Department of Biological Sciences, Purdue University, West Lafayette, IN, USA.
Weldon School of Biomedical Engineering, Purdue University, West Lafayette, IN, USA.
Methods Mol Biol. 2024;2831:265-282. doi: 10.1007/978-1-0716-3969-6_18.
Actin flow refers to the motion of the F-actin cytoskeleton and has been observed in many different cell types, especially in motile cells including neuronal growth cones. The direction of the actin flow is generally retrograde from the periphery toward the center of the cell. Actin flow can be harnessed for forward movement of the cell through substrate-cytoskeletal coupling; thus, a key function of actin flow is in cell locomotion. In this chapter, we illustrate three different methods of quantifying retrograde F-actin flow in growth cones derived from cultured Aplysia bag cell neurons. These methods include tracking the movement of surface marker beads as well as kymograph analysis of time-lapse sequences acquired by differential interference contrast (DIC) imaging or fluorescent speckle microscopy (FSM). Due to their large size, Aplysia neuronal growth cones are uniquely suited for these methods; however, they can also be applied to any other growth cones with clear F-actin-rich peripheral domains.
肌动蛋白流是指 F-肌动蛋白细胞骨架的运动，在许多不同的细胞类型中都有观察到，特别是在运动细胞中，包括神经元生长锥。肌动蛋白流的方向通常是从细胞的外围向中心逆行。肌动蛋白流可以通过基质-细胞骨架耦联来实现细胞的向前运动；因此，肌动蛋白流的一个关键功能是在细胞运动中。在本章中，我们展示了三种不同的方法来量化培养的 Aplysia 袋状神经元衍生的生长锥中的逆行 F-肌动蛋白流。这些方法包括跟踪表面标记珠的运动，以及通过相差差示干涉对比 (DIC) 成像或荧光散斑显微镜 (FSM) 获取的延时序列的动图分析。由于其体积较大，Aplysia 神经元生长锥非常适合这些方法；然而，它们也可以应用于任何具有清晰的富含 F-肌动蛋白的外围区域的其他生长锥。
