Мы благодарим 4 анонимных рецензентов за их работу, чтобы улучшить рукопись. Мы благодарим Национальный центр Научных Исследований за финансовую поддержку в МБГ и Ю.Г. и Университет Бургундии и французское министерство исследований в MMS. Исследования в лаборатории Ю. Г. финансируется Европейского исследовательского совета (ERC Начиная Грант, GliSFCo-311403), Национальное агентство по де-ла-Recherche (ANR-JCJC, GGCB-2010), в Conseil Régional Бургундии (Faber), и CNRS.

1. Перед началом <ol><li> Используйте isogenized опорный запас подшипник в стабильных и надежных поведенческих фенотипов. Там не существует общего правила для выбора этого запаса, поскольку все потенциальные управления может нести неоднородного фона аллели. </li><li> Используйте этот штамм управления для обратного скрещивания все остальные акции, необходимые для более поздних стадий. Этот шаг возвратное скрещивание обычно представляется, по крайней мере 5 последовательных скрещиваний одной девственной самки (чтобы можно кроссинговер между гомологичными хромосомами) до 2-3 изогенных справочных мужчин 5. Этот шаг важен для гомогенизации генетический фон между различными акциями лету. </li><li> Поддержание Drosophila запасов на стандартную кукурузной муки (9%), дрожжи (10%), а также агаровой среде (1,5%), дополненной антибиотиками (0,4% метил пара-гидрокси-бензоат) в 12-час цикле свет / темнота в 25 ° C. </li><li> Достижение хемосенсорных экспериментов в контролируемой температурой комнаты (25 ° C) под дальнего красного света(Для устранения вклад визуальные подсказки, и сосредоточиться на хемосенсорных сигналов). Регулярно обновить воздух в помещении, чтобы проветрить помещение между каждого эксперимента. </li></ol> 2. Обонятельная отклика с помощью Y-лабиринт Пробирной <ol><li> Голодать в течение 16-18 мух ч при 25 ° С в стеклянных трубках, содержащих влажное бумажное полотенце перед испытанием. </li><li> Присоединяйтесь разъем Y-образный для двух стеклянных флаконах и в меньшей пластической флакона (загрузка флаконе). Используйте 1 мл пипетки советы, которые проходят через пены пробками связать разъем с тремя флаконах, и получить плотно закрытой Y-лабиринт. Обрежьте узкие концы двух наконечников пипеток (~ диаметр 2 мм, чтобы избежать любого возвращение лету, когда он сделал свое решение) с образованием двух &quot;ловушки&quot; флаконов, и большой конец одного наконечника пипетки, чтобы сформировать &quot;загрузка&quot; Трубка (фиг. 1A). </li><li> Просто перед подключением «ловушка» ампул (Рисунок 1б), поместите одну ~ 6 мм Диаметр фильтровальную бумагу вкаждый флакон. Добавить 40 мкл одоранта решения на одном фильтровальной бумаги, и 40 мкл соответствующий растворитель на втором фильтровальную бумагу. </li><li> Представьте десяти от 4 до 9 однодневных мух в &quot;загрузки&quot; флаконе. Не используйте CO 2 анестезии во время этого переноса, так как он оказывает сильное влияние на поведение 10. Скорее использовать краткое охлаждении на льду. Правильное манипулирование анестезированных мух важно ограничить нагрузку на субъектов, насколько это возможно. <ol><li> Выполните серию Y-лабиринт испытаний при 25 ° С под дальним красным светом (с использованием светодиодных ламп, чтобы ограничить возможный источник отопления), чтобы избежать визуальных стимулов как можно больше. Будьте осторожны, чтобы чередовать ориентацию на Y-лабиринты (одоранта, содержащий труб слева, или справа, и загрузочной трубы в передней или в задней части; Рисунок 1C). </li><li> Разрешить несколько часов для мух, чтобы войти в ловушки флакон, содержащий одоранта или растворитель. Всего мух после 24 часов, чтобы увеличить participatiна до более чем 80% и обеспечить максимальную обонятельный значение индекса (Симонне, личное сообщение). </li><li> Рассчитайте результате обонятельный индекс по следующей формуле: (количество в запаха трубки - число в трубке растворителя) / общее количество загруженных мух. </li></ol></li><li> Вымойте Y-лабиринт настройки следующим образом: смочите демонтированы настройку в RBS 35 MD ночь. Тщательно промыть проточной водой. Наконец промыть деионизированной водой и дайте высохнуть. </li></ol> 3. Статистический анализ данных <ol><li> Выполните т-тест, односторонний ANOVA или двусторонний ANOVA в зависимости от данных и переменных. </li></ol>

