While the role of cholinergic neurotransmission from motoneurons is well established during neuromuscular development, whether it regulates central nervous system development in the spinal cord is unclear. Zebrafish presents a powerful model to investigate how the cholinergic system is set up and evolves during neural circuit formation. In this study, we carried out a detailed spatiotemporal analysis of the cholinergic system in embryonic and larval zebrafish. In 1-day-old embryos, we show that spinal motoneurons express presynaptic cholinergic genes including choline acetyltransferase (chata), vesicular acetylcholine transporters (vachta, vachtb), high-affinity choline transporter (hacta) and acetylcholinesterase (ache), while nicotinic acetylcholine receptor (nAChR) subunits are mainly expressed in interneurons. However, in 3-day-old embryos, we found an unexpected decrease in presynaptic cholinergic transcript expression in a rostral to caudal gradient in the spinal cord, which continued during development. On the contrary, nAChR subunits remained highly expressed throughout the spinal cord. We found that protein and enzymatic activities of presynaptic cholinergic genes were also reduced in the rostral spinal cord. Our work demonstrating that cholinergic genes are initially expressed in the embryonic spinal cord, which is dynamically downregulated during development suggests that cholinergic signaling may play a pivotal role during the formation of intra-spinal locomotor circuit.

虽然来自运动神经元的胆碱能神经传递在神经肌肉发育过程中的作用已经确立，但它是否调节脊髓中的中枢神经系统发育尚不清楚。斑马鱼提供了一个强大的模型来研究胆碱能系统在神经回路形成过程中是如何建立和进化的。在这项研究中，我们对胚胎和幼虫斑马鱼的胆碱能系统进行了详细的时空分析。在 1 天大的胚胎中，我们发现脊髓运动神经元表达突触前胆碱能基因，包括胆碱乙酰转移酶( chata ) 、囊泡乙酰胆碱转运蛋白( vachta , vachtb ) 、高亲和力胆碱转运蛋白( hacta ) 和乙酰胆碱酯酶 ( ache)，而烟碱乙酰胆碱受体 (nAChR) 亚基主要在中间神经元中表达。然而，在 3 天大的胚胎中，我们发现脊髓的头端到尾端梯度中突触前胆碱能转录物表达意外下降，这在发育过程中持续存在。相反，nAChR 亚基在整个脊髓中保持高度表达。我们发现突触前胆碱能基因的蛋白质和酶活性也在脊髓延髓中降低。我们的工作证明胆碱能基因最初在胚胎脊髓中表达，在发育过程中动态下调表明胆碱能信号可能在脊髓内运动回路的形成过程中发挥关键作用。