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Daniel Hoyer - One of the best experts on this subject based on the ideXlab platform.

  • Drug design at peptide receptors
    Journal of Molecular Neuroscience, 2002
    Co-Authors: Jason P. Hannon, Christian Bruns, Caroline Nunn, Barbara Stolz, Gisbert Weckbecker, Ian Lewis, Thomas Troxler, Konstanze Hurth, Daniel Hoyer
    Abstract:

    Somatostatin (SRIF, somatotropin release inhibiting factor), discovered for its inhibitory action on growth hormone (GH) secretion from pituitary, is an abundant neuropeptide. Two forms, SRIF_14 and SRIF_28 exist. Recently, a second family of peptides with very similar sequences and features was described; the cortistatins (CST), CST_17 and CST_29 which are brain selective. The five cloned SRIF receptors (sst_1–5) belong to the G-protein coupled/heptathelical receptor family. Structural and operational features distinguish two classes of receptors; SRIF_1-sst_2/sst_3/sst_5 (high affinity for octreotide or Seglitide) and SRIF_2=sst_1/sst_4 (very low affinity for the aforementioned ligands). The affinity of SRIF receptors for somatostatins and cortistatins is equally high, and it is not clear whether selective receptors do exist for one or the other of the peptides. Several radiologlands label all SRIF receptors, e.g., [^125I]LTT-SRIF_28, [^125I]CGP23996, [^125I]Tyr^10cortistatin or [^125I]Tyr^11SRIF_14. In contrast, [^125I]Tyr^3octreotide, [^125I]BIM23027, [^125I]MK678 or [^125I]D-Trp^8SRIF_14 label predominantly SRIF_1 sites, especially sst_2 and possibly sst_5 receptors. In brain, [^125I]Tyr^3octreotide binding equates with sst_2 receptor mRNA distribution. Native SRIF_2 receptors can be labeled with [^125I]SRIF_14 in the presence of high NaCl in brain (sst_1) or lung (sst_4) tissue. Short cyclic or linear peptide analogs show selectivity for sst_2/sst_5 (octreotide, lanreotide, BIM 23027), sst_1 (CH-275), sst_3 (sst_3-ODN-8), or sst_5 receptors (BIM 23268); although claims for selectivity have not always been confirmed. Beta peptides with affinity for SRIF receptors are also reported. The general lack of SRIF receptor antagonists is unique for peptide receptors, although CYN 154806 is a selective and potent sst_2 antagonist. Nonpeptide ligands are still rare, although a number of molecules have been reported with selectivity and potency for sst_1 (L 757,519), sst_2 (L 779,976), sst_3 (L 796,778), sst_4 (NNC 26-9100, L 803,087) or sst_1/sst_5 receptors (L 817,018). Such molecules are essential to establish the role of SRIF receptors, e.g., sst_1 in hypothalamic glutamate currents: sst_2 in inhibiting release of GH, glucagon, TSH, gastric acid secretion, pain, seizures and tumor growth, and sst_5 in vascular remodeling and inhibition of insulin and GH release.

  • drug design at peptide receptors somatostatin receptor ligands
    Journal of Molecular Neuroscience, 2002
    Co-Authors: Jason P. Hannon, Caroline Nunn, Barbara Stolz, Gisbert Weckbecker, Ian Lewis, Thomas Troxler, Konstanze Hurth, Christians Bruns, Daniel Hoyer
    Abstract:

    Somatostatin (SRIF, somatotropin release inhibiting factor), discovered for its inhibitory action on growth hormone (GH) secretion from pituitary, is an abundant neuropeptide. Two forms, SRIF14 and SRIF28 exist. Recently, a second family of peptides with very similar sequences and features was described; the cortistatins (CST), CST17 and CST29 which are brain selective. The five cloned SRIF receptors (sst1-5) belong to the G-protein coupled/ heptathelical receptor family. Structural and operational features distinguish two classes of receptors; SRIF1 - sst2/sst3/sst5 (high affinity for octreotide or Seglitide) and SRIF2 = sst1/sst4(very low affinitty for the aforementioned ligands). The affinity of SRIF receptors for somatostatins and cortistatins is equally high, and it is not clear whether selective receptors do exist for one or the other of the peptides. Several radiologlands label all SRIF receptors, e.g., [125]LTT-SRIF28' [l25I]CGP23996, [125]Tyr10cortistatin or [125I]Tyr11SRIF14. In contrast, [125I]Tyr3octreotide, [125I]BIM23027, [125I]MK678 or [125I]D-Trp8SRIF14 label predominantly SRIF1 sites, especially sst2 and possibly sst5 receptors. In brain, [125I]Tyr3octreotide binding equates with sst2 receptor mRNA distribution. Native SRIF2receptors can be labeled with [125I]SRIF14 in the presence of high NaCl in brain (sst1) or lung (sst4) tissue. Short cyclic or linear peptide analogs show selectivity for sst2/sst5 (octreotide, lanreotide, BIM 23027), sst1 (CH-275), sst3 (sst3-ODN-8), or sst5 receptors (BIM 23268); although claims for selectivity have not always been confirmed. Beta peptides ith affinity for SRIF receptors are also reported. The general lack of SRIF receptor antagonists is unique for peptide receptors, although CYN 154806 is a selective and potent sst2 antagonist. Nonpeptide ligands are still rare, although a number of molecules have been reported with selectivity and potency for sst1 (L 757,519), sst2 (L 779,976), sst3 (L 796,778), sst4 (NNC 26-9100, L 803,087) or sst1/sst5 receptors (L 817,018). Such molecules are essential to establish the role of SRIF receptors, e.g., sst1 in hypothalamic glutamate currents: sst2 in inhibiting release of GH, glucagon, TSH, gastric acid secretion, pain, seizures and tumor growth, and sst5 in vascular remodeling and inhibition of insulin and GH release.

  • Characterisation of the fish sst3 receptor, a member of the SRIF1 receptor family : atypical pharmacological features
    Neuropharmacology, 1999
    Co-Authors: Sandra Siehler, Günther K.h. Zupanc, Klaus Seuwen, Daniel Hoyer
    Abstract:

    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≥BIM 23052≥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 Giα and/or Goα. [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.