<ol>
	<li>Mombaerts, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genes+and+ligands+for+odorant,+vomeronasal+and+taste+receptors.">Genes and ligands for odorant, vomeronasal and taste receptors.</a> Nat. Rev. Neurosci. 5, 263-278 (2004).</li><li>Silbering, A. F., Benton, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ionotropic+and+metabotropic+mechanisms+in+chemoreception:+'chance+or+design.">Ionotropic and metabotropic mechanisms in chemoreception: 'chance or design.</a> EMBO Rep. 11, 173-179 (2010).</li><li>Ziegler, A. B., Berthelot-Grosjean, M., Grosjean, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+smell+of+love+in+Drosophila.">The smell of love in Drosophila.</a> Front. Physiol. 4, (2013).</li><li>Silbering, A. 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=Complementary+function+and+integrated+wiring+of+the+evolutionarily+distinct+Drosophila+olfactory+subsystems.">Complementary function and integrated wiring of the evolutionarily distinct Drosophila olfactory subsystems.</a> J. Neurosci. 31, 13357-13375 (2011).</li><li>Grosjean, 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=An+olfactory+receptor+for+food-derived+odours+promotes+male+courtship+in+Drosophila.">An olfactory receptor for food-derived odours promotes male courtship in Drosophila.</a> Nature. 478, 236-240 (2011).</li><li>Woodard, C., Huang, T., Sun, H., Helfand, S. L., Carlson, 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+analysis+of+olfactory+behavior+in+Drosophila:+a+new+screen+yields+the+ota+mutants.">Genetic analysis of olfactory behavior in Drosophila: a new screen yields the ota mutants.</a> Genetics. 123, 315-326 (1989).</li><li>Semmelhack, J. L., Wang, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Select+Drosophila+glomeruli+mediate+innate+olfactory+attraction+and+aversion.">Select Drosophila glomeruli mediate innate olfactory attraction and aversion.</a> Nature. 459, 218-223 (2009).</li><li>Budick, S. A., Dickinson, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Free-flight+responses+of+Drosophila+melanogaster+to+attractive+odors.">Free-flight responses of Drosophila melanogaster to attractive odors.</a> The Journal of Experimental Biology. 209, 3001-3017 (2006).</li><li>Martin, F., Charro, M. J., Alcorta, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mutations+affecting+the+cAMP+transduction+pathway+modify+olfaction+in+Drosophila.">Mutations affecting the cAMP transduction pathway modify olfaction in Drosophila.</a> J. Comp. Physiol. A. 187, 359-370 (2001).</li><li>Barron, A. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Anaesthetising+Drosophila+for+behavioural+studies.">Anaesthetising Drosophila for behavioural studies.</a> Journal of Insect Physiology. 46, 439-442 (2000).</li><li>Ai, M., 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=Acid+sensing+by+the+Drosophila+olfactory+system.">Acid sensing by the Drosophila olfactory system.</a> Nature. 468, 691-695 (2010).</li><li>Becher, P. G., Bengtsson, M., Hansson, B. S., Witzgall, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Flying+the+fly:+long-range+flight+behavior+of+Drosophila+melanogaster+to+attractive+odors.">Flying the fly: long-range flight behavior of Drosophila melanogaster to attractive odors.</a> J. Chem. Ecol. 36, 599-607 (2010).</li></ol>

Авторы не имеют ничего раскрывать.

