http://http://dspace.unitus.it:80 The DSpace digital repository system captures, stores, indexes, preserves, and distributes digital research material. Wed, 19 Jun 2013 18:59:14 GMT 2013-06-19T18:59:14Z http://hdl.handle.net/2067/1444 Title: Pharmacological characterisation of native somatostatin receptors in AtT-20 mouse tumour corticotrophs Authors: Cervia, Davide; Nunn, Caroline; Fehlmann, Dominique; Langenegger, Daniel; Schuepbach, Edi; Hoyer, Daniel Abstract: 1. The mouse corticotroph tumour cell line AtT-20, is a useful model to investigate the physiological role of native somatostatin (SRIF) receptor subtypes (sst1-sst5). The objective of this study was to characterise the pharmacological features and the functional effects of SRIF receptors expressed by AtT-20 cells using radioligand binding and cAMP accumulation. 2. [125I]LTT-SRIF-28, [125I]CGP 23996, [125I]Tyr10-cortistatin-14 and [125I]Tyr3-octreotide labelled SRIF receptor binding sites with high affinity and in a saturable manner (Bmax = 315, 274, 239 and 206 fmol mg-1, respectively). [125I]LTT-SRIF-28 labels significantly more sites than [125I]Tyr10-cortistatin-14 and [125I]Tyr3-octreotide as seen previously in cells expressing pure populations of sst2 or sst5 receptors. 3. SRIF analogues displaced the binding of the four radioligands. sst2/5 receptor-selective ligands showed much higher affinity than sst1/3/4 receptor-selective ligands. The binding profile of [125I]Tyr3-octreotide was different from that of [125I]LTT-SRIF-28, [125I]CGP 23996 and [125I]Tyr10-cortistatin-14. The sst5/1 receptor-selective ligand L-817,818 identified two binding sites, one with subnanomolar affinity (sst5 receptors) and one with micromolar affinity (sst2 receptors), however the proportions were different: 70-80% high affinity with [125I]LTT-SRIF-28, [125I]CGP 23996, [125I]Tyr10-cortistatin-14 but only 20% with [125I]Tyr3-octreotide. 4. SRIF analogues concentration-dependently inhibited the forskolin-stimulated cAMP levels. sst2/5 receptor-selective ligands were highly potent, whereas sst1/3/4 receptor-selective ligands had no significant effects. The sst2 receptor antagonist D-Tyr8-CYN 154806 competitively antagonised the effects of SRIF-14 and sst2 receptor-preferring agonists, but not those of L-817,818. The complex binding properties of SRIF receptor analogues indicates that sst2 and sst5 receptors are the predominant SRIF receptors expressed on AtT-20 cell membranes with no or only negligible presence of sst1, sst3 and sst4 receptors. In the functional studies using cAMP accumulation, only sst2 and sst5 receptors appear to play a role. However, the “predominant” receptor appears to be the sst2 receptor, although sst5 receptors can also mediate the effect, when the ligand is not able to activate sst2 receptors. This clearly adds flexibility to SRIF-mediated functional effects and suggests that the physiological role of SRIF and its analogs may be mediated preferentially via one subtype over another. Description: L'articolo è disponibile sul sito dell'editore http://onlinelibrary.wiley.com/ Tue, 31 Dec 2002 23:00:00 GMT http://hdl.handle.net/2067/1444 2002-12-31T23:00:00Z http://hdl.handle.net/2067/1456 Title: Binding and functional properties of the novel somatostatin analogue KE 108 at native mouse somatostatin receptors Authors: Cervia, Davide; Langenegger, Daniel; Schuepbach, Edi; Cammalleri, Maurizio; Schoeffter, Philippe; Schmid, Herbert A.; Bagnoli, Paola; Hoyer, Daniel Abstract: Clinically used somatostatin (SRIF) analogs octreotide and lanreotide act primarily by binding to SRIF receptor subtype 2 (sst2). In contrast, the recently described multiligand SOM230 binds with high affinity to sst1-3 and sst5 and KE 108 is characterised as a high affinity ligand for all five