  • Somatostatin receptors in the Rhesus monkey brain: localization and pharmacological characterization
    Naunyn-Schmiedeberg's Archives of Pharmacology, 1996
    Co-Authors: V. Silke Thoss, Charles Piwko, Daniel Hoyer
    Abstract:

    To characterize the nature and distribution of somatostatin (SRIF) receptors, radioligand binding studies and in vitro receptor autoradiography were performed in Rhesus monkey brain using either [^125I]LTT-SRIF-28 ([Leu^8,D-Trp^22,^125I-Tyr^25]SRIF-28) alone or in the presence of 3 nM Seglitide (to block sst_2 sites), [^1251]Tyr^3-octreotide or [^125I] CGP 23996 (c[Asu-Lys-Asn-Phe-Phe-Trp-Lys-Thr-Tyr-Thr-Ser]) in buffer containing either 120 mM Na^+ or 5 mM Mg^2+. [^125I]Tyr^3-octreotide labelled an apparently homogeneous population of sites in cerebral and cerebellar cortex ( B _max = 27.3±2.8 fmol/mg protein and 52.6±8.6 fmol/mg protein, pK_d = 9.46±0.03 and 9.93±0.03, respectively). The pharmacological profile of these sites correlated highly significantly with that of human recombinant sst_2 receptors ( r = 0.996), but not or much less with that of human recombinant sst_3 and sst_5 receptors ( r = 0.12 and 0.45, respectively). [^125I]CGP 23996 (in Na^+-buffer) also labelled an apparently homogeneous population of sites in Rhesus monkey cerebral cortex membranes ( B _max = 3.1±0.3 fmol/mg protein, pKd = 10.57±0.08), the pharmacological profile of which was highly significantly correlated with the profiles of human recombinant sst_1 and sst_4 receptors ( r = 0.98 and 0.96, respectively). Using receptor autoradiography, high levels of [^125I]LTT-SRIF-28 and [^125I]Tyr^3-octreotide recognition sites were found in basal ganglia, molecular and granular layers of the cerebellum and layers III, V and VI of entorhinal cortex. In these regions, the addition of 3 nM Seglitide produced a marked decrease of [^125I]LTT-SRIF-28 binding. Low levels of [^125I]LTT-SRIF-28 binding were observed in subiculum, pituitary and choroid plexus. By contrast, [^125I]CGP 23996 labelling in the presence of Mg^2+ as well as Na^+ ions was highest in pituitary and choroid plexus. However, [^125I]CGP 23996 binding was diversely affected by these ionic conditions in several regions of hippocampus and cerebral cortex. Displacement of [^125I]CGP 23996 (in Mg^2+-buffer) with Seglitide in the molecular layer of the cerebellum, deep layers of the entorhinal cortex, layers I, II and V of the insular cortex and frontal pole yielded complex competition curves suggesting the presence of two populations of SRIF receptors. By contrast, [^125I]CGP 23996 binding (in Mg^2+-buffer) in the choroid plexus, hilus of the dentate gyrus and stratum oriens and radiatum of the CA3 field of hippocampus was not affected by Seglitide up to 10 μM, suggesting only sst_1 and/or sst_4 sites which have a negligible affinity for Seglitide to be present in these structures. Taken together, these results suggest that [^125I]CGP 23996 (in the presence of Na^+) labels exclusively SRIF-2 receptors (sst_1 and/or sst_4), whereas in the presence of Mg^2+ ions, [^125I]CGP 23996 labels both SRIF-2 and SRIF-1 receptors (sst_2, sst_3 and sst_5). The present study also demonstrates the presence and differential distribution of sst_2 and sst_1/sst_4 receptors in the Rhesus monkey brain.

  • Somatostatin receptors in the rhesus monkey brain: localization and pharmacological characterization.
    Naunyn-Schmiedeberg's archives of pharmacology, 1996
    Co-Authors: V. Silke Thoss, Charles Piwko, Daniel Hoyer
    Abstract:

    To characterize the nature and distribution of somatostatin (SRIF) receptors, radioligand binding studies and in vitro receptor autoradiography were performed in Rhesus monkey brain using either [125I]LTT-SRIF-28 ([Leu8,D-Trp22,125I-Tyr25]SRIF-28) alone or in the presence of 3 nM Seglitide (to block sst2 sites), [1251]Tyr3-octreotide or [125I] CGP 23996 (c[Asu-Lys-Asn-Phe-Phe-Trp-Lys-Thr-Tyr-Thr-Ser]) in buffer containing either 120 mM Na+ or 5 mM Mg2+. [125I]Tyr3-octreotide labelled an apparently homogeneous population of sites in cerebral and cerebellar cortex (Bmax = 27.3±2.8 fmol/mg protein and 52.6±8.6 fmol/mg protein, pKd = 9.46±0.03 and 9.93±0.03, respectively). The pharmacological profile of these sites correlated highly significantly with that of human recombinant sst2 receptors (r = 0.996), but not or much less with that of human recombinant sst3 and sst5 receptors (r = 0.12 and 0.45, respectively). [125I]CGP 23996 (in Na+-buffer) also labelled an apparently homogeneous population of sites in Rhesus monkey cerebral cortex membranes (Bmax = 3.1±0.3 fmol/mg protein, pKd = 10.57±0.08), the pharmacological profile of which was highly significantly correlated with the profiles of human recombinant sst1 and sst4 receptors (r = 0.98 and 0.96, respectively).

Patrick P.a. Humphrey - One of the best experts on this subject based on the ideXlab platform.

  • Differences in the operational characteristics of the human recombinant somatostatin receptor types, sst1 and sst2, in mouse fibroblast (Ltk-) cells.
    British journal of pharmacology, 1996
    Co-Authors: Sandra W. Castro, Wasyl Feniuk, G. Buell, Patrick P.a. Humphrey
    Abstract:

