somatostatin--tyr(11)- and Pituitary-Neoplasms

Somatostatin-14 (S-14) acts via specific receptors to inhibit basal as well as hormone- and forskolin-stimulated ACTH secretion in tumor cells (AtT-20/D16-16) of mouse anterior pituitary. In addition S-14 inhibits the stimulated but not basal cAMP accumulation. The potency of somatostatin-28 (S-28) for regulating these processes in these tumor cells has not been reported. In this study we have investigated the relationship between receptor-binding affinities of S-14 and S-28 and their biopotency in these cells. Membrane receptors for S-14 characterized using [125I-Tyr11]S-14 as the radioligand [maximum binding capacity (Bmax) = 1.28 +/- 0.1 pmol/mg; dissociation constant (Kd) = 1.1 +/- 0.04 nM] bound S-28 with 3-fold greater affinity than S-14. Binding sites quantitated using an S-28 analog [Leu8, D-Trp22, 125I-Tyr25]S-28 as radioligand (Bmax = 1.18 +/- 0.15 pmol/mg; Kd = 0.08 +/- 0.06 nM) also exhibited greater affinity for S-28 than S-14. Forskolin-stimulated cAMP accumulation and ACTH secretion in these cells were inhibited to a greater extent (4- and 9-fold, respectively) by S-28 than S-14. Preincubation of the cells with S-14 and S-28 (10(-7) M) resulted in a marked decrease (36% and 71%, respectively) of S-14 receptor concentration. Coincubation of the cells with both S-14 and S-28 led to 56% decrease in S-14 receptor binding. The responsiveness of the cells to forskolin stimulation of ACTH secretion and cAMP accumulation was significantly enhanced by preincubation with S-14 (10(-7) M) whereas the responsiveness to forskolin was completely abolished by preincubation with S-28. Simultaneous exposure of the cells to both S-14 and S-28 resulted in a partial reversal of the inhibiting effect of S-28 on forskolin-stimulated cAMP accumulation in these cells but did not result in a partial reversal of the inhibitory effect of S-28 on forskolin-stimulated ACTH secretion in these cells. These results demonstrate that S-28 is more potent than S-14 in AtT-20/D16-16 cells, its greater potency arising from its greater affinity for binding to S-14 receptors. The differential effects of these peptides after preincubation on the responsiveness of ACTH secretion and cAMP accumulation in these cells to forskolin stimulation suggests the possibility of existence of distinct S-14 and S-28 receptors, but these could not be identified by direct binding experiments using the S-14 and S-28 analogs employed in the study as radioligands.(ABSTRACT TRUNCATED AT 400 WORDS)

Topics: Adrenocorticotropic Hormone; Animals; Cell Line; Cell Membrane; Colforsin; Cyclic AMP; Diterpenes; Kinetics; Mice; Peptide Fragments; Pituitary Neoplasms; Receptors, Cell Surface; Receptors, Somatostatin; Somatostatin; Somatostatin-28

Specific somatostatin (SRIH) receptors on human pituitary adenoma cell membranes were characterized using [125I]Tyr11-SRIH as the radioligand. Specific binding of [125I] Tyr11-SRIH to adenoma cell membranes reached a steady state within 30 min at 25 C, and semilogarithmic analysis of the data revealed that the rate of the binding was linear at 25 C with a t1/2 of 13.2 min. Specific binding increased linearly with 5-160 micrograms plasma membrane protein. SRIH-14 and SRIH-28 inhibited [125I]Tyr11-SRIH binding to adenoma cell membranes with ID50S of 0.32 and 0.50 nM, respectively, while secretin, glucagon, gastrin, cholecystokinin-8, bombesin, TRH, LHRH, human GH-releasing factor-(1-44)-NH2, D-Ala2-met-enkephalin, gamma-aminobutyric acid and taurine did not significantly inhibit binding. All of 13 GH-secreting adenomas investigated had specific and high affinity SRIH receptors, with a dissociation constant (Kd) of 0.80 +/- 0.15 nM (mean +/- SEM) and a maximal binding capacity (Bmax) of 234.2 +/- 86.9 fmol/mg protein (mean +/- SEM). Among five of the nonsecreting pituitary adenomas examined, two had SRIH receptors with Kd values of 0.18 and 0.32 nM and Bmax values of 17.2 and 48.0 fmol/mg protein, respectively. In the remaining three, SRIH receptors were not detected. These results indicate that GH-secreting adenomas as well as some nonfunctioning adenomas have specific SRIH receptors, and hence, the function of the adenomas could be altered by SRIH.

Topics: Adenoma; Adult; Cell Membrane; Female; Growth Hormone; Humans; Kinetics; Male; Middle Aged; Pituitary Neoplasms; Protein Binding; Receptors, Cell Surface; Receptors, Somatostatin; Somatostatin