Department of Cell Biology, Neurobiology, and Anatomy and Medical College of Wisconsin, Milwaukee, Wisconsin, USA.
Department of Ophthalmology and Visual Sciences, Medical College of Wisconsin, Milwaukee, Wisconsin, USA.
J Ocul Pharmacol Ther. 2023 Oct;39(8):563-571. doi: 10.1089/jop.2023.0048. Epub 2023 Jul 24.
Penetrating eye injuries commonly cause permanent loss of vision in patients. Unlike mammals, zebrafish can regenerate both damaged tissue and severed axons in the central nervous system. Here, we present a tractable adult zebrafish model to study intraocular axon regeneration after penetrating eye injury. To create consistent penetrating intraocular injuries, pins of standardized diameters were inserted into the eye through the cornea and penetrating the retina but not the underlying sclera. Transgenic g reporter fish were used to preferentially label retinal ganglion cells (RGCs) that respond to injury with regenerating axons. Retinas were fixed and flat mounted at various times postinjury to examine injury size, number of green fluorescent protein (GFP)-positive cells and axons, axonal varicosities, and rate of regeneration to the optic nerve head. Intraocular injection of colchicine was used to inhibit axon outgrow as a proof of principle that this method can be used to screen effects of pharmacological agents on intraocular axon regeneration. Penetrating injury to the zebrafish retina results in robust axon regeneration by RGCs around and beyond the site of injury. The transgene allows visualization of individual or small bundles of axons with varicosities and growth cones easily observable. Regeneration proceeded with most, if not all, axons reaching the optic nerve head by 3-day postinjury. A single intraocular injection of colchicine a day after injury was sufficient to delay axon regeneration at 2-days postinjury. Surprisingly, we identified a stereotypically located population of circumferential projecting neurons within the retina that upregulate expression after injury. Penetrating injury to the adult transgenic zebrafish eye is a model of successful intraocular axon regeneration. The pharmacological and genetic tools available for this organism should make it a powerful tool for dissecting the cellular, molecular, and genetic mechanisms of axon regeneration in the intraocular environment.
穿透性眼外伤常导致患者视力永久丧失。与哺乳动物不同，斑马鱼可以在中枢神经系统中再生受损组织和切断的轴突。在这里，我们提出了一种可行的成年斑马鱼模型，用于研究穿透性眼外伤后的眼内轴突再生。为了创建一致的穿透性眼内损伤，将标准化直径的别针通过角膜插入眼睛，穿透视网膜但不穿透下面的巩膜。使用转基因 g 报告鱼优先标记对再生轴突有反应的视网膜神经节细胞（RGC）。在受伤后不同时间固定和平面安装视网膜，以检查损伤大小、绿色荧光蛋白（GFP）阳性细胞和轴突的数量、轴突膨体以及向视神经头的再生速度。眼内注射秋水仙碱抑制轴突生长，证明该方法可用于筛选药物对眼内轴突再生的影响。 穿透性损伤斑马鱼视网膜会导致 RGC 产生强烈的轴突再生，超出和超出损伤部位。该转基因允许可视化单个或小束轴突，其膨体和生长锥很容易观察到。大多数（如果不是全部）轴突在受伤后 3 天到达视神经头，再生过程就会进行。受伤后第 1 天单次眼内注射秋水仙碱足以在受伤后第 2 天延迟轴突再生。令人惊讶的是，我们在视网膜内鉴定出一个位置固定的环状投射神经元群体，它们在受伤后上调表达。 穿透性损伤成年转基因斑马鱼眼睛是眼内轴突成功再生的模型。该生物体可用的药理学和遗传学工具应该使其成为解析眼内环境中轴突再生的细胞、分子和遗传机制的有力工具。