Наш протокол Y-лабиринт основан на описанной выше протоколом 9. Тем не менее, мы вводим два основных отличия. Во-первых, мы используем узкие наконечники для предотвращения мух от возвращения, как только они решили вступить в пробирку, содержащую растворитель или растворитель плюс ?...

Хеморецепция через вкуса или обоняния является ключевым сенсорные модальности для выживания животных. Это дает жизненные сигналы, необходимые для обнаружения опасности или источников питания, а также для социальных взаимодействий. Он также помогает животным, чтобы найти партнера необходимую секс для их воспроизводства. На протяжении более 20 лет, интенсивные исследования, в том числе лауреат Нобелевской премии работы Ричард Аксель и Линда Бак в 2004 году &quot;за открытия, одоранта рецепторов и организации обонятельной системы&quot;, была проведена выявить молекулярные и клеточные основы обоняние 1,2. Один из любимых животных моделях для ученых рассекать обонятельный восприятие является Д. MELANOGASTER. Это насекомое похожая клеточной и молекулярной запах кодирования стратегию с млекопитающих. Научное сообщество использует различные поведенческие парадигмы, чтобы изучить роль отдушек в этой плодовой мухи. Эти тесты включают мультимодальные анализы, такие как courtshiр Тесты, где различные сенсорные модальности, в том числе обоняния, которые важны для выявления мужского ухаживания 3. Другие анализы также были разработаны для решения роль одорантов более конкретно; к ним относятся Т-лабиринты, Y-лабиринты, ловушки анализы, четыре полевых арен и аэродинамических трубах 4,5,6,7,8. В этой статье мы приведем простой модифицированный Y-лабиринт анализа, которая обеспечивает надежные обонятельные ответы с помощью D. MELANOGASTER. Наш набор использует конечных советы в отличие от ранее описанного метода 9. Таким образом, наше Y-лабиринт имеет два преимущества. Во-первых, он позволяет избежать возвращения в системе после того, как муха сделала свой выбор. Во-вторых, это ограничивает обмен пахучих веществ во всех областях Y-образном лабиринте. Этот последний преимуществом является важным, поскольку Drosophila очень чувствительны к воздушным потоком, которая часто используется, чтобы избежать одоранта насыщенность. Для регулировки экспериментальной установки с воздушным потоком будет время и стоимость трудоемким. Таким образом, наша Y-лабиринт анализ представляет собой эффициента и быстрый способ проверить обонятельный производительность дрозофилы.

<table border="0" cellpadding="0" cellspacing="0" width="835">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr>
			<td>Drosophila Polystyrene tube</td>
			<td>VWR europe</td>
			<td>734-2255</td>
			<td>30 x 25 mm&#160; 
			Y-maze</td>
		</tr>
		<tr>
			<td>Drosophila Borosilicate tube</td>
			<td>Dijon verre</td>
			<td></td>
			<td>95 X 25 mm 
			Y-maze</td>
		</tr>
		<tr>
			<td>Foam stopper</td>
			<td>Dutscher</td>
			<td>999038</td>
			<td> 
			Y-maze</td>
		</tr>
		<tr>
			<td>Y-shaped connector</td>
			<td>Europrix</td>
			<td>11020605</td>
			<td> 
			Y-maze</td>
		</tr>
		<tr>
			<td>100-1000&#181;l pipet tips</td>
			<td>Corning</td>
			<td>4868</td>
			<td>Join the following pipet tips to the Y-shaped connector. Cut 2 pipet tips at 65 mm from the wide end, and connect the narrow end (with a &#8764;2 mm opening) to 2 test vials. These openings will limit the U-turns once the flies enter the tubes containing the odors. Cut 1 pipet tip at 35 mm from the wide end, and connect it to the loading vial. 
			Y-maze</td>
		</tr>
		<tr>
			<td>Far-Red LED Bulb</td>
			<td>Rubin-Lacaque</td>
			<td>0RB180238</td>
			<td>625-630 nm</td>
		</tr>
		<tr>
			<td>Acetic Acid</td>
			<td>Sigma-Aldrich</td>
			<td>45725</td>
			<td></td>
		</tr>
		<tr>
			<td>Phenylacetic Acid</td>
			<td>Sigma-Aldrich</td>
			<td>P16621</td>
			<td></td>
		</tr>
		<tr>
			<td>Yeast</td>
			<td>Sensient Flavors Strasbourg</td>
			<td>1018880464</td>
			<td></td>
		</tr>
		<tr>
			<td>Cornmeal</td>
			<td>eurogerm</td>
			<td>Farine de ma&#239;s</td>
			<td></td>
		</tr>
		<tr>
			<td>Agar</td>
			<td>Kalys</td>
			<td>HP-697-25</td>
			<td></td>
		</tr>
		<tr>
			<td>Methyl hydroxy 4 benzoate</td>
			<td>VWR international</td>
			<td>25605293</td>
			<td></td>
		</tr>
	</tbody>
</table>

