Pituitary secretory vesicle enzymes involved in the processing of pro-opiomelanocortin (POMC, pro-ACTH/endorphin) and pro-vasopressin were studied. A 70,000 mol. wt., paired basic residue specific prohormone converting enzyme (PCE), previously purified and characterized as an aspartyl protease, has recently been shown to be structurally related to Cathepsin D. PCE was shown to be secreted from bovine intermediated lobe together with alpha-MSH, in a co-ordinately regulated manner. Pepstatin A, an inhibitor of PCE, blocked processing of POMC in the mouse intermediate lobe further supporting a physiologic role of the enzyme in vivo. Cloning of this enzyme is in progress. Our previous finding that mice, during salt-loading stress, exhibit an increase in plasma ACTH and anterior pituitary POMC mRNA levels, prompted studies to determine the involvement of CRF and vasopressin (AVP) in mediating this response. Quantitative in situ hybridization analyses of CRF and AVP mRNA levels in hypothalamic neurons indicate no changes in CRF mRNA, but an increase in AVP mRNA after salt-loading. The content of CRFi in the median eminence, the site of release of CRF also showed no change after salt-loading. Plasma AVP was increased, and AVP appears to be the primary regulator of ACTH secretion, in the presence of basal levels of CRF, during salt-loading stress. Studies using dissociated mouse anterior pituitary cells reveal that AVP is highly effective in potentiating CRF action at low doses of CRF. Analysis of the molecular mechanism for the signal transduction of CRF and AVP indicate that CRF acts through cAMP-dependent, and AVP through a phosphoinositol by an increase in cAMP, as in the rat, but perhaps through an increase in the efficacy of phosphorylation of proteins involved in the secretory process.