    1. The human recombinant somatostatin (SRIF) receptors, sst1 and sst2, have been stably expressed in mouse fibroblast (Ltk-) cells. Two stable clones, LSSR 1/20 and LSSR 11/13, expressing sst1 and sst2 receptors, respectively, have been used to characterize these receptor types using radioligand binding assays as well as measurements of changes in extracellular acidification rates using microphysiometry. 2. [125I]-[Tyr11]-SRIF bound to sst1 and sst2 receptors expressed in Ltk- cells with high affinity, Kd values being 1.52 nM, and 0.23 nM respectively. 3. In Ltk- cells expressing sst1 receptors, SRIF, SRIF-28, [D-Trp8]-SRIF and CGP 23996 all displaced [125I]-[Tyr11]-SRIF binding with high potency (IC50 values of 0.43 - 1.27 nM) whilst Seglitide, BIM-23027, BIM-23056 and L-362855 were either weak inhibitors of binding or were ineffective. 4. In contrast MK-678 (Seglitide) and BIM-23027 were the most potent inhibitors of [125I]-[Tyr11]-SRIF binding in Ltk- cells expressing sst2 receptors with IC50 values of 0.014 and 0.035 nM, respectively. 5. SRIF and a number of SRIF agonists, including Seglitide and BIM-23027, caused concentration-dependent increases in extracellular acidification rates in Ltk- cells expressing sst2 receptors but not in Ltk- cells expressing sst1 receptors. The maximum increase in acidification rate produced by SRIF was 11.3 +/- 0.7% above baseline (0.1-0.28 pH unit min-1). The relative potencies of the SRIF agonists examined in causing increases in extracellular acidification rates in Ltk- cells expressing sst2 receptors correlated well with their relative potencies in inhibiting [125I]-[Tyr11] -SRIF binding (r = 0.94). 6. The increase in extracellular acidification produced by SRIF was markedly inhibited by pretreatment of cells with pertussis toxin (100 ng ml-1) indicating the involvement of pertussis toxin-sensitive G proteins. 7. SRIF (1 microM) had no effect on basal cyclic AMP levels in Ltk- cells expressing sst1 or sst2 receptors nor did it inhibit forskolin stimulated increases in cyclic AMP levels in either cell type. 8. The results from the present study describe the operational characteristics of human sst2 receptors expressed in Ltk- cells where receptor activation causes increases in extracellular acidification rates. This receptor is coupled to a pertussis toxin-sensitive G protein. In contrast, activation of sst1 receptors, at a similar transfection density, did not cause increases in extracellular acidification rates.

  • Somatostatin sst2 receptor-mediated inhibition of parietal cell function in rat isolated gastric mucosa.
    British journal of pharmacology, 1996
    Co-Authors: Mark Wyatt, Wasyl Feniuk, Emma M. Jarvie, Patrick P.a. Humphrey
    Abstract:

    1. The aim of this study was to determine the location and functional characteristics of the somatostatin (SRIF) receptor type(s) which mediate inhibition of acid secretion in rat isolated gastric mucosa. 2. Gastrin (1 nM-1 microM), dimaprit (10 microM-300 microM) and isobutyl methylxanthine (IBMX, 1 microM-100 microM) all caused concentration-dependent increases in acid output. Responses to gastrin were almost completely inhibited by ranitidine (10 microM) at a concentration which abolished the secretory response to dimaprit. In contrast, responses to IBMX were not changed by ranitidine suggesting that IBMX acts directly on the parietal cell and not indirectly by releasing histamine from enterochromaffin-like (ECL) cells. 3. SRIF-14 (1 nM-1 microM) had no effect on basal acid output, but inhibited acid output produced by gastrin, dimaprit and IBMX in a concentration-dependent manner with respective EC50 values of 46, 54 and 167 nM. The peptidase inhibitors, amastatin (10 microM) and phosphoramidon (1 microM), had no effect on SRIF-induced inhibition of dimaprit stimulated gastric acid secretion. 4. The inhibitory effect of a range of SRIF analogues on gastrin-, dimaprit- and IBMX-induced acid secretion was also studied. Irrespective of the secretagogue used to increase acid output, the rank order of potencies was similar (BIM-23027 = Seglitide = octreotide > SRIF-14 = SRIF-28 > L-362,855). The linear peptide BIM-23056 was devoid of agonist or antagonist activity in concentrations up to 1 microM. 5. The sst2 receptor selective peptides, BIM-23027, Seglitide and octreotide were the most potent inhibitors of gastrin-, dimaprit- and IBMX-induced acid secretion suggesting that SRIF receptors resembling the recombinant sst2 receptors are involved. Furthermore, since dimaprit and IBMX stimulate gastric acid secretion independently of histamine release, sst2 receptor-mediated inhibition must occur at the level of the parietal cell itself.

  • Somatostatin-induced inhibition of neurotransmission in the mouse isolated vas deferens is resistant to pertussis toxin
    European journal of pharmacology, 1994
    Co-Authors: Wasyl Feniuk, Patrick P.a. Humphrey
    Abstract:

    The potential effects of pertussis toxin pretreatment on the inhibitory effect of somatostatin (SRIF) and the selective SRIF receptor agonist, Seglitide, were studied in mouse vas deferens and these were compared with its effect on the negative chronotropic action of carbachol in mouse atria. Somatostatin and Seglitide caused a concentration-dependent inhibition of neurogenically mediated contractile responses in the vas deferens (EC50 values of 15 and 0.6 nM respectively). There was no difference in their potencies in preparations removed from pertussis toxin pretreated mice. In contrast, the negative chronotropic action of carbachol in mouse atria was abolished by pretreatment with pertussis toxin. We conclude that, in contrast to muscarinic receptor activation in mouse atria, the inhibitory effect of somatostatin in the vas deferens is not mediated by a pertussis toxin sensitive G-protein. The high potency of Seglitide suggests that the SRIF receptor involved is of the SRIF1 type.

  • Mediation by SRIF1 receptors of the contractile action of somatostatin in rat isolated distal colon; studies using some novel SRIF analogues.
    British journal of pharmacology, 1994
    Co-Authors: E.s. Mckeen, Wasyl Feniuk, Patrick P.a. Humphrey
    Abstract:

    1. The motor effects of somatostatin-14 (SRIF), and several SRIF peptide analogues were investigated on the rat isolated distal colon. The objective of these studies was to characterize the receptor mediating the contractile action of SRIF by comparing the relative agonist potencies of a range of SRIF analogues. 2. SRIF (1 nM-1 microM) produced concentration-dependent contractions with an EC50 value of approximately 10 nM. Contractile responses induced by SRIF were insensitive to atropine (1 microM) or naloxone (1 microM) but abolished by tetrodotoxin (1 microM). Somatostatin-28 (SRIF28), also induced concentration-dependent contractions and was equipotent with SRIF. Phosphoramidon (1 microM) and amastatin (10 microM) did not increase the potency of either SRIF or SRIF28. 3. The SRIF peptide analogues, octreotide, SRIF25, Seglitide, angiopeptin and CGP23996 (1 nM-1 microM) produced contractile responses in the rat distal colon, each having similar potency and maximal activity relative to SRIF. The SSTR2 receptor-selective hexapeptide, BIM23027 (0.1 nM-1 microM), and the SRIF stereoisomer, D-Trp8-SRIF (0.1 nM-1 microM), were the most potent agonists examined being approximately 12 and 7 times more potent than SRIF, respectively. In contrast, the SSTR5 receptor-selective analogue, L362,855, was approximately 120 times weaker than SRIF, whilst the SSTR3 receptor-selective analogue, BIM23056, was inactive at concentrations up to 3 microM. 4. The putative SRIF receptor antagonist, (cyclo(7-aminoheptanoyl Phe-D-Trp-Lys-Thr[Bzl]))(CPP) (1 microM), had no agonist activity and had no effect on contractions induced by SRIF. 5. The contractile actions of BIM23027 and Seglitide were subject to pronounced desensitization. Desensitization of preparations by BIM23027 (0.3 JIM) abolished the contractile action of SRIF andSRIF28 but had no effect on contractions produced by acetylcholine (0.1 nM-I1M), suggesting thatBIM23027, SRIF and SRIF28 act via a common receptor mechanism.6. In conclusion, the rat isolated distal colon contracts in response to SRIF and a number of SRIF analogues. Seglitide and octreotide exhibited similar potency and maximal activity relative to SRIF,suggesting that in the rat colon the receptor mediating contraction belongs to the SRIF,-receptor group,of which the recombinant SSTR2, SSTR3 and SSTR5 receptors appear to be subtypes. The high potency of BIM23027, the weak agonist activity of L362,855 and the lack of activity exhibited by BIM23056suggests that the SRIF receptor mediating contraction in the rat distal colon is similar to there combinant SSTR2 receptor.

  • Antagonist effects of Seglitide (MK 678) at somatostatin receptors in guinea-pig isolated right atria
    British journal of pharmacology, 1993
    Co-Authors: J. Dimech, Wasyl Feniuk, Patrick P.a. Humphrey
    Abstract:

    Somatostatin (SS) exerts a negative inotropic effect in isolated atria. Here we report that in guinea-pig isolated right atria, Seglitide, a potent cyclic hexapeptide somatostatin agonist, behaves as a competitive somatostatin receptor antagonist with pA2 values against SS14, SS25 and SS28, of 6.50 +/- 0.40, 6.24 +/- 0.08 and 6.09 +/- 0.06, respectively. Seglitide had little or no effect on the negative inotropic action of carbachol or N6-cyclohexyladenosine. Our findings indicate that the receptor-response coupling characteristics of guinea-pig atria are such that in this preparation Seglitide has low intrinsic activity and behaves specifically as a somatostatin receptor antagonist.

Wasyl Feniuk - One of the best experts on this subject based on the ideXlab platform.

  • Differences in the operational characteristics of the human recombinant somatostatin receptor types, sst1 and sst2, in mouse fibroblast (Ltk-) cells.
    British journal of pharmacology, 1996
    Co-Authors: Sandra W. Castro, Wasyl Feniuk, G. Buell, Patrick P.a. Humphrey
    Abstract:

    1. The human recombinant somatostatin (SRIF) receptors, sst1 and sst2, have been stably expressed in mouse fibroblast (Ltk-) cells. Two stable clones, LSSR 1/20 and LSSR 11/13, expressing sst1 and sst2 receptors, respectively, have been used to characterize these receptor types using radioligand binding assays as well as measurements of changes in extracellular acidification rates using microphysiometry. 2. [125I]-[Tyr11]-SRIF bound to sst1 and sst2 receptors expressed in Ltk- cells with high affinity, Kd values being 1.52 nM, and 0.23 nM respectively. 3. In Ltk- cells expressing sst1 receptors, SRIF, SRIF-28, [D-Trp8]-SRIF and CGP 23996 all displaced [125I]-[Tyr11]-SRIF binding with high potency (IC50 values of 0.43 - 1.27 nM) whilst Seglitide, BIM-23027, BIM-23056 and L-362855 were either weak inhibitors of binding or were ineffective. 4. In contrast MK-678 (Seglitide) and BIM-23027 were the most potent inhibitors of [125I]-[Tyr11]-SRIF binding in Ltk- cells expressing sst2 receptors with IC50 values of 0.014 and 0.035 nM, respectively. 5. SRIF and a number of SRIF agonists, including Seglitide and BIM-23027, caused concentration-dependent increases in extracellular acidification rates in Ltk- cells expressing sst2 receptors but not in Ltk- cells expressing sst1 receptors. The maximum increase in acidification rate produced by SRIF was 11.3 +/- 0.7% above baseline (0.1-0.28 pH unit min-1). The relative potencies of the SRIF agonists examined in causing increases in extracellular acidification rates in Ltk- cells expressing sst2 receptors correlated well with their relative potencies in inhibiting [125I]-[Tyr11] -SRIF binding (r = 0.94). 6. The increase in extracellular acidification produced by SRIF was markedly inhibited by pretreatment of cells with pertussis toxin (100 ng ml-1) indicating the involvement of pertussis toxin-sensitive G proteins. 7. SRIF (1 microM) had no effect on basal cyclic AMP levels in Ltk- cells expressing sst1 or sst2 receptors nor did it inhibit forskolin stimulated increases in cyclic AMP levels in either cell type. 8. The results from the present study describe the operational characteristics of human sst2 receptors expressed in Ltk- cells where receptor activation causes increases in extracellular acidification rates. This receptor is coupled to a pertussis toxin-sensitive G protein. In contrast, activation of sst1 receptors, at a similar transfection density, did not cause increases in extracellular acidification rates.

