseases using Drosophila as a model system1. Most of ...

科研

神经科学

Selective Viral Transduction of Adult-born Olfactory Neurons for Chronic in vivo Optogenetic Stimulation

Local interneurons are continuously regenerated in the olfactory bulb of adult rodents1-3. In this process, called adult neurogenesis, neural stem cells in the walls of the lateral ventricle give rise to neuroblasts that migrate for several millimeters along the rostral migratory stream (RMS) to reach and incorporate into the olfactory bulb. To study the different steps and the impact of adult-born neuron integration into preexisting olfactory circuits, it is necessary to selectively label and manipulate the activity of this specific population of neurons. The recent development of optogenetic technologies offers the opportunity to use light to precisely activate this specific cohort of neurons without affecting surrounding neurons4,5. Here, we present a series of procedures to virally express Channelrhodopsin2(ChR2)-YFP in a temporally restricted cohort of neuroblasts in the RMS before they reach the olfactory bulb and become adult-born neurons. In addition, we show how to implant and calibrate a miniature LED for chronic in vivo stimulation of ChR2-expressing neurons.

Selective Viral Transduction of Adult-born Olfactory Neurons for Chronic in vivo Optogenetic Stimulation

Adult-born neurons of the olfactory bulb can be optogenetically controlled using Channelrhodopsin2-expressing lentiviral injection in the rostral migratory stream and chronic photostimulation with an implanted miniature LED.

慢性成人出生的嗅觉神经元的选择性病毒转导在体内 Optogenetic刺激

Local interneurons are continuously regenerated in the olfactory bulb of adult rodents1-3. In this process, called adult neurogenesis, neural stem cells ...

Appetitive Associative Olfactory Learning in Drosophila Larvae

In the following we describe the methodological details of appetitive associative olfactory learning in Drosophila larvae. The setup, in combination with genetic interference, provides a handle to analyze the neuronal and molecular fundamentals of specifically associative learning in a simple larval brain.
Organisms can use past experience to adjust present behavior. Such acquisition of behavioral potential can be defined as learning, and the physical bases of these potentials as memory traces1-4. Neuroscientists try to understand how these processes are organized in terms of molecular and neuronal changes in the brain by using a variety of methods in model organisms ranging from insects to vertebrates5,6. For such endeavors it is helpful to use model systems that are simple and experimentally accessible. The Drosophila larva has turned out to satisfy these demands based on the availability of robust behavioral assays, the existence of a variety of transgenic techniques and the elementary organization of the nervous system comprising only about 10,000 neurons (albeit with some concessions: cognitive limitations, few behavioral options, and richness of experience questionable)7-10.
Drosophila larvae can form associations between odors and appetitive gustatory reinforcement like sugar11-14. In a standard assay, established in the lab of B. Gerber, animals receive a two-odor reciprocal training: A first group of larvae is exposed to an odor A together with a gustatory reinforcer (sugar reward) and is subsequently exposed to an odor B without reinforcement 9. Meanwhile a second group of larvae receives reciprocal training while experiencing odor A without reinforcement and subsequently being exposed to odor B with reinforcement (sugar reward). In the following both groups are tested for their preference between the two odors. Relatively higher preferences for the rewarded odor reflect associative learning - presented as a performance index (PI). The conclusion regarding the associative nature of the performance index is compelling, because apart from the contingency between odors and tastants, other parameters, such as odor and reward exposure, passage of time and handling do not differ between the two groups9.

Appetitive Associative Olfactory Learning in Drosophila Larvae

Drosophila larvae are able to associate odor stimuli with gustatory reward. Here we describe a simple behavioral paradigm that allows the analysis of appetitive associative olfactory learning.

欲求联想嗅觉学习<em&gt;果蝇</em&gt;幼虫

In the following we describe the methodological  details of appetitive associative olfactory learning in Drosophila larvae. The setup, in combination with ...

Dissection and Immunohistochemistry of Larval, Pupal and Adult Drosophila Retinas

The compound eye of Drosophila melanogaster consists of about 750 ommatidia (unit eyes). Each ommatidium is composed of about 20 cells, including lens-secreting cone cells, pigment cells, a bristle cell and eight photoreceptors (PRs) R1-R8 2. The PRs have specialized microvillar structures, the rhabdomeres, which contain light-sensitive pigments, the Rhodopsins (Rhs). The rhabdomeres of six PRs (R1-R6) form a trapezoid and contain Rh1 3 4. The rhabdomeres of R7 and R8 are positioned in tandem in the center of the trapezoid and share the same path of light. R7 and R8 PRs stochastically express different combinations of Rhs in two main subtypes5: In the 'p' subtype, Rh3 in pR7s is coupled with Rh5 in pR8s, whereas in the 'y' subtype, Rh4 in yR7s is associated with Rh6 in yR8s 6 7 8.
Early specification of PRs and development of ommatidia begins in the larval eye-antennal imaginal disc, a monolayer of epithelial cells. A wave of differentiation sweeps across the disc9 and initiates the assembly of undifferentiated cells into ommatidia10-11. The 'founder cell' R8 is specified first and recruits R1-6 and then R7 12-14. Subsequently, during pupal development, PR differentiation leads to extensive morphological changes 15, including rhabdomere formation, synaptogenesis and eventually rh expression.
In this protocol, we describe methods for retinal dissections and immunohistochemistry at three defined periods of retina development, which can be applied to address a variety of questions concerning retinal formation and developmental pathways. Here, we use these methods to visualize the stepwise PR differentiation at the single-cell level in whole mount larval, midpupal and adult retinas (Figure 1).

