Effect of the bombesin receptor blockers [Leu13,ψCH2NH-Leu14]bombesin and N-pivaloyl GRP(20–25) alkylamide (L 686,095-001C002) on basal and neuromedin C-stimulated PRL and GH release in pituitary cell aggregates

doi:10.1016/0196-9781(91)90028-N

Peptides

Volume 12, Issue 2, March–April 1991, Pages 371-374

https://doi.org/10.1016/0196-9781(91)90028-N Get rights and content

Abstract

Perifusion of rat anterior pituitary cell aggregates, cultured in estrogen-supplemented serum-free medium with 1 nM of the bombesin (BBN)-like peptide, neuromedin C (NMC), significantly stimulates GH and PRL release. This effect is dose-dependently inhibited by the BBN receptor blocker L 686,095-001C002 [an N-pivaloyl-gastrin-releasing-peptide(20–25) alkylamide]. The IC₅₀ was 0.20 nM in the case of the GH response and 0.16 nM in the case of the PRL response. The antagonist has no effect on basal PRL or GH release. [Leu¹³,ψCH₂NH-Leu¹⁴]BBN (ψBBN) displays an IC₅₀ of 0.41 μM for inhibiting the GH response and 0.36 μM for inhibiting the PRL response to NMC. At a concentration of 0.5 μM or 5 μM, however, the latter antagonist stimulates PRL and GH release when perifused alone. This stimulatory effect is dose dependent, augments when aggregates are cultured in 1 nM E₂ (as is the case for NMC) and is abolished by 2 nM L 686,095-001C002. It is concluded that L 686,095-001C002 is a potent and pure antagonist of pituitary BBN receptors mediating PRL and GH release, whereas ψBBN is a relatively weak antagonist with considerable partial agonist activity.

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Cited by (12)

