Measurement of Carbon Dioxide Production from Radiolabeled Substrates in Drosophila melanogaster
Measurement of Carbon Dioxide Production from Radiolabeled Substrates in Drosophila melanogaster
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DOI：
10.3791/54045
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发表时间：
2016-06-01
影响因子：
1.2
通讯作者：
Bland, Michelle L.
中科院分区：
文献类型：
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作者：
Bland, Michelle L.
果蝇遗传学的力量正越来越多地被应用于激素信号和代谢的问题，以及在这种生物体中开发人类疾病模型。需要测量代谢率等参数的灵敏方法来推动对果蝇等小动物生理学和疾病的理解。本文描述的方法评估燃料氧化在少量成年苍蝇喂养的食物含有微量的C-14标记的底物，如葡萄糖或脂肪酸。在喂食期和任何额外的实验操作后，将苍蝇转移到用网盖的短管中，然后将其置于含有KOH饱和滤纸的玻璃小瓶中，该滤纸捕获放射性标记底物氧化为碳酸氢钾KHCO 3产生的呼出的放射性标记CO2。通过闪烁计数测量放射性标记的碳酸氢盐。这是一种定量的、可重复的、简单的研究燃料氧化的方法。使用放射性标记的葡萄糖、脂肪酸或氨基酸可以确定这些不同燃料源在不同条件下（如进食和禁食）以及不同遗传背景下对能量代谢的贡献。这补充了用于测量体内能量代谢的其他方法，并应进一步了解代谢调节。
The power of Drosophila genetics is increasingly being applied to questions of hormone signaling and metabolism and to the development of models of human disease in this organism. Sensitive methods for measurements of parameters such as metabolic rates are needed to drive the understanding of physiology and disease in small animals such as the fruit fly. The method described here assesses fuel oxidation in small numbers of adult flies fed food containing trace amounts of C-14-labeled substrates such as glucose or fatty acid. After the feeding period and any additional experimental manipulations, flies are transferred to short tubes capped with mesh, which are then placed in glass vials containing KOH-saturated filter paper that traps exhaled, radiolabeled CO2 generated from oxidation of radiolabeled substrates as potassium bicarbonate, KHCO3. This radiolabeled bicarbonate is measured by scintillation counting. This is a quantitative, reproducible, and simple approach for the study of fuel oxidation. The use of radiolabeled glucose, fatty acids, or amino acids allows determination of the contribution of these different fuel sources to energy metabolism under different conditions such as feeding and fasting and in different genetic backgrounds. This complements other approaches used to measure in vivo energy metabolism and should further the understanding of metabolic regulation.