Chandel Ishita, Baker Ryan, Nakamura Naosuke, Panin Vlad
Department of Biochemistry and Biophysics, Texas A&M University.
Department of Biochemistry and Biophysics, Texas A&M University;
J Vis Exp. 2019 Jul 9(149). doi: 10.3791/59404.
Coordinated muscle contractions are a form of rhythmic behavior seen early during development in Drosophila embryos. Neuronal sensory feedback circuits are required to control this behavior. Failure to produce the rhythmic pattern of contractions can be indicative of neurological abnormalities. We previously found that defects in protein O-mannosylation, a posttranslational protein modification, affect the axon morphology of sensory neurons and result in abnormal coordinated muscle contractions in embryos. Here, we present a relatively simple method for recording and analyzing the pattern of peristaltic muscle contractions by live imaging of late stage embryos up to the point of hatching, which we used to characterize the muscle contraction phenotype of protein O-mannosyltransferase mutants. Data obtained from these recordings can be used to analyze muscle contraction waves, including frequency, direction of propagation and relative amplitude of muscle contractions at different body segments. We have also examined body posture and taken advantage of a fluorescent marker expressed specifically in muscles to accurately determine the position of the embryo midline. A similar approach can also be utilized to study various other behaviors during development, such as embryo rolling and hatching.
协调性肌肉收缩是果蝇胚胎发育早期出现的一种节律性行为形式。需要神经元感觉反馈回路来控制这种行为。无法产生有节律的收缩模式可能表明存在神经学异常。我们之前发现，蛋白质O-甘露糖基化（一种翻译后蛋白质修饰）缺陷会影响感觉神经元的轴突形态，并导致胚胎中出现异常的协调性肌肉收缩。在这里，我们提出了一种相对简单的方法，通过对晚期胚胎直至孵化点进行实时成像来记录和分析蠕动性肌肉收缩模式，我们用该方法来表征蛋白质O-甘露糖基转移酶突变体的肌肉收缩表型。从这些记录中获得的数据可用于分析肌肉收缩波，包括频率、传播方向以及不同身体节段肌肉收缩的相对幅度。我们还检查了身体姿势，并利用在肌肉中特异性表达的荧光标记物来准确确定胚胎中线的位置。类似的方法也可用于研究发育过程中的各种其他行为，如胚胎翻滚和孵化。