  • Somatostatin sst2 receptor-mediated inhibition of parietal cell function in rat isolated gastric mucosa.
    British journal of pharmacology, 1996
    Co-Authors: Mark Wyatt, Wasyl Feniuk, Emma M. Jarvie, Patrick P.a. Humphrey
    Abstract:

    1. The aim of this study was to determine the location and functional characteristics of the somatostatin (SRIF) receptor type(s) which mediate inhibition of acid secretion in rat isolated gastric mucosa. 2. Gastrin (1 nM-1 microM), dimaprit (10 microM-300 microM) and isobutyl methylxanthine (IBMX, 1 microM-100 microM) all caused concentration-dependent increases in acid output. Responses to gastrin were almost completely inhibited by ranitidine (10 microM) at a concentration which abolished the secretory response to dimaprit. In contrast, responses to IBMX were not changed by ranitidine suggesting that IBMX acts directly on the parietal cell and not indirectly by releasing histamine from enterochromaffin-like (ECL) cells. 3. SRIF-14 (1 nM-1 microM) had no effect on basal acid output, but inhibited acid output produced by gastrin, dimaprit and IBMX in a concentration-dependent manner with respective EC50 values of 46, 54 and 167 nM. The peptidase inhibitors, amastatin (10 microM) and phosphoramidon (1 microM), had no effect on SRIF-induced inhibition of dimaprit stimulated gastric acid secretion. 4. The inhibitory effect of a range of SRIF analogues on gastrin-, dimaprit- and IBMX-induced acid secretion was also studied. Irrespective of the secretagogue used to increase acid output, the rank order of potencies was similar (BIM-23027 = Seglitide = octreotide > SRIF-14 = SRIF-28 > L-362,855). The linear peptide BIM-23056 was devoid of agonist or antagonist activity in concentrations up to 1 microM. 5. The sst2 receptor selective peptides, BIM-23027, Seglitide and octreotide were the most potent inhibitors of gastrin-, dimaprit- and IBMX-induced acid secretion suggesting that SRIF receptors resembling the recombinant sst2 receptors are involved. Furthermore, since dimaprit and IBMX stimulate gastric acid secretion independently of histamine release, sst2 receptor-mediated inhibition must occur at the level of the parietal cell itself.

  • Somatostatin-induced inhibition of neurotransmission in the mouse isolated vas deferens is resistant to pertussis toxin
    European journal of pharmacology, 1994
    Co-Authors: Wasyl Feniuk, Patrick P.a. Humphrey
    Abstract:

    The potential effects of pertussis toxin pretreatment on the inhibitory effect of somatostatin (SRIF) and the selective SRIF receptor agonist, Seglitide, were studied in mouse vas deferens and these were compared with its effect on the negative chronotropic action of carbachol in mouse atria. Somatostatin and Seglitide caused a concentration-dependent inhibition of neurogenically mediated contractile responses in the vas deferens (EC50 values of 15 and 0.6 nM respectively). There was no difference in their potencies in preparations removed from pertussis toxin pretreated mice. In contrast, the negative chronotropic action of carbachol in mouse atria was abolished by pretreatment with pertussis toxin. We conclude that, in contrast to muscarinic receptor activation in mouse atria, the inhibitory effect of somatostatin in the vas deferens is not mediated by a pertussis toxin sensitive G-protein. The high potency of Seglitide suggests that the SRIF receptor involved is of the SRIF1 type.

  • Mediation by SRIF1 receptors of the contractile action of somatostatin in rat isolated distal colon; studies using some novel SRIF analogues.
    British journal of pharmacology, 1994
    Co-Authors: E.s. Mckeen, Wasyl Feniuk, Patrick P.a. Humphrey
    Abstract:

    1. The motor effects of somatostatin-14 (SRIF), and several SRIF peptide analogues were investigated on the rat isolated distal colon. The objective of these studies was to characterize the receptor mediating the contractile action of SRIF by comparing the relative agonist potencies of a range of SRIF analogues. 2. SRIF (1 nM-1 microM) produced concentration-dependent contractions with an EC50 value of approximately 10 nM. Contractile responses induced by SRIF were insensitive to atropine (1 microM) or naloxone (1 microM) but abolished by tetrodotoxin (1 microM). Somatostatin-28 (SRIF28), also induced concentration-dependent contractions and was equipotent with SRIF. Phosphoramidon (1 microM) and amastatin (10 microM) did not increase the potency of either SRIF or SRIF28. 3. The SRIF peptide analogues, octreotide, SRIF25, Seglitide, angiopeptin and CGP23996 (1 nM-1 microM) produced contractile responses in the rat distal colon, each having similar potency and maximal activity relative to SRIF. The SSTR2 receptor-selective hexapeptide, BIM23027 (0.1 nM-1 microM), and the SRIF stereoisomer, D-Trp8-SRIF (0.1 nM-1 microM), were the most potent agonists examined being approximately 12 and 7 times more potent than SRIF, respectively. In contrast, the SSTR5 receptor-selective analogue, L362,855, was approximately 120 times weaker than SRIF, whilst the SSTR3 receptor-selective analogue, BIM23056, was inactive at concentrations up to 3 microM. 4. The putative SRIF receptor antagonist, (cyclo(7-aminoheptanoyl Phe-D-Trp-Lys-Thr[Bzl]))(CPP) (1 microM), had no agonist activity and had no effect on contractions induced by SRIF. 5. The contractile actions of BIM23027 and Seglitide were subject to pronounced desensitization. Desensitization of preparations by BIM23027 (0.3 JIM) abolished the contractile action of SRIF andSRIF28 but had no effect on contractions produced by acetylcholine (0.1 nM-I1M), suggesting thatBIM23027, SRIF and SRIF28 act via a common receptor mechanism.6. In conclusion, the rat isolated distal colon contracts in response to SRIF and a number of SRIF analogues. Seglitide and octreotide exhibited similar potency and maximal activity relative to SRIF,suggesting that in the rat colon the receptor mediating contraction belongs to the SRIF,-receptor group,of which the recombinant SSTR2, SSTR3 and SSTR5 receptors appear to be subtypes. The high potency of BIM23027, the weak agonist activity of L362,855 and the lack of activity exhibited by BIM23056suggests that the SRIF receptor mediating contraction in the rat distal colon is similar to there combinant SSTR2 receptor.

  • Antagonist effects of Seglitide (MK 678) at somatostatin receptors in guinea-pig isolated right atria
    British journal of pharmacology, 1993
    Co-Authors: J. Dimech, Wasyl Feniuk, Patrick P.a. Humphrey
    Abstract:

    Somatostatin (SS) exerts a negative inotropic effect in isolated atria. Here we report that in guinea-pig isolated right atria, Seglitide, a potent cyclic hexapeptide somatostatin agonist, behaves as a competitive somatostatin receptor antagonist with pA2 values against SS14, SS25 and SS28, of 6.50 +/- 0.40, 6.24 +/- 0.08 and 6.09 +/- 0.06, respectively. Seglitide had little or no effect on the negative inotropic action of carbachol or N6-cyclohexyladenosine. Our findings indicate that the receptor-response coupling characteristics of guinea-pig atria are such that in this preparation Seglitide has low intrinsic activity and behaves specifically as a somatostatin receptor antagonist.

