This application is to study the in vivo biosynthesis of the somatostatin family of peptides in the hypothalamus. The pathway involves initial synthesis from messenger RNA of a prosomatostatin (proSRIF) molecule, which is then cleaved by proteolytic enzymes into a variety of smaller peptides, including somatostatin-28 (SRIF-28) and somatostatin-14; (SRIF-14; the original growth hormone inhibiting hormone of the hypothalamus). Over the past few years, we have developed a method for measuring SRIF-14 and SRIF-28 synthesis in vivo in rat hypothalamus, and have used it to study several aspects of somatostatin biosynthesis. The method involves administering (35S)cysteine into the third ventricle, and then removing the hypothalamus several hours later and measuring labeled cysteine incorporation into SRIF 14 and SRIF-28. The peptides are isolated by high-performance liquid chromatography (HPLC) prior to label quantitation. We propose to continue to study factors governing SRIF-14 and SRIF-28 biosynthesis in hypothalamus. We will investigate the effects on SRIF synthesis of drugs, hormones, and other treatments known to modify immunoreactive SRIF levels in hypothalamus. We will develop HPLC methods for isolating and quantitating proSRIF. We then plan to study labeled cysteine incorporation into proSRIF, and the effects of physiologic, pharmacologic, and hormonal treatments on its production and processing to SRIF-14 and SRIF-28. Finally, we will begin to study SRIF synthesis from (35S)cysteine during postnatal development in hypothalamus, and also begin to investigate SRIF synthesis in vivo and in vitro in another portion of the central nervous system (CNS) that contains SRIF-immunoreactive neurons having no endocrine functions: the retina. This latter CNS region has been chosen in particular because it readily lends itself to in vitro biochemical study with minimal disruption to the tissue. The results of these investigations should provide information on the factors governing the synthesis of the somatostatin peptides in the CNS, and help in understanding the functional roles of SRIF neurons in overall CNS function. In addition, they should help in building of a foundation of basic information that will help in understanding the etiologies of CNS diseases associated with abnormal GH secretion, and perhaps also with dementias and other neurological diseases associated with forebrain deterioration (and loss of somatostatin neurons).
