vasoactive-intestinal-peptide and 2--5--dideoxyadenosine

We investigated the effect of neuropeptides, which are vasoactive intestinal polypeptide (VIP), substance P, (SP), neuropeptide Y (NPY), neurokinin A (NKA), somatostatin (SOM), calcitonin gene-related peptide (CGRP), and leucine-enkephalin (L-ENK), on the invasion of murine Colon 26-L5 adenocarcinoma cells through a reconstituted basement membrane (Matrigel) using a Transwell cell culture chamber assay. VIP, SP, NPY, and L-ENK reduced invasive potential of tumor cells in a concentration-dependent manner, whereas SOM, CGRP, and NKA had no effect. Especially, VIP showed the most effective in inhibiting tumor invasion, and achieved 50% reduction at 10(-6) M. A similar effect by VIP was also observed in cell migration to fibronectin. VIP had no effect on the growth of tumor cells at the concentrations ranging from 10(-10) to 10(-6) M. The suppressed ability of the tumor cell motility by VIP (10(-6) M) was practically recovered by co-treatment with 2',5'-dideoxyadenosine, an adenylate cyclase inhibitor. These results indicate that VIP, among the neuropeptides used, could inhibit Matrigel invasion of Colon 26-L5 carcinoma cells through partial suppression of their motility, and the reduction was associated with an intracellular cAMP-mediated pathway.

Topics: Adenocarcinoma; Animals; Biocompatible Materials; Calcitonin Gene-Related Peptide; Cell Division; Cell Movement; Colforsin; Collagen; Colonic Neoplasms; Dideoxyadenosine; Dose-Response Relationship, Drug; Drug Combinations; Enkephalins; Laminin; Leucine; Mice; Neoplasm Invasiveness; Neurokinin A; Neuropeptide Y; Neuropeptides; Proteoglycans; Somatostatin; Substance P; Tumor Cells, Cultured; Vasoactive Intestinal Peptide

The hormonal regulation of Na+-dependent phosphate transport was studied in opossum kidney (OK) cells. PTH caused time- and concentration-dependent decreases in Na+-dependent phosphate transport, with 10 pM PTH-(1-34) producing a 19% decline in phosphate transport. The EC50 for PTH inhibition of phosphate transport was 50 pM. Kinetic analyses of phosphate transport indicated that PTH decreased the maximum velocity without affecting the Km for phosphate. PTH increased cAMP formation with an EC50 of 10 nM. 8-Bromo-cAMP and (Bu)2cAMP also inhibited phosphate transport. Forskolin increased cAMP formation and decreased phosphate transport, whereas the cyclase-inactive forskolin analog 1,9-dideoxyforskolin also inhibited phosphate transport. The PTH analog [8,18-norleucine,34-tyrosinamide]PTH-(3-34) reduced phosphate transport at concentrations from 10 nM to 30 microM, but did not increase cAMP formation at concentrations up to 10 microM. The adenylate cyclase inhibitor 2',5'-dideoxyadenosine produced concentration-dependent decreases in PTH-stimulated cAMP formation, but did not influence PTH inhibition of Na+-dependent phosphate transport. Vasoactive intestinal polypeptide and prostaglandin E1 increased cAMP formation in OK cells, but were weak inhibitors of phosphate transport. This study suggests that cAMP may not be the only transmembrane signaling mechanism involved in the regulation of Na+-dependent phosphate transport by PTH-(1-34) in OK cells.

Topics: 8-Bromo Cyclic Adenosine Monophosphate; Alprostadil; Animals; Biological Transport; Bucladesine; Cell Line; Colforsin; Cyclic AMP; Deoxyadenosines; Dideoxyadenosine; Kidney; Kinetics; Opossums; Parathyroid Hormone; Peptide Fragments; Phosphates; Sodium; Teriparatide; Vasoactive Intestinal Peptide