Charles Piwko - One of the best experts on this subject based on the ideXlab platform.

  • Somatostatin receptors in the Rhesus monkey brain: localization and pharmacological characterization
    Naunyn-Schmiedeberg's Archives of Pharmacology, 1996
    Co-Authors: V. Silke Thoss, Charles Piwko, Daniel Hoyer
    Abstract:

    To characterize the nature and distribution of somatostatin (SRIF) receptors, radioligand binding studies and in vitro receptor autoradiography were performed in Rhesus monkey brain using either [^125I]LTT-SRIF-28 ([Leu^8,D-Trp^22,^125I-Tyr^25]SRIF-28) alone or in the presence of 3 nM Seglitide (to block sst_2 sites), [^1251]Tyr^3-octreotide or [^125I] CGP 23996 (c[Asu-Lys-Asn-Phe-Phe-Trp-Lys-Thr-Tyr-Thr-Ser]) in buffer containing either 120 mM Na^+ or 5 mM Mg^2+. [^125I]Tyr^3-octreotide labelled an apparently homogeneous population of sites in cerebral and cerebellar cortex ( B _max = 27.3±2.8 fmol/mg protein and 52.6±8.6 fmol/mg protein, pK_d = 9.46±0.03 and 9.93±0.03, respectively). The pharmacological profile of these sites correlated highly significantly with that of human recombinant sst_2 receptors ( r = 0.996), but not or much less with that of human recombinant sst_3 and sst_5 receptors ( r = 0.12 and 0.45, respectively). [^125I]CGP 23996 (in Na^+-buffer) also labelled an apparently homogeneous population of sites in Rhesus monkey cerebral cortex membranes ( B _max = 3.1±0.3 fmol/mg protein, pKd = 10.57±0.08), the pharmacological profile of which was highly significantly correlated with the profiles of human recombinant sst_1 and sst_4 receptors ( r = 0.98 and 0.96, respectively). Using receptor autoradiography, high levels of [^125I]LTT-SRIF-28 and [^125I]Tyr^3-octreotide recognition sites were found in basal ganglia, molecular and granular layers of the cerebellum and layers III, V and VI of entorhinal cortex. In these regions, the addition of 3 nM Seglitide produced a marked decrease of [^125I]LTT-SRIF-28 binding. Low levels of [^125I]LTT-SRIF-28 binding were observed in subiculum, pituitary and choroid plexus. By contrast, [^125I]CGP 23996 labelling in the presence of Mg^2+ as well as Na^+ ions was highest in pituitary and choroid plexus. However, [^125I]CGP 23996 binding was diversely affected by these ionic conditions in several regions of hippocampus and cerebral cortex. Displacement of [^125I]CGP 23996 (in Mg^2+-buffer) with Seglitide in the molecular layer of the cerebellum, deep layers of the entorhinal cortex, layers I, II and V of the insular cortex and frontal pole yielded complex competition curves suggesting the presence of two populations of SRIF receptors. By contrast, [^125I]CGP 23996 binding (in Mg^2+-buffer) in the choroid plexus, hilus of the dentate gyrus and stratum oriens and radiatum of the CA3 field of hippocampus was not affected by Seglitide up to 10 μM, suggesting only sst_1 and/or sst_4 sites which have a negligible affinity for Seglitide to be present in these structures. Taken together, these results suggest that [^125I]CGP 23996 (in the presence of Na^+) labels exclusively SRIF-2 receptors (sst_1 and/or sst_4), whereas in the presence of Mg^2+ ions, [^125I]CGP 23996 labels both SRIF-2 and SRIF-1 receptors (sst_2, sst_3 and sst_5). The present study also demonstrates the presence and differential distribution of sst_2 and sst_1/sst_4 receptors in the Rhesus monkey brain.

  • Somatostatin receptors in the rhesus monkey brain: localization and pharmacological characterization.
    Naunyn-Schmiedeberg's archives of pharmacology, 1996
    Co-Authors: V. Silke Thoss, Charles Piwko, Daniel Hoyer
    Abstract:

    To characterize the nature and distribution of somatostatin (SRIF) receptors, radioligand binding studies and in vitro receptor autoradiography were performed in Rhesus monkey brain using either [125I]LTT-SRIF-28 ([Leu8,D-Trp22,125I-Tyr25]SRIF-28) alone or in the presence of 3 nM Seglitide (to block sst2 sites), [1251]Tyr3-octreotide or [125I] CGP 23996 (c[Asu-Lys-Asn-Phe-Phe-Trp-Lys-Thr-Tyr-Thr-Ser]) in buffer containing either 120 mM Na+ or 5 mM Mg2+. [125I]Tyr3-octreotide labelled an apparently homogeneous population of sites in cerebral and cerebellar cortex (Bmax = 27.3±2.8 fmol/mg protein and 52.6±8.6 fmol/mg protein, pKd = 9.46±0.03 and 9.93±0.03, respectively). The pharmacological profile of these sites correlated highly significantly with that of human recombinant sst2 receptors (r = 0.996), but not or much less with that of human recombinant sst3 and sst5 receptors (r = 0.12 and 0.45, respectively). [125I]CGP 23996 (in Na+-buffer) also labelled an apparently homogeneous population of sites in Rhesus monkey cerebral cortex membranes (Bmax = 3.1±0.3 fmol/mg protein, pKd = 10.57±0.08), the pharmacological profile of which was highly significantly correlated with the profiles of human recombinant sst1 and sst4 receptors (r = 0.98 and 0.96, respectively).

