atrial-natriuretic-factor and Testicular-Neoplasms

The ability of ANP to inhibit the hydrolysis of phosphoinositides was examined in [3H] myoinositol-labeled intact murine Leydig tumor (MA-10) cells. Arginine vasopressin (AVP) stimulated the formation of inositol monophosphate (IP1), inositol bisphosphate (IP2), and inositol trisphosphate (IP3) both in a time-and dose-dependent manner in MA-10 cells. ANP inhibited the AVP-induced formation of IP1, IP2, and IP3 in these cells. The inhibitory effect of ANP on the AVP-stimulated formation of IP1, IP2, and IP3 accounted for 30%, 38% and 42%, respectively, which was observed at the varying concentrations of AVP. ANP caused a dose-dependent attenuation in AVP-stimulated production of IP1, IP2 and IP3 with maximum inhibition at 100 nM concentration of ANP. The production of inositol phosphates was inhibited in the presence of 8-bromo cGMP in a dose-dependent manner, whereas dibutyryl-cAMP had no effect on the generation of these metabolites. The LY 83583, an inhibitor of guanylyl cyclase and cGMP production, abolished the inhibitory effect of ANP on the AVP-stimulated production of inositol phosphates. Furthermore, 10 microM LY 83583 also inhibited the ANP-stimulated guanylyl cyclase activity and the intracellular accumulation of cGMP by more than 65-70%. The inhibition of cGMP-dependent protein kinase by H-8, significantly restored the levels of AVP-stimulated inositol phosphates in the presence of either ANP or exogenous 8-bromo cGMP. The results of this study suggest that ANP exerts an inhibitory effect on the production of inositol phosphates in murine Leydig tumor (MA-10) cells by mechanisms involving cGMP and cGMP-dependent protein kinase.

Topics: Aminoquinolines; Animals; Arginine Vasopressin; Atrial Natriuretic Factor; Cyclic GMP; Enzyme Inhibitors; Guanylate Cyclase; Hydrolysis; Inositol; Leydig Cell Tumor; Male; Mice; Phosphatidylinositols; Testicular Neoplasms; Tumor Cells, Cultured

Atrial natriuretic factor (ANF) is a peptide hormone discovered recently from the heart atrium that possesses potent natriuretic and vasorelaxant activities. Recently we found that ANF markedly stimulates intracellular cGMP and almost completely inhibits cAMP accumulation in testicular interstitial tumor cells [Pandey, K. N., Kovacs, W. J., & Inagami, T. (1985) Biochem. Biophys. Res. Commun. 133, 800-806]. These actions of ANF suggest the presence of ANF receptors in testicular interstitial cells. In this study, cultured murine Leydig tumor cells have been shown to contain specific binding sites for ANF. Saturation binding studies indicated a single class of binding sites with a Kd of 5 X 10(-9) M at a density of 2 X 10(6) sites/cell. The binding of mono[125I]iodo-ANF (125I-ANF) was competed by unlabeled ANF in a dose-dependent manner. Hormones unrelated to ANF such as angiotensin I, bovine luteinizing hormone, and human chorionic gonadotropin were not able to compete against 125I-ANF. The binding of 125I-ANF was rapid, reaching maximum levels in 15 min at 4 degrees C. At 37 degrees C, the cell-bound 125I label was quickly decreased. Pretreatment of cells with NH4Cl, chloroquine, or NaN3 resulted in significant increases in maximum levels of the cell-bound 125I radioactivity. A photoaffinity reagent for ANF receptor was prepared by reacting ANF with succinimido 4-azidobenzoate, and resultant 4-azidobenzoyl- (AZB-) ANF was purified by high-performance liquid chromatography (HPLC). AZB-ANF was radioiodinated by use of chloramine T and purified again by HPLC.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Atrial Natriuretic Factor; Binding, Competitive; Cell Line; Kinetics; Leydig Cell Tumor; Male; Mice; Receptors, Atrial Natriuretic Factor; Receptors, Cell Surface; Testicular Neoplasms

We have found that atrial natriuretic factor (ANF) has a profound effect on testicular cells in altering intracellular cyclic nucleotide levels as well as progesterone secretion. Using clonal cultured Leydig tumor cells we found that 1 X 10(-8)M ANF caused a two thousand-fold elevation in the accumulation of cellular cGMP and inhibited cAMP in treated cells by more than 90% as compared to the controls. ANF (1 X 10(-8)M) also significantly inhibited gonadotropin-stimulated accumulation of cAMP in response to bovine luteinizing hormone (bLH) or human chorionic gonadotropin (hCG). Gonadotropin-stimulated progesterone secretion was inhibited by ANF (1 X 10(-10) - 1 X 10(-9)M) in these cultured Leydig tumor cells. Approximately 50% inhibition of progesterone secretion was observed at the peptide concentration of 1 X 10(-9) M.

Topics: Animals; Atrial Natriuretic Factor; Chorionic Gonadotropin; Clone Cells; Cyclic AMP; Cyclic GMP; Gonadotropins; Leydig Cell Tumor; Luteinizing Hormone; Male; Mice; Progesterone; Testicular Neoplasms