The mechanism by which somatostatin acts to modulate cholinergic transmission is not clear. In this study we investigated the role of the adenosine 3',5'-cyclic monophosphate (cAMP) system in mediating cholinergic transmission in the guinea pig myenteric plexus and examined the ability of somatostatin to alter acetylcholine (ACh) release stimulated by various cAMP agonists. Forskolin, 8-bromo-cAMP, vasoactive intestinal peptide (VIP), and cholera toxin each stimulated the release of [3H]ACh in a dose-related manner. Addition of theophylline enhanced the release of [3H]ACh stimulated by these cAMP agonists. In contrast 2',5'-dideoxyadenosine, an inhibitor of adenylate cyclase, antagonized the action of forskolin, VIP, and cholera toxin but had no effect on that evoked by 8-bromo-cAMP. These observations suggest that cAMP may serve as a physiological mediator for ACh release from myenteric neurons. Somatostatin inhibited release of [3H]ACh evoked by various cAMP agonists in a dose-related manner. Maximal inhibition, observed in the presence of 10(-6) M somatostatin was 48 +/- 5, 47 +/- 9, and 43 +/- 12% of control for forskolin-, VIP-, and cholera toxin-evoked release of [3H]ACh. In contrast somatostatin at 10(-6) M inhibited only 20 +/- 5% of the release of [3H]ACh stimulated by 8-bromo-cAMP. Pretreatment with pertussis toxin antagonized the inhibitory effect of somatostatin on the release of [3H]ACh evoked by forskolin, VIP, or cholera toxin but had no effect on the inhibitory action of somatostatin on the release of [3H]ACh evoked by 8-bromo-cAMP.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 8-Bromo Cyclic Adenosine Monophosphate; Acetylcholine; Adenylate Cyclase Toxin; Animals; Cholera Toxin; Colforsin; Cyclic AMP; Deoxyadenosines; Dideoxyadenosine; Guinea Pigs; Male; Myenteric Plexus; Pertussis Toxin; Somatostatin; Synaptic Transmission; Theophylline; Vasoactive Intestinal Peptide; Virulence Factors, Bordetella

Although purine nucleosides have been shown to regulate the secretion of several peptide and steroid hormones, effects on pituitary hormone release have not been reported. We show here that in the clonal GH4C1 pituitary cell line maximal concentrations of adenosine (greater than or equal to 50 microM) inhibited PRL and GH secretion by 40%. Adenosine deaminase abolished the inhibitory effect of adenosine but not that of SRIF or (-)N6(R-2-phenylisopropyl)adenosine (PIA), a nonhydrolyzable adenosine analog. Furthermore, this enzyme increased basal secretion by 50%, and analysis of the incubation medium by HPLC demonstrated that the cells secreted biologically effective concentrations of adenosine. These results indicate that adenosine produced in culture tonically inhibits hormone release. In other target cells, adenosine inhibition is mediated by two types of binding sites: an extracellular Ri-site requiring an intact ribose moiety or an intracellular P-site requiring an intact purine ring. Four lines of evidence indicate that in GH4C1 cells, adenosine acts at an Ri-site. PIA, an Ri-site-specific agonist, was a potent inhibitor of hormone release (ED50 = 30 nM). Theophylline, an Ri-site antagonist, competitively inhibited the action of PIA (Ki = 2.4 microM). 3) 2'5'-Dideoxyadenosine, a P-site-specific agonist, did not inhibit PRL release even at a concentration of 1 mM. 4) Dipyridamole, an adenosine uptake inhibitor, did not reduce adenosine inhibition. In addition to its effect on basal secretion, PIA inhibited stimulation of hormone release by vasoactive intestinal peptide and TRH. PIA also reduced vasoactive intestinal peptide-stimulated cAMP accumulation by 75%, consistent with its action to inhibit adenylate cyclase via Ri receptors in other targets. Since PIA inhibition of PRL release and cAMP accumulation was not additive with the effects of SRIF and carbamyl choline, these inhibitors may act via a common rate-limiting step. Our results demonstrate that adenosine activates an Ri-type of adenosine receptor in GH4C1 cells and that the production of adenosine under normal culture conditions causes autocrine inhibition of secretion.

Topics: Adenine; Adenosine; Adenosine Deaminase; Animals; Carbachol; Cell Line; Chromatography, High Pressure Liquid; Cyclic AMP; Deoxyadenosines; Dideoxyadenosine; Dipyridamole; Growth Hormone; Phenylisopropyladenosine; Pituitary Gland; Pituitary Neoplasms; Prolactin; Rats; Thyrotropin-Releasing Hormone; Vasoactive Intestinal Peptide