testing drosophila olfaction with a y-maze assay

Обнаружение сигналов из окружающей среды имеет важное значение для животных, чтобы обеспечить их выживание. С этой целью они используют экологические сигналы, такие как видение, mechanoreception, слуха и chemoperception через вкус, через прямой контакт или через обоняния, который представляет ответ на летучих молекулы действующего на большей дальности. Летучие химические молекулы очень важные сигналы для большинства животных в обнаружении опасности, источник питания, или для обмена данными между физическими лицами. Дрозофилы является одним из наиболее распространенных биологических моделей для ученых, чтобы исследовать клеточные и молекулярные основы обоняния. Чтобы подчеркнуть обонятельные способности этого маленького насекомого, мы описываем модифицированный выбор протокол, основанный на тесте на Y-лабиринт классически используется с мышами. Данные, полученные с Y-лабиринты дать ценную информацию, чтобы лучше понять, как животные иметь дело с их постоянно меняющихся условиях. Введем протокол шаг за шагом изучатьВоздействие пахучих веществ на лету разведочного ответ, используя эту Y-лабиринт анализа.

Рисунок 1D показывает два представительства ответы с помощью этой Y-лабиринт анализа. Canton-S самцы сильно избежать 10% уксусной кислоты, разбавленного дистиллированной водой, в то время как они существенно не избежать 10% фенилуксусной кислоты. Эти анализы основаны на 10 мужчинах в ?...

Watch this Scientific Journal Video about Testing Drosophila Olfaction with a Y-maze Assay at JoVE.com

Testing Drosophila Olfaction with a Y-maze Assay

Мы представляем тест выбор, чтобы выявить влияние пахучих веществ на Drosophila поведения с помощью Y-лабиринт анализа.

Тестирование<em&gt; Дрозофилы</em&gt; Обоняние с Y-лабиринт Пробирной

Detecting signals from the environment is essential for animals to ensure their survival. To this aim, they use environmental cues such as vision, ...

testing-drosophila-olfaction-with-a-y-maze-assay

Research

JoVE Journal

Neuroscience

Testing Drosophila Olfaction with a Y-maze Assay

14.8K Views.  UMR-6265 CNRS.  Мы представляем тест выбор, чтобы выявить влияние пахучих веществ на Drosophila поведения с помощью Y-лабиринт анализа. 

