Adrenal Medullary Enkephalin-Containing Peptides

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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.