Dissection and Immunohistochemistry of Larval, Pupal and Adult Drosophila Retinas

The Drosophila retina is a crystal-like lattice composed of a small number of cell types that are generated in a stereotyped manner 1. Its amenability to sophisticated genetic analysis allows the study of complex developmental programs. This protocol describes dissections and immunohistochemistry of retinas at three discrete developmental stages, with a focus on photoreceptor differentiation.

幼虫，蛹和成人的解剖和免疫组化<em&gt;果蝇</em&gt;视网膜

The compound eye of Drosophila melanogaster consists of about 750 ommatidia (unit eyes). Each ommatidium is composed of about 20 cells, including ...

Whole Mount Immunolabeling of Olfactory Receptor Neurons in the Drosophila Antenna

Odorant molecules bind to their target receptors in a precise and coordinated manner. Each receptor recognizes a specific signal and relays this information to the brain. As such, determining how olfactory information is transferred to the brain, modifying both perception and behavior, merits investigation. Interestingly, there is emerging evidence that cellular transduction and transcriptional factors are involved in the diversification of olfactory receptor neuron. Here we provide a robust whole mount immunological labeling method to assay in vivo olfactory receptor neuron organization. Using this method, we identified all olfactory receptor neurons with anti-ELAV antibody, a known pan-neural marker and Or49a-mCD8::GFP, an olfactory receptor neuron specifically expressed in Nba neuron using anti-GFP antibody.

Whole Mount Immunolabeling of Olfactory Receptor Neurons in the Drosophila Antenna

Herein we describe the process of whole mount immunostaining of Drosophila antennae, which enables us to better understand the molecular mechanisms involved in the diversification of olfactory receptor neurons (ORN)s.

嗅觉受体神经元在全山免疫标记<em&gt;果蝇</em&gt;天线

Odorant molecules bind to their target receptors in a precise and coordinated manner. Each receptor recognizes a specific signal and relays this ...

Electrophysiological Method for Recording Intracellular Voltage Responses of Drosophila Photoreceptors and Interneurons to Light Stimuli In Vivo

Voltage responses of insect photoreceptors and visual interneurons can be accurately recorded with conventional sharp microelectrodes. The method described here enables the investigator to measure long-lasting (from minutes to hours) high-quality intracellular responses from single Drosophila R1-R6 photoreceptors and Large Monopolar Cells (LMCs) to light stimuli. Because the recording system has low noise, it can be used to study variability among individual cells in the fly eye, and how their outputs reflect the physical properties of the visual environment. We outline all key steps in performing this technique. The basic steps in constructing an appropriate electrophysiology set-up for recording, such as design and selection of the experimental equipment are described. We also explain how to prepare for recording by making appropriate (sharp) recording and (blunt) reference electrodes. Details are given on how to fix an intact fly in a bespoke fly-holder, prepare a small window in its eye and insert a recording electrode through this hole with minimal damage. We explain how to localize the center of a cell's receptive field, dark- or light-adapt the studied cell, and to record its voltage responses to dynamic light stimuli. Finally, we describe the criteria for stable normal recordings, show characteristic high-quality voltage responses of individual cells to different light stimuli, and briefly define how to quantify their signaling performance. Many aspects of the method are technically challenging and require practice and patience to master. But once learned and optimized for the investigator's experimental objectives, it grants outstanding in vivo neurophysiological data.

Electrophysiological Method for Recording Intracellular Voltage Responses of Drosophila Photoreceptors and Interneurons to Light Stimuli In Vivo

Sharp microelectrodes enable accurate electrophysiological characterization of photoreceptor and visual interneuron output in living Drosophila. Here we show how to use this method to record high-quality voltage responses of individual cells to controlled light stimulation. This method is ideal for studying neural information processing in insect compound eyes.

为记录细胞内电压响应电法<em&gt;果蝇</em&gt;光感受器和中间对光刺激<em&gt;在体内</em

Voltage responses of insect photoreceptors and visual interneurons can be accurately recorded with conventional sharp microelectrodes. The method ...

