Many environmental stimuli influence endocrine and neural homeostasis. These effects may be manifested by changes in chemical communication in these systems, particularly as they relate to reproduction. Since peptides convey much of this information and protein kinase C (PKC) is a major second messenger in these systems, in vivo and cell culture studies have been used to examine the role that PKC plays in pro-LHRH gene expression, peptide biosynthesis, processing and secretion. In rat hypothalamus and an immortalized LHRH neuronal cell line, the LHRH precursor is processed to several different peptides. These studies and Northern blots suggest that prohormone convertase (PC), carboxypeptidase, glutaminyl cyclase, and alpha-amidating enzyme may catalyze the reactions. In order to test the role of the PCs more directly, BSC-40 kidney cells were infected with vacinnia virus containing LHRH or in combination with PC1, PC2 or furin. Biochemical studies indicate that pro-LHRH is most efficiently cleaved by PC2. Hence, these studies and Northerns from LHRH neurons suggest that PC2 may be the first enzyme in the pro-LHRH metabolic cascade. In secretion studies, the immortalized LHRH neurons secrete LHRH in a pulsatile fashion on the same time-scale as that found in vivo. Since synaptic and gap- junctional connections are formed among these cells, the observed rhythmic behavior may be located within this neuronal network. The immortalized cells appear to secrete LHRH by both the constitutive and regulated pathways. Agents which activate protein kinase C and A used the regulated pathway. Chronic administration of phorbol esters downregulates PKC-alpha, -beta I, - beta II, - delta, - epsilon, but not -zeta in these neurons. These agents also profoundly depress LHRH gene expression. In an attempt to functionally characterize the PKC isoenzymes, we have shown that some of these isoforms show heterogeneity of size and of tissue and cellular distribution. These changes are particularly marked in brain. Future studies will attempt to identify which environmental stimuli affect reproduction through alterations both in PKC and in pro-LHRH gene expression, biosynthesis, processing and secretion.
