Adrenal Medullary Enkephalin-Containing Peptides

虽然前脑啡肽的氨基酸序列和许多加工产物的特性是已知的，但关于前脑啡肽的合成和降解速率或前脑啡肽的加工和储存细节的信息很少。这个项目的第一个目标是确定前脑啡肽的合成和降解率。牛肾上腺髓质嗜铬细胞将与放射性标记的氨基酸孵育，并将它们掺入不同前脑啡肽片段中的脑啡肽序列，然后用高效液相色谱分离。第二个目的是确定脑啡肽原的加工途径。嗜铬细胞内脑啡肽原序列的脉冲追逐放射性标记将被用来研究这一途径。第三个目的是确定负责调节前脑啡肽合成和加工的第二信使(S)。将评估钙、环磷酸腺苷和包括糖皮质激素在内的其他药物的作用。第四个目的是确定前脑啡肽与嗜铬小泡合成的关系。来自放射性标记的嗜铬细胞的囊泡将经过密度梯度离心法；部分组分将分析新合成的脑啡肽序列和其他囊泡成分。这些研究将阐明神经递质-激素合成和储存的基本机制。

Although the amino acid sequence and the identities of many processing products of proenkephalin are known, little information is available on the rates of proenkephalin synthesis and degradation or on the details of proenkephalin processing and storage. The first aim of this project is to determine the rates of proenkephalin synthesis and degradation. Bovine adrenal medullary chromaffin cells will be incubated with radiolabeled amino acids, and their incorporation into enkephalin sequences in the various proenkephalin fragments will be determined following fractionation by high performance liquid chromatography. The second aim is to determine the pathway of proenkephalin processing. Pulse-chase radiolabeling of enkephalin sequences within chromaffin cell proenkephalin will be used to study this pathway. The third aim is to determine the second messenger(s) responsible for regulation of proenkephalin synthesis and processing. The roles of calcium, cyclic AMP, and other agents, including glucocorticiods, will be evaluated. The forth aim is to determine the relationship between proenkephalin and chromaffin vesicle synthesis. Vesicles from radiolabeled chromaffin cells will be subjected to density gradient centrifugation; fractions will be analyzed for newly synthesized enkephalin sequences and other vesicle components. These studies will elucidate the basic mechanisms of neurotransmitter-hormone synthesis and storage.