Insights into bombesin receptors and ligands: Highlighting recent advances
2015, Peptides
This following article is written for Prof. Abba Kastin's Festschrift, to add to the tribute to his important role in the advancement of the role of peptides in physiological, as well as pathophysiological processes. There have been many advances during the 35 years of his prominent role in the Peptide field, not only as editor of the journal Peptides, but also as a scientific investigator and editor of two volumes of the Handbook of Biological Active Peptides [146], [147]. Similar to the advances with many different peptides, during this 35 year period, there have been much progress made in the understanding of the pharmacology, cell biology and the role of (bombesin) Bn receptors and their ligands in various disease states, since the original isolation of bombesin from skin of the European frog Bombina bombina in 1970 [76]. This paper will briefly review some of these advances over the time period of Prof. Kastin 35 years in the peptide field concentrating on the advances since 2007 when many of the results from earlier studies were summarized [128], [129]. It is appropriate to do this because there have been 280 articles published in Peptides during this time on bombesin-related peptides and it accounts for almost 5% of all publications. Furthermore, 22 Bn publications we have been involved in have been published in either Peptides [14], [39], [55], [58], [81], [92], [93], [119], [152], [216], [225], [226], [231], [280], [302], [309], [355], [361], [362] or in Prof. Kastin's Handbook of Biological Active Peptides [137], [138], [331].
Characterization of putative GRP- and NMB-receptor antagonist's interaction with human receptors
2009, Peptides
The mammalian bombesin (Bn) peptides neuromedin B (NMB) and gastrin-releasing peptide (GRP) actions are mediated by two receptors (NMB-receptor, GRP-receptor) which are widely distributed in the GI tract and CNS. From primarily animal studies NMB/GRP-receptor activation has physiological/pathophysiological effects in the CNS and GI tract including stimulating of growth of cancers and normal tissues. Whereas these Bn-receptors’ effects have been extensively studied in nonhuman cells and animals, little is known of the physiological/pathological role(s) in humans, largely due to lack of potent antagonists. To address this issue we compared NMB/GRP-receptor affinity/potency of 10 chemical classes of putative antagonists (35 compounds) for human Bn-receptors by performing binding studies or assessing abilities to activate hGRP/hNMB-receptor [assessing phospholipase C activation] in four different cells containing native Bn-receptors or transfected receptors. From binding studies 23 were GRP-receptor-preferring, 4 were NMB-receptor, and 8 nonselective. For the hGRP-receptor-preferring analogues none showed hGRP-receptor agonist activity, but 13 were full or partial hNMB-receptor agonists at hNMB-receptors. For hNMB-receptor-preferring analogues none were agonists. Analogue #24 ([(3-Ph-Pr⁶), His⁷, d-Ala¹¹, d-Pro¹³, Ψ(13-14), Phe¹⁴]Bn(6-14)NH₂) and analogue #7 [d-Phe⁶, Leu¹³, Ψ(CH₂NH), Cpa¹⁴]Bn(6-14) were the most potent (0.2–1.4 nM) and selective (>10,000-fold) for the hGRP-receptor with analogue #7.5 [d-Tpi⁶, Leu¹³, Ψ(CH₂NH), Leu¹⁴]Bn(6-14)[RC-3095] (0.2–1.4 nM) slightly less selective. Analogue #34 (PD168368) had the highest affinity for hNMB-receptor (1.32–1.58 nM) and the greatest selectivity (2298–6952-fold) for the hNMB-receptor. These results demonstrate numerous putative hGRP/hNMB-receptor antagonists identified in nonhuman cells and/or animals have agonist activity at the hNMB-receptor, limiting their potential usefulness. However, a number were identified which were potent/selective for human Bn-receptors and should be useful for investigating their roles in human physiological/pathophysiological conditions.
Bombesin regulation of adrenocorticotropin release from ovine anterior pituitary cells
1997, Peptides
AU, C. L., B. J. CANNY, P. G. FARNWORTH AND A. S. GIRAUD.
Bombesin regulation of adrenocorticotropin release from ovine anterior pituitary cells.
PEPTIDES 18(7) 995–1000, 1997.—Mammalian members of the bombesin-like peptide family (gastrin releasing peptides; GRP) have been localized in the ovine median eminence and in hypophysial-portal blood, suggesting a role in the regulation of anterior pituitary function. In this study we have shown that although bombesin cannot stimulate ACTH secretion alone, it potentiates release by ovine CRF, an effect blocked by the GRP receptor antagonist d-Tyr⁶bombesin 6, 7, 8, 9, 10, 11, 12, 13propylamide. Bombesin did not potentiate AVP-stimulated ACTH release; instead release was attenuated when bombesin was given at a 10-fold or greater molar excess over AVP, with no interaction seen at lower concentrations. We conclude that ovine corticotrophs express bombesin receptors, and that GRP may act in concert with other hypothalamic releasing factors to regulate ACTH secretion.
Bombesin receptor antagonists
1996, Critical Reviews in Oncology/Hematology
Characterization and distribution of bombesin binding sites in the goldfish hypothalamic feeding center and pituitary
1995, Regulatory Peptides
Bombesin (BBS)/gastrin-releasing peptide (GRP) binding sites were characterized and their distribution examined in the goldfish brain and pituitary by radioligand binding and autoradiography. Binding of ¹²⁵I-[Tyr⁴]-BBS-14 to tissue sections was found to be saturable, reversible, time-dependent and displaceable by BBS/GRP-like peptides. Analysis of saturable equilibrium binding revealed a one-site model fit with a K_d of 0.665 ± 0.267 nM. This binding site displayed high affinity for members of the BBS subfamily of peptides, including GRP10 (K_i; 0.292 ± 0.038 nM) and GRP27 (K_i; 2.034 ± 1.597 nM), but showed no affinity for the BBS_8–14 fragment. While an approximate 100-fold lower binding affinity was displayed by the binding site for neuromedin B (K_i; 61.5 ± 28.2 nM), litorin was highly effective in displacing radiolabeled BBS binding (K_i; 1.469 ± 0.427 nM). The localization of saturable and high affinity BBS/GRP binding sites in specific areas of the goldfish brain and pituitary generally revealed a similar anatomical distribution to BBS/GRP-like immunoreactive material reported previously by our laboratory. Quantitative densitometric analysis of radiolabeled BBS binding to brain nuclei and the pituitary revealed a moderate concentration of BBS/GRP binding sites in the hypothalamic feeding area, including the nucleus diffusus lobi inferioris, nucleus recessus lateralis, nucleus lateral tuberis, and nucleus anterior tuberis. Other brain nuclei known to influence the brain feeding center which contained a high density of BBS/GRP binding sites included nuclei of the dorsal and ventro-medial telencephalon, the preoptic hypothalamus, and the optic tectum. High densities of BBS/GRP binding sites were also localized in the dorsal cerebellum, and nucleus habenularis. In the pituitary, BBS/GRP binding sites were present in high concentration in the neurointermediate lobe, with a relatively lower density localized in the pars distalis. The present study further supports a role for BBS/GRP-like peptides in the regulation of feeding behavior and anterior pituitary hormone secretion in teleosts.
Autoradiographic demonstration of <sup>125</sup>I-Tyr<sup>4</sup>-bombesin binding sites on rat anterior pituitary cells
1994, Peptides
Because several direct effects of bombesin related peptides on pituitary hormone release have been demonstrated, we chose to study the presence of bombesin binding sites in the adult male rat anterior pituitary in aggregate cell culture by autoradiographic localization of ¹²⁵I-tyrosine⁴-bombesin (¹²⁵I-Tyr⁴-BBN) binding and immunocytochemical localization of the anterior pituitary hormones. When aggregates were cultured in medium without hormonal supplements, the number of cells with detectable ¹²⁵I-Tyr⁴-BBN binding was below 1%. In cell aggregates cultured in the presence of 1 nM estradiol (E₂) ¹²⁵I-Tyr⁴-BBN binding was detected on $5.4 ± 0.8%$ of the cells after redispersion and on $5.8 ± 1.1%$ of the cells in sections of paraffin embedded aggregates. The binding cell types were mainly lactotrophs and somatotrophs. The binding of ¹²⁵I-Tyr⁴-BBN (3 or 5 nM) was specific because it was inhibited by the addition of an excess of unlabelled Tyr⁴-BBN or the bombesin receptor antagonist L686,095.
In aggregates cultured in the presence of 1 nM E₂ and 4 nM dex, the percentage of cells with detectable ¹²⁵I-Tyr⁴-BBN binding was significantly lower than that in aggregates cultured in the presence of 1 nM E₂ alone. Binding on somatotrophes almost completely disappeared.
The present data show that specific ¹²⁵I-Tyr⁴-BBN binding sites are present on anterior pituitary cells, are detectable mainly on a small subpopulation of lactotrophs and somatotrophs, and are affected by hormonal conditions. These data are consistent with the previously observed effects of bombesin-related peptides on growth hormone and prolactin release and the modulation of these effects by E₂ and dexamethasone and indicate that these effects are brought about by a direct action on lactotrophs and somatotrophs.

