测量并改变配套驱动马累<em>果蝇</em
Summary
本文介绍了采用果蝇 melanogaste R以学习动机男性交配驱动器中的行为分析。利用这种方法，研究人员可以利用先进的飞神经源性技术来揭开背后这个动机的遗传，分子和细胞机制。
Abstract
尽管几十年的调查，动机状态的神经细胞和分子基础仍然神秘。我们最近开发了一种新型，还原，和可扩展的系统使用雄性果蝇 （果蝇）的交配驱动动机进行了深入调查，采用何种方法在这里，我们的细节。行为模式集中在雄性交配驱动器一起生育过多次交配的过程中减少并恢复了〜3天的发现。在这个系统中，在飞可用的有力神经源性工具收敛与遗传访问性和可用于性行为推定接线图。这种融合允许快速隔离和小神经元群体具有特定功能的动机审讯。在这里，我们详细地是，用于测量和在雄性苍蝇改变求偶动机的饱足感测定的设计和执行。使用此实验中，我们还表明，男性低交配驱动器可通过刺激多巴胺能神经元加以克服。饱腹法简单，价格适中，和强大的遗传背景的影响。我们预计饱腹感检测，以产生许多新的见解激励状态的神经生物学。
Introduction
在果蝇的工作提供了深刻的开拓洞察许多生物现象，包括基因1的性质，胚胎发育2，昼夜节律3，和神经系统4，5，6的发展和布线的原则。动机仍然因为上迄今已研究了系统的限制可能远不如比这些现象可知，。动机在飞主要研究了饥饿，它提出了许多挑战的背景下，由于每次喂食发作和外骨骼这就排除脂肪沉积明显的迹象了微乎其微的食物摄入量。因此，有必要扩大用于研究动机在飞的系统。
我们描述一个行为框架，配合驱动器的研究果蝇 。该系统以在飞以及辅助7，8，9，10，11，12和其性二态电路8,13的推定连接组优势的神经发生的工具。此外，许多先天的14，15，16，17，18，19，20，21和22的教训，23，24感觉运动电路控制求爱已详细制定出来，提供了一个难得的机遇找到赖以动机撞击的精确电路节点。我们最近报道，在飞，如在人类中，多巴胺水平是中央交配驱动装置25，26，27。我们已经获得了相关产生多巴胺和飞接受神经元，促进详细molecular-和电路级的遗传访问使用试验，我们在这里介绍25这种现象的保守分析。
我们添加到张某等人的行为分析。 25一个新的平行为竞技场，允许视频得分，我们称之为一个2维（2-D）的饱足感测定中，在以前的方法的一个重要的改进。因此，新的检测更可扩展性和可量化的，因此更适合参与积极性的基因和神经元的遗传筛选。我们使用这种新的检测，加上求爱分析和neurogeNETIC操作，演示了如何衡量和改变苍蝇交配的车程。
Protocol
备注：本协议描述的准备（第1 – 3），执行（第4节），和分析的2-D饱足感检测（第4节）。然后，用多巴胺能刺激作为一个例子，第5显示了如何thermogenetic刺激的2-D饱足测定结合来诱导亢进。第6节说明3种方式来验证的2-D饱足感测定的结果。最后，第7展示了如何衡量雄蝇交配驱动器的恢复。 1.编造8位和32室行为阿里纳斯注意：每个行为竞技场包括在每个四?…
Representative Results
为了描述果蝇交配的车程，3日龄，广州WT-S男性被在2-D饱法检测。过测定（4.5小时）的过程中，雄性交配的4.8±0.3的平均（平均均值，SEM±标准误差）次。交配发起大多在最初2小时（78％）（ 图6A，6B）和变得较不频繁的测定法的进展（ 图6A，6B）。这种减少不是由于缺乏配合伙伴的（74％的女性在整个试验保持未配合）或肉体疲劳（ 图…
Discussion
动机状态可以满足，维护，并回收34。我们提出的2-D饱足测定迅速，鲁棒测量所有的在飞配合驱动器的这些方面。该测定中开辟了采用先进的飞基因操作来研究动机的行为的分子和电路元件的可能性。
饱腹感检测依赖于男性的能力，成功地法院和交配，并在适当的时候终止交配。虽然hyposexual苍蝇少法庭，低求爱苍蝇不一定hyposexual;他们可能，例如，具有难以…
Disclosures
The authors have nothing to disclose.
Acknowledgements
The authors thank Mike Crickmore, Dragana Rogulja, and Michelle Frank for comments on the manuscript. Pavel Gorelik provided technical support for manufacturing the behavioral arenas. This work was conducted in Mike Crickmore’s lab and is also supported by the Whitehall Foundation (Principal Investigator: Dragana Rogulja). S.X.Z. is a Stuart H.Q. and Victoria Quan Fellow at Harvard Medical School.
Materials
| 1/16 inch clear acrylic | McMaster-Carr | 8589K12 | Used to make arenas; see Supplemental Material 1 for designs. |
| 1/8 inch clear acrylic | McMaster-Carr | 8589K42 | Used to make arenas; see Supplemental Material 1 for designs. |
| 3/16 inch clear acrylic | McMaster-Carr | 8560K219 | Used to make arenas; see Supplemental Material 1 for designs. |
| 1/32 inch black delrin | McMaster-Carr | 8575K132 | Used to make arenas; see Supplemental Material 1 for designs. |
| Hex screws, 1 inch long (50x) | McMaster-Carr | 92314A115 | Used to make arenas. Can be replaced by 3/4 inch screws (92314A113, McMaster-Carr) for 32-chamber arenas. |
| Thumb nuts (25x) | McMaster-Carr | 92741A100 | Used to make arenas. Can be replaced by regular hex nuts (90480A005, McMaster-Carr). |
| Camcorder | Canon | Vixia HF R700 | Can be replaced by any consumer comcorder. |
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Cite This Article
Boutros, C. L., Miner, L. E., Mazor, O., Zhang, S. X. Measuring and Altering Mating Drive in Male Drosophila melanogaster. J. Vis. Exp. (120), e55291, doi:10.3791/55291 (2017).