Background

Among animal phyla, some of the least studied nervous systems are those of the phylum Echinodermata. Part of the problem lies in that most of their nervous components are embedded in the body wall that has calcareous skeletal components.

New method

We have developed a novel technique for the successful isolation of the radial nerve cords (RNCs) and an in vitro system where the isolated RNCs can be cultured and are amenable to experimental manipulation. Here we use this system to isolate the RNC of the sea cucumber Holothuria glaberrima as a way to extend our studies on its regeneration capabilities.

Results

The RNCs can be isolated from the surrounding tissues by collagenase treatment. The explants obtained following enzymatic dissociation can be kept in culture for up to 2 weeks. Histological and immunohistochemical studies show that the explants maintain a stable number of cells with little proliferation or apoptosis throughout the culture incubation period. The main change observed in RNCs in vitro is a progressive dedifferentiation of radial glia-like cells. This dedifferentiation corresponds to the first step in the regeneration response to injury that has been described in vivo.

Comparison with existing methods

There are no existing methods to isolate and culture echinoderm radial nerve cord.

Conclusions

The described protocol provides a unique tool to obtain easily accessible RNC from holothurians to perform cellular, biochemical, and genomic experiments in the echinoderm nervous system without interference of adjacent tissues. The technique provides a unique opportunity to study the dedifferentiation response associated with the regeneration of the nervous system in echinoderms.

中文翻译：

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在海参桡神经索的体外外植体中观察到放射状胶质细胞样细胞的去分化

背景

在动物门中，一些研究最少的神经系统是棘皮动物门的神经系统。部分问题在于它们的大部分神经成分都嵌在具有钙质骨骼成分的体壁中。

新方法

我们开发了一种成功分离桡神经索 (RNC) 的新技术，以及一种可以培养分离的 RNC 并适合实验操作的体外系统。在这里，我们使用该系统分离海参Holothuria glaberrima的 RNC，作为扩展我们对其再生能力研究的一种方式。

结果

RNCs 可以通过胶原酶处理从周围组织中分离出来。酶促解离后获得的外植体可在培养物中保存长达 2 周。组织学和免疫组织化学研究表明，外植体在整个培养孵育期间保持稳定的细胞数量，几乎没有增殖或凋亡。在体外RNC 中观察到的主要变化是放射状胶质细胞样细胞的进行性去分化。这种去分化对应于体内描述的损伤再生反应的第一步。

与现有方法的比较

目前尚无分离培养棘皮动物桡神经索的方法。

结论

所描述的协议提供了一种独特的工具，可以从海参中获取易于访问的 RNC，以便在棘皮动物神经系统中进行细胞、生化和基因组实验，而不会干扰相邻组织。该技术提供了一个独特的机会来研究与棘皮动物神经系统再生相关的去分化反应。