dibutyryl-cyclic-gmp and Pituitary-Neoplasms

Controversies remain whether atrial natriuretic factor (ANF) may play a role in modulating the release of POMC derived peptides from pituitary corticotrophs. Employing AtT-20 mouse pituitary tumour cells, we report here the effects of rat ANF(1-28) and sodium nitroprusside (SNP), both of which augment cellular levels of cGMP through activating particulate and soluble guanylyl cyclases respectively, on the expression of POMC mRNA abundance. Furthermore, the cellular contents and secretion of (beta endorphin-like immunoreactivity) beta EP-LI from these cultures were also examined. Whereas the abundance of POMC mRNA was found to be markedly suppressed following 4h of incubation with rANP(1-28) (0.01 to 1 microM), SNP (0.1 to 10 microM) and dibutyryl-cGMP (1 to 100 microM) in a dose related manner, only a modest reduction in the release and cell contents of beta EP-LI was found in some of these cultures. It is also of interest to note that in all the cases examined, the inhibitory effect was associated with a significant suppression of cAMP levels in the cultures. Taken together, our present findings suggest that ANF may play a more important role in suppressing the production than the release of POMC related peptides from AtT-20 cells. Thus, it raises the possibility that hypothalamic ANF may likewise modulate the function of the pituitary-adrenal axis through exerting a greater effect on inhibiting the production than the secretion of pituitary ACTH.

Topics: Animals; Atrial Natriuretic Factor; beta-Endorphin; Blotting, Northern; Dibutyryl Cyclic GMP; Dose-Response Relationship, Drug; Gene Expression Regulation; Mice; Nitroprusside; Pituitary Neoplasms; Pro-Opiomelanocortin; RNA, Messenger; Tumor Cells, Cultured

Although C-type natriuretic peptide (CNP) has been shown to exist at the highest concentration in the anterior pituitary in rat tissues, its physiological role(s) there is (are) not clear. In this study, we report a novel function of CNP examined with anterior pituitary-derived cell lines, GH3 and AtT20/D16v-F2. Both CNP and atrial natriuretic peptide (ANP) increased cellular cGMP levels in both cell lines in dose-dependent manners. CNP, but not ANP, stimulated growth hormone (GH) release from GH3 cells. In contrast, neither ANP nor CNP had any significant effect on the corticotropin release from AtT20/D16v-F2 cells. An activator for cGMP-dependent protein kinase (cGK), dibutyryl cGMP, mimicked the stimulation of GH release from GH3 cells by CNP. Constitutive GH release from GH3 cells was greatly diminished in the presence of inhibitors for cAMP-dependent protein kinase, while stimulative GH release by CNP was not affected. However, inhibitors which can block cGK almost completely diminished the stimulative effect of CNP. An inhibitor for protein kinase C did not show any effect on either constitutive or CNP-stimulative GH release. Our observations indicate that the stimulation of GH release from GH3 cells by CNP is mediated mainly by the cGK signal-transduction pathway, not by cAMP-dependent protein kinase or protein kinase C, through a CNP-specific receptor (possibly ANP-B receptor). Thus, CNP may act as a local modulator in the anterior pituitary.

Topics: Animals; Atrial Natriuretic Factor; Bucladesine; Cell Line; Cyclic AMP; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Dibutyryl Cyclic GMP; Dose-Response Relationship, Drug; Growth Hormone; Kinetics; Natriuretic Peptide, C-Type; Pituitary Gland, Anterior; Pituitary Neoplasms; Protein Kinase Inhibitors; Proteins; Rats

The effects of cyclic AMP treatment on total cAMP-dependent protein kinase activity in GH3 pituitary tumor cells have been studied. Incubation of cells for 24 h with 1 microM forskolin resulted in a 50% decrease in total cAMP-dependent protein kinase activity which was reversible upon removal of forskolin from culture media. A similar response was observed in GH3 cells treated with 5 ng/ml cholera toxin and 0.5 mM dibutyryl cAMP but not 0.5 mM dibutyryl cGMP. Northern blot analysis demonstrated that the steady-state level of the mRNA for each of the six kinase subunit isoforms studied was not detectably altered after treatment with 1 microM forskolin for 24 h. The concentration of catalytic subunit was also assessed by binding studies using a radiolabeled heat-stable protein kinase inhibitor. Treatment of GH3 cells with 1 microM forskolin for 24 h reduced protein kinase inhibitor binding activity by 50%, consistent with the observed forskolin-induced decrease in total kinase activity. Analysis of endogenous heat-stable protein kinase inhibitor activity in GH3 cell extracts showed no significant difference between forskolin-treated cells and cells maintained under control conditions. To assess possible effects on catalytic subunit degradation, pulse-chase experiments were performed and radiolabeled catalytic subunit was isolated by affinity chromatography. The results demonstrated that treatment of cells with chlorophenylthio-cAMP detectably increased the apparent degradation of radiolabeled catalytic subunit. The increased degradation of the catalytic subunit was sufficient to account for the observed decreases in kinase activity. These results suggest that relatively long term cAMP treatment can alter total cAMP-dependent protein kinase activity through effects to alter the degradation of the catalytic subunit of the enzyme.

Topics: Animals; Bucladesine; Cell Line; Cholera Toxin; Colforsin; Cyclic AMP; Cyclic GMP; Dibutyryl Cyclic GMP; Gene Expression Regulation, Enzymologic; Kinetics; Macromolecular Substances; Pituitary Neoplasms; Protein Kinases; Protein Processing, Post-Translational; RNA, Messenger