Here, we report the findings after application of neurobiotin tracing to pallial and stellar nerves in the mantle of the cephalopod mollusk Octopus vulgaris and to the axial nerve cord in its arm. Neurobiotin backfilling is a known technique in other molluscs, but it is applied to octopus for the first time to be best of our knowledge. Different neural tracing techniques have been carried out in cephalopods to study the intricate neural connectivity of their nervous system, but mapping the nervous connections in this taxon is still incomplete, mainly due to the absence of a reliable tracing method allowing whole-mount imaging. In our experiments, neurobiotin backfilling allowed: (1) imaging of large/thick samples (larger than 2 mm) through optical clearing; (2) additional application of immunohistochemistry on the backfilled tissues, allowing identification of neural structures by coupling of a specific antibody. This work opens a series of future studies aimed to the identification of the neural diagram and connectome of octopus nervous system.

在这里，我们报告了应用神经生物素追踪头足类软体动物普通章鱼外套膜和星状神经及其手臂轴神经索后的发现。神经生物素回填在其他软体动物中是一种已知的技术，但据我们所知，它首次应用于章鱼。人们已经在头足类动物中进行了不同的神经追踪技术，以研究其神经系统复杂的神经连接，但绘制该分类单元中的神经连接图谱仍然不完整，这主要是由于缺乏允许整体成像的可靠追踪方法。在我们的实验中，神经生物素回填允许：（1）通过光学透明对大/厚样品（大于2毫米）进行成像； (2)在回填组织上额外应用免疫组织化学，通过特异性抗体的偶联来识别神经结构。这项工作开启了一系列旨在识别章鱼神经系统的神经图和连接组的未来研究。