Video: Testing Drosophila Olfaction with a Y-maze Assay

Paired Nanoinjection and Electrophysiology Assay to Screen for Bioactivity of Compounds using the Drosophila melanogaster Giant Fiber System

Screening compounds for in vivo activity can be used as a first step to identify candidates that may be developed into pharmacological agents1,2. We developed a novel nanoinjection/electrophysiology assay that allows the detection of bioactive modulatory effects of compounds on the function of a neuronal circuit that mediates the escape response in Drosophila melanogaster3,4. Our in vivo assay, which uses the Drosophila Giant Fiber System (GFS, Figure 1) allows screening of different types of compounds, such as small molecules or peptides, and requires only minimal quantities to elicit an effect. In addition, the Drosophila GFS offers a large variety of potential molecular targets on neurons or muscles. The Giant Fibers (GFs) synapse electrically (Gap Junctions) as well as chemically (cholinergic) onto a Peripheral Synapsing Interneuron (PSI) and the Tergo Trochanteral Muscle neuron (TTMn)5. The PSI to DLMn (Dorsal Longitudinal Muscle neuron) connection is dependent on D&alpha;7 nicotinic acetylcholine receptors (nAChRs)6. Finally, the neuromuscular junctions (NMJ) of the TTMn and the DLMn with the jump (TTM) and flight muscles (DLM) are glutamatergic7-12. Here, we demonstrate how to inject nanoliter quantities of a compound, while obtaining electrophysiological intracellular recordings from the Giant Fiber System13 and how to monitor the effects of the compound on the function of this circuit. We show specificity of the assay with methyllycaconitine citrate (MLA), a nAChR antagonist, which disrupts the PSI to DLMn connection but not the GF to TTMn connection or the function of the NMJ at the jump or flight muscles.
Before beginning this video it is critical that you carefully watch and become familiar with the JoVE video titled "Electrophysiological Recordings from the Giant Fiber Pathway of D. melanogaster " from Augustin et al7, as the video presented here is intended as an expansion to this existing technique. Here we use the electrophysiological recordings method and focus in detail only on the addition of the paired nanoinjections and monitoring technique.

Paired Nanoinjection and Electrophysiology Assay to Screen for Bioactivity of Compounds using the Drosophila melanogaster Giant Fiber System

A rapid in vivo assay to test for neuromodulatory compounds using the Giant Fiber System (GFS) of Drosophila melanogaster is described. Nanoinjections in the head of the animal along with electrophysiological recordings of the GFS can reveal bioactivity of compounds on neurons or muscles.

Парные Nanoinjection и электрофизиологии Анализ для выявления биологической активности соединений с использованием DROSOPHILA MELANOGASTER Гигантские системы волоконно

Screening compounds for in vivo activity can be used as a first step to identify candidates that may be developed into pharmacological agents1,2. We ...

Methods to Assay Drosophila Behavior

Drosophila melanogaster, the fruit fly, has been used to study molecular mechanisms of a wide range of human diseases such as cancer, cardiovascular disease and various neurological diseases1. We have optimized simple and robust behavioral assays for determining larval locomotion, adult climbing ability (RING assay), and courtship behaviors of Drosophila. These behavioral assays are widely applicable for studying the role of genetic and environmental factors on fly behavior. Larval crawling ability can be reliably used for determining early stage changes in the crawling abilities of Drosophila larvae and also for examining effect of drugs or human disease genes (in transgenic flies) on their locomotion. The larval crawling assay becomes more applicable if expression or abolition of a gene causes lethality in pupal or adult stages, as these flies do not survive to adulthood where they otherwise could be assessed. This basic assay can also be used in conjunction with bright light or stress to examine additional behavioral responses in Drosophila larvae. Courtship behavior has been widely used to investigate genetic basis of sexual behavior, and can also be used to examine activity and coordination, as well as learning and memory. Drosophila courtship behavior involves the exchange of various sensory stimuli including visual, auditory, and chemosensory signals between males and females that lead to a complex series of well characterized motor behaviors culminating in successful copulation. Traditional adult climbing assays (negative geotaxis) are tedious, labor intensive, and time consuming, with significant variation between different trials2-4. The rapid iterative negative geotaxis (RING) assay5 has many advantages over more widely employed protocols, providing a reproducible, sensitive, and high throughput approach to quantify adult locomotor and negative geotaxis behaviors. In the RING assay, several genotypes or drug treatments can be tested simultaneously using large number of animals, with the high-throughput approach making it more amenable for screening experiments.

