title: Characterisation of the fish sst3 receptor, a member of the SRIF1 receptor family: atypical pharmacological features.
abstract: The first cloned non-mammalian somatostatin (somatostatin release-inhibiting factor = SRIF) receptor previously obtained from the teleost fish Apteronotus albifrons and generically named somatostatin receptor 3 (fsst3), was stably expressed and characterised in Chinese hamster lung fibroblast (CCL39) cells. Radioligand binding studies were performed with four radioligands selective for SRIF receptors in CCL39 cells expressing the fsst3 receptors; [125I]LTT-SRIF28 ([Leu8, D-Trp22, 125I-Tyr25]-SRIF28), [125I]Tyr10-cortistatin, [125I]CGP 23996, and [125I]Tyr3-octreotide labelled the fsst3 receptor with high affinity (pKd values: 10.47, 10.87, 9.59 and 9.57) and in a saturable manner, but defined different Bmax values; 4500, 4000, 3400 and 1500 fmol/mg, respectively. The affinities of SRIF peptides and analogues determined for fsst3 receptors displayed the following rank order of potency: seglitide = SRIF25 > SRIF14 = SRIF28 > cortistatin 14 > BIM 23014 > RC160 = L361,301 = octreotide > or = BIM 23052 > or = L362,855 > CGP23996 > BIM 23056 > BIM 23030 = cycloantagonist > SRIF22. The pharmacological profiles determined with [125I]LTT-SRIF28, [125I]CGP 23996 and [125I]Tyr10-cortistatin correlated highly significantly (r = 0.96-0.99), whereas [125I]Tyr3-octreotide binding was rather divergent (r = 0.78-0.81). Further, [125I]Tyr3-octreotide- and [125I]CGP 23996-labelled sites showed higher affinity for the various peptides than [125I]LTT-SRIF28 and [125I]Tyr10-cortistatin-labelled sites, although there were exceptions. [125I]LTT-SRIF28-binding to fsst3 receptors and human sst1-5 receptors was compared; the fsst3 binding profile correlated better with the hsst5- than with the hsst3 receptor profile. SRIF inhibited potently forskolin-stimulated adenylate cyclase activity in fsst3 transfected CCL39 cells; this effect was blocked by pertussis toxin, suggesting coupling of the fsst3 receptor to Gialpha and/or Goalpha. [125I]LTT-SRIF28 binding was detected in fish brain, liver, heart, spleen, and stomach, but not in gut. The pharmacological profile of [125I]LTT-SRIF28-labelled sites in brain, but not in liver, correlated significantly with the recombinant fsst3 receptor, in agreement with expression of the fsst3 receptor gene found by RT-PCR in the brain. However, biphasic binding curves obtained with two SRIF-analogues in brain, as well as the distinct pharmacological profile of the liver SRIF receptor, suggest the existence of several yet to be defined SRIF receptor subtypes in fish. The present data demonstrate that the recombinantly expressed fsst3 receptor has a pharmacological profile compatible with that of a SRIF1 receptor, although the rank order of affinity of fsst3 is closer to that of hsst5 than hsst3 receptors, as may be found when comparing very distantly related species. The fsst3 receptor expressed in CCL39 cells, is negatively coupled to adenylate cyclase activity via pertussis toxin-sensitive G-proteins, like mammalian sst3 receptors. Radioligand binding performed with fish tissue suggests the presence of a native sst3 receptor in brain as well as other yet to be defined SRIF receptor subtypes.
authors.affiliation: Nervous System Research, Novartis Pharma AG, Basel, Switzerland.
authors.forename: S
authors.initials: S
authors.lastname: Siehler
authors.forename: G K
authors.initials: GK
authors.lastname: Zupanc
authors.forename: K
authors.initials: K
authors.lastname: Seuwen
authors.forename: D
authors.initials: D
authors.lastname: Hoyer
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D001667:Binding, Competitive; D001921:Brain / Q000378:metabolism; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000378:metabolism; D005399:Fishes; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000378:metabolism; D008168:Lung; D009928:Organ Specificity; D010446:Peptide Fragments / Q000378:metabolism / Q000494:pharmacology; D011869:Radioligand Assay; D017481:Receptors, Somatostatin / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013004:Somatostatin / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology*; D014162:Transfection
chemical_list: D010446:Peptide Fragments; D017481:Receptors, Somatostatin; D011994:Recombinant Proteins; C101130:somatostatin receptor type 1; D013004:Somatostatin; D000242:Cyclic AMP; D000262:Adenylyl Cyclases