Olfactory Behaviors Assayed by Computer Tracking Of Drosophila in a Four-quadrant Olfactometer

A key challenge in neurobiology is to understand how neural circuits function to guide appropriate animal behaviors. Drosophila melanogaster is an excellent model system for such investigations due to its complex behaviors, powerful genetic techniques, and compact nervous system. Laboratory behavioral assays have long been used with Drosophila to simulate properties of the natural environment and study the neural mechanisms underlying the corresponding behaviors (e.g. phototaxis, chemotaxis, sensory learning and memory)1-3. With the recent availability of large collections of transgenic Drosophila lines that label specific neural subsets, behavioral assays have taken on a prominent role to link neurons with behaviors4-11. Versatile and reproducible paradigms, together with the underlying computational routines for data analysis, are indispensable for rapid tests of candidate fly lines with various genotypes. Particularly useful are setups that are flexible in the number of animals tested, duration of experiments and nature of presented stimuli. The assay of choice should also generate reproducible data that is easy to acquire and analyze. Here, we present a detailed description of a system and protocol for assaying behavioral responses of Drosophila flies in a large four-field arena. The setup is used here to assay responses of flies to a single olfactory stimulus; however, the same setup may be modified to test multiple olfactory, visual or optogenetic stimuli, or a combination of these. The olfactometer setup records the activity of fly populations responding to odors, and computational analytical methods are applied to quantify fly behaviors. The collected data are analyzed to get a quick read-out of an experimental run, which is essential for efficient data collection and the optimization of experimental conditions.

Olfactory Behaviors Assayed by Computer Tracking Of Drosophila in a Four-quadrant Olfactometer

We describe here a behavioral setup and data analysis method for assaying olfactory responses of up to 100 vinegar flies (Drosophila melanogaster). This system may be used with single or multiple olfactory stimuli, and adaptable for optogenetic activation or silencing of neuronal subsets.

通过计算机跟踪检测大鼠嗅觉行为<em&gt;果蝇</em&gt;在四象限嗅

A key challenge in neurobiology is to understand how neural circuits function to guide appropriate animal behaviors. Drosophila melanogaster is an ...

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 ...

Drosophila Courtship Conditioning As a Measure of Learning and Memory

Many insights into the molecular mechanisms underlying learning and memory have been elucidated through the use of simple behavioral assays in model organisms such as the fruit fly, Drosophila melanogaster. Drosophila is useful for understanding the basic neurobiology underlying cognitive deficits resulting from mutations in genes associated with human cognitive disorders, such as intellectual disability (ID) and autism. This work describes a methodology for testing learning and memory using a classic paradigm in Drosophila known as courtship conditioning. Male flies court females using a distinct pattern of easily recognizable behaviors. Premated females are not receptive to mating and will reject the male&#39;s copulation attempts. In response to this rejection, male flies reduce their courtship behavior. This learned reduction in courtship behavior is measured over time, serving as an indicator of learning and memory. The basic numerical output of this assay is the courtship index (CI), which is defined as the percentage of time that a male spends courting during a 10 min interval. The learning index (LI) is the relative reduction of CI in flies that have been exposed to a premated female compared to na&#239;ve flies with no previous social encounters. For the statistical comparison of LIs between genotypes, a randomization test with bootstrapping is used. To illustrate how the assay can be used to address the role of a gene relating to learning and memory, the pan-neuronal knockdown of Dihydroxyacetone phosphate acyltransferase (Dhap-at) was characterized here. The human ortholog of Dhap-at, glyceronephosphate O-acyltransferase (GNPT), is involved in rhizomelic chondrodysplasia punctata type 2, an autosomal-recessive syndrome characterized by severe ID. Using the courtship conditioning assay, it was determined that Dhap-at is required for long-term memory, but not for short-term memory. This result serves as a basis for further investigation of the underlying molecular mechanisms.

Drosophila Courtship Conditioning As a Measure of Learning and Memory

This protocol describes a Drosophila learning and memory assay called courtship conditioning. This classic assay is based on a reduction of male courtship behavior after sexual rejection by a non-receptive premated female. This natural form of behavioral plasticity can be used to test learning, short-term memory, and long-term memory.

果蝇调理作为学习和记忆的措施

Many insights into the molecular mechanisms underlying learning and memory have been elucidated through the use of simple behavioral assays in model ...

An Objective and Reproducible Test of Olfactory Learning and Discrimination in Mice

Olfaction is the predominant sensory modality in mice and influences many important behaviors, including foraging, predator detection, mating, and parenting. Importantly, mice can be trained to associate novel odors with specific behavioral responses to provide insight into olfactory circuit function. This protocol details the procedure for training mice on a Go/No-Go operant learning task. In this approach, mice are trained on hundreds of automated trials daily for 2&#8211;4 weeks and can then be tested on novel Go/No-Go odor pairs to assess olfactory discrimination, or be used for studies on how odor learning alters the structure or function of the olfactory circuit. Additionally, the mouse olfactory bulb (OB) features ongoing integration of adult-born neurons. Interestingly, olfactory learning increases both the survival and synaptic connections of these adult-born neurons. Therefore, this protocol can be combined with other biochemical, electrophysiological, and imaging techniques to study learning and activity-dependent factors that mediate neuronal survival and plasticity.

Here, we train mice on an associative learning task to test odor discrimination. This protocol also allows for studies on learning-induced structural changes in the brain.