Methods to Assay Drosophila Behavior

Drosophila melanogaster is a genetically and behaviorally tractable model system that has been used to understand the molecular and cellular basis of many important biological processes for over a century 1. Drosophila has been well exploited to gain insights into the genetic basis of fly behavior.

Методы Анализ Drosophila Поведение

Drosophila melanogaster, the fruit fly, has been used to study molecular mechanisms of a wide range of human diseases such as cancer, cardiovascular ...

A Single-fly Assay for Foraging Behavior in Drosophila

For many animals, hunger promotes changes in the olfactory system in a manner that facilitates the search for appropriate food sources. In this video article, we describe an automated assay to measure the effect of hunger or satiety on olfactory dependent food search behavior in the adult fruit fly Drosophila melanogaster. In a light-tight box illuminated by red light that is invisible to fruit flies, a camera linked to custom data acquisition software monitors the position of six flies simultaneously. Each fly is confined to walk in individual arenas containing a food odor at the center. The testing arenas rest on a porous floor that functions to prevent odor accumulation. Latency to locate the odor source, a metric that reflects olfactory sensitivity under different physiological states, is determined by software analysis. Here, we discuss the critical mechanics of running this behavioral paradigm and cover specific issues regarding fly loading, odor contamination, assay temperature, data quality, and statistical analysis.

A Single-fly Assay for Foraging Behavior in Drosophila

In this video article, we describe an automated assay to measure the effect of hunger or satiety on olfactory dependent food search behavior in the adult fruit fly Drosophila melanogaster.

Одно-муха Анализ на нагула поведения в<em&gt; Дрозофилы</em

For many animals, hunger promotes changes in the olfactory system in a manner that facilitates the search for appropriate food sources. In this video ...

A Simple Behavioral Assay for Testing Visual Function in Xenopus laevis

Measurement of the visual function in the tadpoles of the frog, Xenopus laevis, allows screening for blindness in live animals. The optokinetic response is a vision-based, reflexive behavior that has been observed in all vertebrates tested. Tadpole eyes are small so the tail flip response was used as alternative measure, which requires a trained technician to record the subtle response. We developed an alternative behavior assay based on the fact that tadpoles prefer to swim on the white side of a tank when placed in a tank with both black and white sides. The assay presented here is an inexpensive, simple alternative that creates a response that is easily measured. The setup consists of a tripod, webcam and nested testing tanks, readily available in most Xenopus laboratories. This article includes a movie showing the behavior of tadpoles, before and after severing the optic nerve. In order to test the function of one eye, we also include representative results of a tadpole in which each eye underwent retinal axotomy on consecutive days. Future studies could develop an automated version of this assay for testing the vision of many tadpoles at once.

A Simple Behavioral Assay for Testing Visual Function in Xenopus laevis

Xenopus laevis tadpoles prefer swimming on the white side of a black/white tank. This behavior is guided by their vision. Based on this behavior, we present a simple assay to test the visual function of tadpoles.

Простой Поведенческая Анализ для тестирования зрительной функции в<em&gt; Xenopus Laevis</em

Measurement of the visual function in the tadpoles of the frog, Xenopus laevis, allows screening for blindness in live animals. The optokinetic response ...

Live Imaging of Drosophila Larval Neuroblasts

Stem cells divide asymmetrically to generate two progeny cells with unequal fate potential: a self-renewing stem cell and a differentiating cell. Given their relevance to development and disease, understanding the mechanisms that govern asymmetric stem cell division has been a robust area of study. Because they are genetically tractable and undergo successive rounds of cell division about once every hour, the stem cells of the Drosophila central nervous system, or neuroblasts, are indispensable models for the study of stem cell division. About 100 neural stem cells are located near the surface of each of the two larval brain lobes, making this model system particularly useful for live imaging microscopy studies. In this work, we review several approaches widely used to visualize stem cell divisions, and we address the relative advantages and disadvantages of those techniques that employ dissociated versus intact brain tissues. We also detail our simplified protocol used to explant whole brains from third instar larvae for live cell imaging and fixed analysis applications.

