Chuckowree Jyoti A, Vickers James C
Discipline of Pathology, Faculty of Health Sciences, University of Tasmania, Hobart, Tasmania, 7000, Australia.
J Neurosci. 2003 May 1;23(9):3715-25. doi: 10.1523/JNEUROSCI.23-09-03715.2003.
We examined the cytoskeletal dynamics that characterize neurite sprouting after axonal injury to cortical neurons maintained in culture for several weeks and compared these with initial neurite development. Cultured neocortical neurons, derived from embryonic day 18 rats, were examined at 3 d in vitro (DIV) and at various time points after axotomy at 21 DIV. The postinjury neuritic response was highly dynamic, progressing through an initial phase of retraction, followed by substantial axonal sprouting within 4-6 hr. Postinjury sprouts were motile and slender with expanded growth cone-like end structures. Microtubule markers were localized to sprout shafts and the proximal regions of putative growth cones and filamentous actin was distributed throughout growth cones, whereas neurofilament proteins were restricted to sprout shafts. A similar distribution of cytoskeletal proteins was present in developing neurites at 3 DIV. Exposure of developing and mature, injured cultures to the microtubule stabilizing agent taxol (10 microg/ml) caused growth inhibition, process distension, the transformation of growth cones into bulbous structures, and abnormal neurite directionality. Microtubule and neurofilament segregation occurred after taxol exposure in developing neurites and postinjury sprouts. Exposure to the microtubule destabilizing agent nocodazole (100 microg/ml) resulted in substantial morphological alteration of developing neurons and inhibited neurite growth and postinjury axonal sprouting. Our results indicate that the axons of cortical neurons have an intrinsic ability to sprout after transection, and similar cytoskeletal dynamics underlie neurite development and postinjury axonal sprouting.
我们研究了培养数周的皮质神经元轴突损伤后神经突萌发所特有的细胞骨架动力学，并将其与最初的神经突发育进行了比较。对源自胚胎第18天大鼠的培养新皮质神经元在体外培养3天（DIV）时以及在21 DIV进行轴突切断后的不同时间点进行了检查。损伤后的神经突反应具有高度动态性，经历了一个最初的回缩阶段，随后在4 - 6小时内出现大量轴突发芽。损伤后的芽是可移动的且细长，末端有类似生长锥的扩张结构。微管标记物定位于芽轴和假定生长锥的近端区域，丝状肌动蛋白分布于整个生长锥，而神经丝蛋白则局限于芽轴。在3 DIV时发育中的神经突中也存在类似的细胞骨架蛋白分布。将发育中的和成熟的、受伤的培养物暴露于微管稳定剂紫杉醇（10微克/毫升）会导致生长抑制、突起扩张、生长锥转变为球茎状结构以及神经突方向性异常。在发育中的神经突和损伤后的芽中，紫杉醇处理后微管和神经丝发生了分离。将其暴露于微管去稳定剂诺考达唑（100微克/毫升）会导致发育中的神经元出现显著的形态改变，并抑制神经突生长和损伤后的轴突发芽。我们的结果表明，皮质神经元的轴突在横断后具有内在的发芽能力，并且类似的细胞骨架动力学是神经突发育和损伤后轴突发芽的基础。
