Development of the technique for expression of peptide receptors in the oocyte membrane

虽然肽受体细胞广泛分布于中枢和外周神经系统，但这些细胞太小，无法分析与受体激活相关的详细离子机制。本研究旨在分析直径约1 mm的非洲爪哇卵母细胞中表达的多肽受体激活的离子机制。为此，将从大鼠脑或牛胃提取的mRNA注射到卵母细胞中，并在电压钳下记录注射的卵母细胞在孵育几天后对各种多肽的反应。虽然通过这种技术可以表达五种不同多肽的受体，但研究集中在P物质受体上。P物质是速激肽家族的一员。目前，除了P物质外，哺乳动物神经系统中还存在另外两种速激肽，即K物质和Neuromedin K。与MUL…的存在一致更多的速激肽，已经被认为存在三种亚型的速激肽受体。通过不同速激肽激动剂的效力排名，可以区分速激肽受体的三个亚型。在此基础上，发现由大鼠脑mRNA诱导的速激肽受体对应于SP-P速激肽受体，而由牛胃mRNA诱导的速激肽受体对应于SP-K速激肽受体。因此，在同一表达系统中可以诱导两种不同类型的速激肽受体，这取决于所注入的外源mRNA的来源。这表明这两种受体由不同的分子组成。因此，这两种受体在性质上的差异不能归因于仅发生在其目的组织的可能的翻译后修饰。另一方面，这两种受体激活的离子机制都是基于&lt；Cl^-&gt；电导的增加。这表明，这两种受体的激活使用相同的细胞内第二信使，而第二信使又激活卵母细胞膜上的离子通道。第二信使的性质在卵母细胞和哺乳动物神经元中可能是不同的。因此，在卵母细胞中分析的离子机制不能概括为激活其他物种中存在的速激肽受体。较少

While peptide-receptive cells are widely distributed in both the central and peripheral nervous systems, these cells are too small to analyze detailed ionic mechanisms associated with activation of the receptors. The present study has been addressed to the analysis of ionic mechanisms underlying activation of the peptide receptors expressed in Xenopus oocytes which have a diameter of about 1 mm. For this purpose, mRNA isolated from the rat brain or the bovine stomach was injected into oocytes, and the response of the injected oocytes to a variety of peptides was recorded under voltage clamp following incubation for a few days. While the receptors to five different peptides could be expressed by this technique, the study was focused on the substance P receptor. Substance P is a member of the tachykinin family. At present, in addition to substance P, two other tachykinins, substance K and neuromedin K, are known to exist in the mammalian nervous system. In accord with the presence of mul … More tiple tachykinins, the existence of three subtypes of tachykinin receptors has been suggested. The three subtypes of tachykinin receptors could be distinguished by potency ranking of different tachykinin agonists. On this basis, the tachykinin receptor induced by rat brain mRNA was found to correspond to the SP-P tachykinin receptor, whereas that directed by bovine stomach mRNA corresponded to the SP-K tachykinin receptor. Thus, two different types of tachykinin receptors could be induced in the same expression system, depending upon the source of exogenous mRNA injected. This indicates that the two receptors are made of different molecules. Therefore, the difference in nature between the two receptors cannot be attributed to the possible posttranslational modification occurring at their destination tissue alone. On the other hand, the ionic mechanisms responsible for activation of the two receptors are both based on an increase in the <Cl^-> conductance. This suggests that activation of the two receptors uses the same intracellular second messenger, which in turn, activates <Cl^-> ionic channels in the oocyte membrane. The nature of the second messenger may be different between oocytes and mammalian neurons. Therefore, the ionic mechanisms analyzed in oocytes cannot be generalized to activation of tachykinin receptors present in other species. Less