Live Imaging of Drosophila Larval Neuroblasts

This protocol details a streamlined method used to conduct live cell imaging in the context of an intact larval brain. Live cell imaging approaches are invaluable for the study of asymmetric neural stem cell divisions as well as other neurogenic and developmental processes, consistently uncovering mechanisms that were previously overlooked.

Живая съемка<em&gt; Дрозофилы</em&gt; Личиночные нейробласты

Stem cells divide asymmetrically to generate two progeny cells with unequal fate potential: a self-renewing stem cell and a differentiating cell. Given ...

Straightforward Assay for Quantification of Social Avoidance in Drosophila melanogaster

Drosophila melanogaster is an emerging model to study different aspects of social interactions. For example, flies avoid areas previously occupied by stressed conspecifics due to an odorant released during stress known as the Drosophila stress odorant (dSO). Through the use of the T-maze apparatus, one can quantify the avoidance of the dSO by responder flies in a very affordable and robust assay. Conditions necessary to obtain a strong performance are presented here. A stressful experience is necessary for the flies to emit dSO, as well as enough emitter flies to cause a robust avoidance response to the presence of dSO. Genetic background, but not their group size, strongly altered the avoidance of the dSO by the responder flies. Canton-S and Elwood display a higher performance in avoiding the dSO than Oregon and Samarkand strains. This behavioral assay will allow identification of mechanisms underlying this social behavior, and the assessment of the influence of genes and environmental conditions on both emission and avoidance of the dSO. Such an assay can be included in batteries of simple diagnostic tests used to identify social deficiencies of mutants or environmental conditions of interest.

Straightforward Assay for Quantification of Social Avoidance in Drosophila melanogaster

Here, we present a protocol to quantify the avoidance of stressed individuals. This paradigm is powerful yet user-friendly and can be used to assess the influence of genes and environment on one kind of social interaction in Drosophila melanogaster.

Простой Анализ для количественного определения социального избегания у<em&gt; Дрозофилы</em

Drosophila melanogaster is an emerging model to study different aspects of social interactions. For example, flies avoid areas previously occupied by ...

A Behavioral Assay for Mechanosensation of MARCM-based Clones in Drosophila melanogaster

Because of the structural and functional homology to the hair cells of the mammalian inner ear, the neurons that innervate the Drosophila external sense organs provide an excellent model system for the study of mechanosensation. This protocol describes a simple touch behavior in fruit flies which can be used to identify mutations that interfere with mechanosensation. The tactile stimulation of a macrochaete bristle on the thorax of flies elicits a grooming reflex from either the first or third leg. Mutations that interfere with mechanotransduction (such as NOMPC), or with other aspects of the reflex arc, can inhibit the grooming response. A traditional screen of adult behaviors would have missed mutants that have essential roles during development. Instead, this protocol combines the touch screen with mosaic analysis with a repressible cell marker (MARCM) to allow for only limited regions of homozygous mutant cells to be generated and marked by the expression of green fluorescent protein (GFP). By testing MARCM clones for abnormal behavioral responses, it is possible to screen a collection of lethal p-element mutations to search for new genes involved in mechanosensation that would have been missed by more traditional methods.

A Behavioral Assay for Mechanosensation of MARCM-based Clones in Drosophila melanogaster

In order to identify novel mutations affecting mechanosensation, we designed an assay that measures the behavioral response to tactile stimulation of fly bristles in mutant clones generated by the MARCM method. The combination of techniques allows for the identification of mechanosensitive mutations that would otherwise be lethal.

