Phylogeny and comparative anatomy of the cholinergic neuron in invertebrate and vertebrates

胆碱乙酰转移酶（ChAT）是乙酰胆碱的生物合成酶，是胆碱能神经元最特异的标志物。利用分子生物学技术，我们先前制备了大鼠ChAT的抗血清，并建立了免疫组织化学。最近我们发现抗血清可在石蜡包埋切片中染色胆碱能神经元，这使我们能够研究胆碱能神经元的发生和比较解剖。然而，我们发现抗血清的跨种反应性不足以完成本项目。因此，我们生产了一种抗血清囊泡乙酰胆碱转运蛋白（VAChT），另一种特异性蛋白的胆碱能神经元，并检查它是否可以用于本项目。为此，我们使用抗血清和地高辛标记的RNA证明ChAT和VAChT，通过免疫组织化学和原位杂交组织化学，单独和与它们的各种组合，广泛比较了ChAT和VAChT的mRNA和相应蛋白的分布，并证明VAChT的抗血清可用于本项目。

Choline acetyltransferase (ChAT), the biosynthetic enzyme for ACh, is the most specific marker for the cholinergic neuron. Using the molecular biological techniques, we previously produced an antiserum to rat ChAT and established the immunohistochemistry. Recently we found that the antiserum stains cholinergic neurons in paraffin-embedded sections, and this enables us to study the phylogeny and comparative anatomy of the cholinergic neuron. We found, hoever, that cross-species reactivity of the antiserum is not wide enough to complete the present project. We, therefore, produced an antiserum to vesicular ACh transporter (VAChT), another specific protein for the cholinergic neuron, and examined whether or not it can be used for the present project. For this, using antisear to and digoxigenin-laveled RNA proves for ChAT and VAChT,we compared extensively the distribution of mRNAs and the corresponding proteins for ChAT and VAChT by immunohistochemistry and in situ hybridization histochemistry, both alone and with various combinations of them, and demonstrated that the antiserum to VAChT can be used for the present project.