Troppmann Britta, Balfanz Sabine, Krach Christian, Baumann Arnd, Blenau Wolfgang
Institute of Biochemistry and Biology, University of Potsdam, Potsdam D-14476, Germany.
Int J Mol Sci. 2014 Jan 6;15(1):629-53. doi: 10.3390/ijms15010629.
We have isolated a cDNA coding for a putative invertebrate-type dopamine receptor (Peadop2) from P. americana brain by using a PCR-based strategy. The mRNA is present in samples from brain and salivary glands. We analyzed the distribution of the PeaDOP2 receptor protein with specific affinity-purified polyclonal antibodies. On Western blots, PeaDOP2 was detected in protein samples from brain, subesophageal ganglion, thoracic ganglia, and salivary glands. In immunocytochemical experiments, we detected PeaDOP2 in neurons with their somata being located at the anterior edge of the medulla bilaterally innervating the optic lobes and projecting to the ventro-lateral protocerebrum. In order to determine the functional and pharmacological properties of the cloned receptor, we generated a cell line constitutively expressing PeaDOP2. Activation of PeaDOP2-expressing cells with dopamine induced an increase in intracellular cAMP. In contrast, a C-terminally truncated splice variant of this receptor did not exhibit any functional property by itself. The molecular and pharmacological characterization of the first dopamine receptor from P. americana provides the basis for forthcoming studies focusing on the significance of the dopaminergic system in cockroach behavior and physiology.
我们采用基于聚合酶链反应（PCR）的策略，从美洲大蠊脑中分离出了一个编码假定无脊椎动物型多巴胺受体（Peadop2）的互补脱氧核糖核酸（cDNA）。该信使核糖核酸（mRNA）存在于脑和唾液腺的样本中。我们用特异性亲和纯化的多克隆抗体分析了Peadop2受体蛋白的分布。在蛋白质印迹法中，在脑、咽下神经节、胸神经节和唾液腺的蛋白质样本中检测到了Peadop2。在免疫细胞化学实验中，我们在神经元中检测到了Peadop2，这些神经元的胞体位于髓质前缘，双侧支配视叶并投射到腹外侧原脑。为了确定克隆受体的功能和药理学特性，我们构建了一个组成性表达Peadop2的细胞系。用多巴胺激活表达Peadop2的细胞会导致细胞内环磷酸腺苷（cAMP）增加。相比之下，该受体的C末端截短剪接变体自身不表现出任何功能特性。对美洲大蠊首个多巴胺受体的分子和药理学特性进行表征，为即将开展的聚焦多巴胺能系统在蟑螂行为和生理学中意义的研究奠定了基础。