Поведенческая Анализ на Mechanosensation из MARCM основе Клоны в<em&gt; Дрозофилы</em

Because of the structural and functional homology to the hair cells of the mammalian inner ear, the neurons that innervate the Drosophila external sense ...

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.

Вкус Предпочтение анализа для взрослых<em&gt; Drosophila</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 ...

The CApillary FEeder Assay Measures Food Intake in Drosophila melanogaster

For most animals, feeding is an essential behavior for securing survival, and it influences development, locomotion, health and reproduction. Ingestion of the right type and quantity of food therefore has a major influence on quality of life. Research on feeding behavior focuses on the underlying processes that ensure actual feeding and unravels the role of factors regulating internal energy homeostasis and the neuronal bases of decision-making. The model organism Drosophila melanogaster, with its great variety of genetically traceable tools for labeling and manipulating single neurons, allows mapping of neuronal networks and identification of molecular signaling cascades involved in the regulation of food intake. This report demonstrates the CApillary FEeder assay (CAFE) and shows how to measure food intake in a group of flies for time spans ranging from hours to days. This easy-to-use assay consists of glass capillaries filled with liquid food that flies can freely access and feed on. Food consumption in the assay is accurately determined using simple measurement tools. Herein we describe step-by-step the method from setup to successful execution of the CAFE assay, and provide practical examples to analyze the food intake of a group of flies under controlled conditions. The reader is guided through possible limitations of the assay, and advantages and disadvantages of the method compared to other feeding assays in D. melanogaster are evaluated.

The CApillary FEeder Assay Measures Food Intake in Drosophila melanogaster

The CApillary FEeder (CAFE) assay is a simple, budget-friendly, highly reliable method for investigating mechanisms underlying food intake. Used with the highly versatile genetic model organism Drosophila melanogaster, it provides a powerful means of gaining new insights into regulatory mechanisms of food intake.

Капиллярных фидерных Анализ мер по обеспечению продовольственной приема внутрь<em&gt; Дрозофилы</em

For most animals, feeding is an essential behavior for securing survival, and it influences development, locomotion, health and reproduction. Ingestion of ...

Novel Assay for Cold Nociception in Drosophila Larvae

How organisms sense and respond to noxious temperatures is still poorly understood. Further, the mechanisms underlying sensitization of the sensory machinery, such as in patients experiencing peripheral neuropathy or injury-induced sensitization, are not well characterized. The genetically tractable Drosophila model has been used to study the cells and genes required for noxious heat detection, which has yielded multiple conserved genes of interest. Little is known however about the cells and receptors important for noxious cold sensing. Although, Drosophila does not survive prolonged exposure to cold temperatures (&#8804;10 &#186;C), and will avoid cool, preferring warmer temperatures in behavioral preference assays, how they sense and possibly avoid noxious cold stimuli has only recently been investigated.
Here we describe and characterize the first noxious cold (&#8804;10 &#186;C) behavioral assay in Drosophila. Using this tool and assay, we show an investigator how to qualitatively and quantitatively assess cold nociceptive behaviors. This can be done under normal/healthy culture conditions, or presumably in the context of disease, injury or sensitization. Further, this assay can be applied to larvae selected for desired genotypes, which might impact thermosensation, pain, or nociceptive sensitization. Given that pain is a highly conserved process, using this assay to further study&#160;thermal nociception will likely glean important understanding of pain processes in other species, including vertebrates.

Novel Assay for Cold Nociception in Drosophila Larvae

Here we demonstrate a novel assay to study cold nociception in Drosophila larvae. This assay utilizes a custom-built Peltier probe capable of applying a focal noxious cold stimulus and results in quantifiable cold-specific behaviors. This technique will allow further cellular and molecular dissection of cold nociception.

Новый анализ для холодной ноцицепции в<em&gt; Drosophila</em&gt; личинки

How organisms sense and respond to noxious temperatures is still poorly understood. Further, the mechanisms underlying sensitization of the sensory ...