SRIF receptors. In tumoural mouse corticotrophs (AtT-20 cells) and in mouse hippocampus, binding and functional features of KE 108 were examined and compared to SRIF-14, octreotide and SOM230. In AtT-20 cells, KE 108 bound with high affinity at [125I]LTT-SRIF-28-labelled sites similarly to SRIF, octreotide and SOM230. At the functional level, all four ligands increased guanosine-5’--(3-35Sthio)-triphosphate binding and decreased cAMP accumulation or intracellular Ca2+ concentration through Gi/o proteins. In hippocampal slices, KE 108, octreotide and SOM230 also bound with high affinity at [125I]LTT-SRIF-28-labelled sites similarly to SRIF, but KE108, octreotide or SOM230 did not influence spontaneous epileptiform activity which was, in contrast, inhibited by SRIF. In conclusion, this study demonstrates that KE 108 has high affinity for native mouse SRIF receptors. Functionally, KE 108 mediates SRIF action at sst2/5 in corticotrophs whereas it does not mimic the SRIF-induced inhibition of hippocampal excitation suggesting that the high potency and efficacy of a synthetic ligand to all known SRIF receptors may not reproduce entirely the effects of the natural SRIF. Description: L'articolo è disponibile sul sito dell'editore http://www.sciencedirect.com/ Fri, 31 Dec 2004 23:00:00 GMT http://hdl.handle.net/2067/1456 2004-12-31T23:00:00Z http://hdl.handle.net/2067/1443 Title: Biological activity of somatostatin receptors in GC rat tumour somatotrophs: evidence with sst1-sst5 receptor-selective nonpeptidyl agonists Authors: Cervia, Davide; Zizzari, P.; Pavan, Barbara; Schuepbach, Edi; Langenegger, Daniel; Hoyer, Daniel; Biondi, Carla; Epelbaum, Jacques; Bagnoli, Paola Abstract: The physiological actions of somatostatin-14 (SRIF) receptor subtypes (sst1-sst5), which are endogenously expressed in GC cells, have not yet been elucidated, although there is evidence that sst2 receptors are negatively coupled to cytosolic free Ca2+ concentration ([Ca2+]i) and cAMP accumulation. In addition, both sst1 and sst2 receptors are negatively coupled to growth hormone (GH) secretion in GC cells. Here we report on studies concerning the expression, the pharmacology and the functional role of native SRIF receptors in GC cells with the use of five nonpeptidyl agonists, highly selective for each of the SRIF receptors. Radioligand binding studies show that sst2 and sst5 receptors are present at different relative densities, while the presence of sst3 and sst4 receptors appears to be negligible. The absence of sst1 receptor binding was unexpected in view of sst1 receptor functional effects on GH secretion. This suggests very efficient receptor-effector coupling of a low density population of sst1 receptors. Functionally, only sst2 receptors are coupled to the inhibition of [Ca2+]i and cAMP accumulation and the selective activation of sst5 receptors facilitates the stimulation of adenylyl cyclase activity through Gi/o proteins. This effect was not observed when sst2 and sst5 receptors were simultaneously activated, suggesting that there is a functional interaction between sst2 and sst5 receptors. In addition, sst1, sst2 and sst5 receptor activation inhibits GH release, further indicating that SRIF can modulate GH secretion in GC cells through mechanisms both dependent and independent on [Ca2+]i and cAMP-dependent pathways. The present data suggest SRIF-mediated functional effects in GC cells to be very diverse and provide compelling arguments to propose that multiple native SRIF receptors expressed in the same cells are not simply redundant, but contribute to marked signalling diversity. Description: L'articolo è disponibile sul sito dell'editore http://www.sciencedirect.com/ Tue, 31 Dec 2002 23:00:00 GMT http://hdl.handle.net/2067/1443 2002-12-31T23:00:00Z