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^☆: This work was supported by grants from the Geconcerteerde Onderzoeksacties and the Belgian Fund for Scientific Research.

²: Research Fellow of the Belgian Fund for Scientific Research.

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Effect of the bombesin receptor blockers [Leu13,ψCH2NH-Leu14]bombesin and N-pivaloyl GRP(20–25) alkylamide (L 686,095-001C002) on basal and neuromedin C-stimulated PRL and GH release in pituitary cell aggregates☆

Abstract

J. Biol. Chem.

Biochem. Biophys. Res. Commun.

Peptides

Life Sci.

J. Biol. Chem.

Behavioral effects of bombesin

Ann. NY Acad. Sci.

Studies in vivo with Leu13-ψ-[CH2NH]-Leu14-bombesin, a proposed competitive antagonist of bombesin

FASEB J.

Potentiation by bombesin of corticotropin-releasing factor-stimulated ACTH release is dependent on the presence of glucocorticoids

Ann. NY Acad. Sci.

Design and evaluation of novel gastrin releasing peptide antagonists for the treatment of small cell lung cancer

Effect of the bombesin receptor blockers [Leu¹³,ψCH₂NH-Leu¹⁴]bombesin and N-pivaloyl GRP(20–25) alkylamide (L 686,095-001C002) on basal and neuromedin C-stimulated PRL and GH release in pituitary cell aggregates☆

Studies in vivo with Leu¹³-ψ-[CH₂NH]-Leu¹⁴-bombesin, a proposed competitive antagonist of bombesin