Dores R M, McDonald L K, Purdom L C, Sei C A
University of Denver, Department of Biological Sciences, CO 80208.
Brain Behav Evol. 1993;42(2):69-76. doi: 10.1159/000114141.
Acid extracts of the brain of a urodele amphibian, Ambystoma tigrinum, were screened with radioimmunoassays specific for enkephalin-related products and dynorphin-related products. Following Sephadex G-50 column chromatography a peak of enkephalin-sized immunoreactive material was detected near the total volume of the column. The enkephalin-sized immunoreactivity was further analyzed by reversed phase HPLC. This analysis detected peaks of authentic Met-enkephalin and Leu-enkephalin. However, the molar ratio of Met-enkephalin to Leu-enkephalin in the brain of this amphibian was approximately 80:1. These observations would suggest that the Leu-enkephalin detected in the brain of Ambystoma may be derived from a source other than the Proenkephalin precursor. Neither Met-enkephalin-RGL or Met-enkephalin-RF were detected by radioimmunoassay in brain extracts from this urodele. However, following digestion with trypsin and carboxypeptidase B, a novel peak of C-terminally extended Met-enkephalin was detected. Two peaks of Prodynorphin-related products were also detected following gel filtration chromatography. These immunoreactive forms were detected using antisera specific for alpha-neo-endorphin and dynorphin B(1-13). No immunoreactive forms with antigenic determinants similar to mammalian dynorphin A(1-17) or dynorphin A(1-8) were detected in this species. Reversed phase HPLC analysis indicated that the major form of urodele alpha-neo-endorphin eluted with the same retention time as synthetic mammalian alpha-neo-endorphin. Urodele dynorphin B(1-13)-related immunoreactivity eluted as a single peak, however, this form did not elute with the same retention time as synthetic mammalian dynorphin B(1-13).(ABSTRACT TRUNCATED AT 250 WORDS)
用针对脑啡肽相关产物和强啡肽相关产物的放射免疫分析法，对有尾两栖动物虎纹钝口螈大脑的酸性提取物进行了筛选。经葡聚糖凝胶G - 50柱色谱分离后，在柱总体积附近检测到一个脑啡肽大小的免疫反应性物质峰。通过反相高效液相色谱法对脑啡肽大小的免疫反应性进行了进一步分析。该分析检测到了天然甲硫氨酸脑啡肽和亮氨酸脑啡肽的峰。然而，这种两栖动物大脑中甲硫氨酸脑啡肽与亮氨酸脑啡肽的摩尔比约为80:1。这些观察结果表明，在虎纹钝口螈大脑中检测到的亮氨酸脑啡肽可能并非源自脑啡肽原前体。在这种有尾两栖动物的脑提取物中，放射免疫分析法未检测到甲硫氨酸脑啡肽 - RGL或甲硫氨酸脑啡肽 - RF。然而，用胰蛋白酶和羧肽酶B消化后，检测到了一个C末端延伸的甲硫氨酸脑啡肽新峰。凝胶过滤色谱分离后还检测到了两个强啡肽原相关产物峰。使用针对α - 新内啡肽和强啡肽B(1 - 13)的抗血清检测到了这些免疫反应形式。在该物种中未检测到具有与哺乳动物强啡肽A(1 - 17)或强啡肽A(1 - 8)相似抗原决定簇的免疫反应形式。反相高效液相色谱分析表明，有尾两栖动物α - 新内啡肽的主要形式与合成的哺乳动物α - 新内啡肽在相同保留时间洗脱。有尾两栖动物强啡肽B(1 - 13)相关的免疫反应性以单峰形式洗脱，然而，这种形式与合成的哺乳动物强啡肽B(1 - 13)在不同保留时间洗脱。（摘要截短于250字）