  • Localization and pharmacological characterization of somatostatin recognition sites in the human cerebellum.
    Neuropharmacology, 1996
    Co-Authors: Charles Piwko, Alphonse Probst, Daniel Hoyer
    Abstract:

    Abstract Radioligand binding studies were performed in membranes of human cerebellum using [ 125 I][Tyr 3 ]octreotide also known as [ 125 I]204-090, [ 125 I]LTT-SRIF-28 ([Leu 8 , d -Trp 22 , 125 I-Tyr 25 ]SRIF-28) and [ 125 I]CGP 23996 ([ 125 I]c[Asu-Lys-Asn-Phe-Phe-Trp-Lys-Thr-Tyr-Thr-Ser]) to characterize the nature of cerebellar somatostatin receptors. Saturation experiments performed with [ 125 I]204-090 suggest the presence of a single class of binding sites with high affinity: B max = 55.7 ± 9.7 fmol/mg protein , p K d = 9.57 ± 0.04. The pharmacological profile of [ 125 I]204-090 and [ 125 I]LTT-SRIF-28 labelled sites in human cerebellar membranes was overlapping (correlation coefficient r = 0.998) and correlated very significantly with that of recombinant human sst 2 receptors ( r = 0.987). By contrast, there was very little correlation with those of recombinant human sst 3 ( r = 0.208) or human sst 5 receptors ( r = 0.547). In contrast to [ 125 I]204-090 or [ 125 I]LTT-SRIF-28 binding, [ 125 I]CGP 23996 binding (in 5 mM MgCl 2 buffer) in cerebellar membranes was heterogeneous as indicated by biphasic competition curves produced by sst 2 receptor selective ligands such as Seglitide or octreotide. The pharmacological profile of the major component was closely correlated with that of human sst 2 receptors ( r = 0.989), whereas the minor component correlated equally well with human sst 1 or sst 4 receptors ( r = 0.902 and 0.941, respectively). In vitro autoradiographic studies performed in cerebellar slices using [ 125 I]204-090 and [ 125 I]LTT-SRIF-28 demonstrated the presence of binding sites predominantly in the molecular layer, whereas weaker labelling was detected in the granular layer. The distribution of sites labelled by both radioligands was very similar. Using [ 125 I]CGP 23996 (in 120 mM NaCl buffer), no clear difference between labeling of the molecular and granular layers was detectable; the dentate nucleus demonstrated binding sites for [ 125 I]CGP 23996, in contrast to the very low level of binding observed with both, [ 125 I]204-090 and [ 125 I]LTT-SRIF-28. Together, the present data demonstrate the presence of SRIF receptors in the adult human cerebellar cortex which are, for the major population, best characterized as sst 2 . The SRIF receptors in the minor population of the cerebellar cortex and the dentate nucleus most probably represent sst 1 and/or sst 4 sites.

  • Localization and pharmacological characterization of somatostatin sst_2 sites in the rat cerebellum
    Naunyn-Schmiedeberg's Archives of Pharmacology, 1995
    Co-Authors: Charles Piwko, V. Silke Thoss, Daniel Hoyer
    Abstract:

    Radioligand binding studies were performed in membranes of rat cerebellum using [^125I]-[Tyr^3]octreotide ([^125I]204-090) to characterize the nature of cerebellar somatostatin receptors. Saturation experiments suggest the presence of a single class of binding sites with high affinity, pK_d = 9.53 ± 0.11, but low receptor density, B _max = 12.7 ± 1.0 fmol/mg protein. The pharmacological profile of [^125I]204-090 sites in cerebellar membranes was established using a range of ligands known to interact with SSTR-2 (now called sst_2) and other somatostatin (SRIF) receptors. SRIF analogues such as octreotide (SMS 201-995), Seglitide (MK 678) and somatuline (BIM 23014) displayed very high affinity for cerebellar [^125I]204-090 binding sites. The data were compared to results obtained using the same ligand in rat cerebral cortex membranes known to represent sst_2 binding. The pharmacological characteristics of the cerebellar sites were in close correlation with those of the cerebral cortex ( r = 0.976, n = 19, p < 0.001) and CHO-cells expressing human recombinant sst_2 receptor ( r = 0.977, n = 19, p < 0.001). By contrast, there was very little correlation between cerebellar binding and published affinities for rat sst_5 receptors ( r = 0.465), for which octreotide has also high affinity. In vitro autoradiographic studies performed in cerebellar slices using [^125I]204-090 demonstrated the presence of binding sites in the molecular layer of the rat cerebellum. In situ hybridization studies using sst_2 receptor mRNA selective oligoprobes confirmed the presence of sst_2 receptor mRNA in the rat cerebellum. Together, the present data demonstrate the presence of a low density of SRIF receptors in the molecular layer of the adult rat cerebellum which are best characterized as sst_2. This is the first pharmacological characterization and localization of sst_2 receptors in the adult rat cerebellum.

Julio Perez - One of the best experts on this subject based on the ideXlab platform.

  • functional effects of d phe c cys tyr d trp lys val cys trp nh2 and differential changes in somatostatin receptor messenger rnas binding sites and somatostatin release in kainic acid treated rats
    Neuroscience, 1995
    Co-Authors: Julio Perez, Annamaria Vezzani, Gianluca Civenni, Piotr Tutka, Massimo Rizzi, E Schupbach, Daniel Hoyer
    Abstract:

    In situ hybridization histochemistry for somatostatin receptors-1, -2, -3 and -4 section and receptor autoradiography using [125I]CGP 23996, [125I]somatostatin-28, [125I]Seglitide and [125I]Tyr3 octreotide were carried out to determine the expression of somatostatin receptor messenger RNAs and binding sites in the hippocampus and cerebral cortex of rats 21 days following generalized limbic seizures induced by subcutaneous injection of 12mg/kg kainic acid. In control rats, somatostatin-1 to somatostatin-4 receptor messenger RNAs were found in the pyramidal layer and granule cell layer of the dentate gyrus. After kainate treatment, the CA1 subfield displayed a selective decrease in somatostatin-3 and somatostatin-4 receptor hybridization signals of 35 and 41%, respectively, whereas no changes were observed in the remaining hippocampal areas. Somatostatin-1 and somatostatin-2 receptor messenger RNA expression in the hippocampus remained unaffected by kainate treatment. No effect of kainate was observed in the expression of somatostatin receptor messenger RNAs in the cerebral cortex. In control rats, the selective somatostatin-2 receptor ligands, [125I]Seglitide and [125I]Tyr3 octreotide and the non-selective somatostatin receptor ligands [125I]CGP 23996 and [125I]somatostatin-28, labelled preferentially the stratum oriens and radiatum CA1, the granule and molecular layers of the dentate gyrus and the deep layers of the cerebral cortex. [125I]somatostatin-28 and [125I]CGP 23996 labelled sites were selectively decreased by 32 and 39%, respectively, in the stratum radiatum CA1 after kainate treatment. [125I]CGP 23996 binding was also decreased by 35% in the stratum oriens CA1 and by 36% on average in the stratum oriens and radiatum CA3. [125I]Seglitide and [125I]Tyr3 octreotide binding was not affected by kainate in any hippocampal region. The granule and molecular layers of the hippocampus and the layers IV-VI of the cerebral cortex did not show changes in binding sites for any of the radioligands analysed. A 18 and 35% decrease in the spontaneous and 50 mM KCl-induced somatostatin release from hippocampal slices was found two days after kainate, a likely reflection of neuronal cell loss. No differences in somatostatin release were observed 21 days after kainate treatment. At this latter time, the rats had an enhanced susceptibility to tonic-clonic seizures induced by intraperitoneal injection of 30 mg/kg pentylenetetrazol, a subconvulsant dose in naive rats. Bilateral infusion of 6 micrograms RC 160, a selective somatostatin-2 receptor agonist, in the dentate gyrus 21 days after kainate, significantly reduced (P < 0.05) the number of animals with tonic-clonic seizures induced by pentylenetetrazol.(ABSTRACT TRUNCATED AT 400 WORDS)

  • Characterization and distribution of somatostatin SS-1 and SRIF-1 binding sites in rat brain: identitity with SSTR-2 receptors
    European journal of pharmacology, 1995
    Co-Authors: Philippe Schoeffter, Julio Perez, E Schupbach, Daniel Langenegger, Ionel Bobirnac, Hermann Lübbert, Christian Bruns, Daniel Hoyer
    Abstract:

    Somatostatin (SRIF) SS-1 binding sites were initially defined in radioligand binding studies performed in rat brain cerebral cortex membranes using [125I]204-090 (a radiolabelled Tyr3 analogue of SMS 201-995, octreotide). SRIF-1 recognition sites were defined in binding studies performed with [125I]MK 678 (Seglitide). Both SS-1 and SRIF-1 sites were characterized by their high affinity for SRIF-14, SRIF-28 and for cyclic peptides such as octreotide and Seglitide, in marked contrast to SS-2 and SRIF-2 sites which have very low affinity for these synthetic SRIF analogues. In the present study, SS-1 and SRIF-1 radioligand binding studies were performed in rat cortex membranes and compared to results obtained in cloned Chinese hamster ovary cells expressing human SSTR-2 receptors using [125I]204-090 and/or [125I]MK-678. The rank orders of affinity of a variety of SRIF analogues and synthetic peptides for SS-1/SRIF-1 binding sites and recombinant SSTR-2 receptors were very similar and correlated highly significantly (r = 0.94–0.99); by contrast, correlation between SS-1 and SSTR-5 (r = 0.44) or SSTR-3 binding (r = 0.07) was not significant. Autoradiographic studies were performed in rat brain using both radioligands [125I]204-090 and [125I]MK-678 and compared with the distribution of SSTR-2 receptor mRNA determined using in situ hybridization. A clear overlap was observed between the distribution of SSTR-2 mRNA and binding sites labelled with both radioligands. SSTR-2 receptor-mediated inhibition of forskolin-stimulated adenylate cyclase in Chines hamster ovary cells by a variety of SRIF analogues and short synthetic peptides displayed a rank order of potency highly similar to their rank order of affinity at SS-1/SRIF-1 binding sites. It is concluded that SS-1 and SRIF-1 binding sites respectively labelled with [125I]204-090 and [125I]MK 678, both display the pharmacological profile of SSTR-2 receptors, that the distribution of [125I]204-090 and [125I]MK-678 binding sites in rat brain is superimposable and largely comparable to that of SSTR-2 mRNA expression. It is also shown that neither [125I]204-090 nor [125I]MK-678 label SSTR-3 or SSTR-5 receptors in rat brain. Finally, it is demonstrated that SSTR-2 receptors can very efficiently couple to adenylate cyclase activity in an inhibitory manner.

  • Pharmacological identity between somatostatin SS-2 binding sites and SSTR-1 receptors
    European journal of pharmacology, 1995
    Co-Authors: Daniel Hoyer, Julio Perez, E Schupbach, Philippe Schoeffter, Daniel Langenegger, Hermann Lübbert, Christian Bruns, Klemens Kaupmann, Jean Claude Reubi
    Abstract:

    Somatostatin (SRIF) SS-2 binding sites were originally defined in rat brain cerebral cortex membranes using [125I]Tyr11-SRIF-14 in the presence of 120 mM NaCl. These sites were characterized by their high affinity for SRIF-14 and SRIF-28, but very low affinity for cyclic peptides such as octreotide (SMS 201-995) and Seglitide (MK 678). The characteristics of SS-2 sites are reminiscent of 125I]CGP 23996-labelled sites in rat brain which have been termed SRIF-2 sites. In the present study, the pharmacological profile of SS-2 sites was determined in radioligand binding studies performed in rat cortex membranes using [125I]SRIF-14 in the presence of 120 mM NaCl and compared to that of human SSTR-1 receptors expressed in human embryonic kidney (HEK 293) cells, using [125I]SRIF-14. The rank orders of affinity of a variety of SRIF analogues and synthetic peptides for SS-2 binding sites and recombinant human SSTR-1 receptors were very similar and correlated highly significantly (r = 0.99). However, SS-2 binding correlated also with binding to recombinant SSTR-4 receptors (r = 0.91). Autoradiographic studies were performed using the radioligand [125I]CGP 23996 which has been claimed to label selectively SRIF-2 binding sites and compared with the distribution of SSTR-1 receptor mRNA determined using in situ hybridization in rat brain. Although some overlap was observed between the distribution of SSTR-1 mRNA and [125I]CGP 23996 binding sites, the latter were clearly more widespread, suggesting this ligand to label SSTR-1 and other sites. In addition, inhibition of forskolin-stimulated adenylate cyclase was investigated in HEK 293 cells transfected with human SSTR-1 receptors; a variety of SRIF analogues and short synthetic peptides behaved as agonists at adenylate cyclase and displayed a rank order of potency highly similar to that observed for these compounds at SS-2 binding sites. Seglitide acted as an antagonist at SSTR-1 receptor mediated inhibition of adenylate cyclase activity with a pKB of 4.42. It is concluded that the pharmacological profile of SS-2 binding sites resembles most closely that of SSTR-1 receptors (although similarities with SSTR-4 receptors were observed), that [125I]CGP 23996 labels presumably several SRIF receptors in rat brain, and that SSTR-1 receptors are negatively and efficiently coupled to adenylate cyclase activity.