Co-operative regulation of ligand binding to melanocortin receptor subtypes: Evidence for interacting binding sites

Kopanchuk, Sergei; Veikšina, Santa; Petrovska, Ramona; Mutule, Ilze; Szardenings, Michael; Rinken, Ago; Wikberg, Jarl E. S.

doi:10.1016/j.ejphar.2005.02.021

Cited by 32 publications

(31 citation statements)

References 37 publications

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“…That was also the case in our system, where 5-HT caused concentration dependent binding of [ 35 [26,27], but considerably lower than it could be expected from the radioligand binding (Table 2). Here it would be assumed that the discrepancies found between functional properties of the ligands and their abilities to compete with the radioligand binding can be caused by the kinetics of conformational isomerizations as it has been found for adenosine A 2A receptors [28] or by cooperative regulation of interacting binding sites as it has been found for melanocortin receptors [29] or by peculiarities of signal transduction system [30].…”

Section: Resultsmentioning

confidence: 97%

Characteristics of Binding of [3H]WAY100635 to Rat Hippocampal Membranes

Parkel

Rinken

2006

Neurochem Res

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Kinetic analysis of binding of [(3)H][N-[2-[4-(2-[O-methyl-(3)H]methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl)cyclohexane carboxamide ([(3)H]WAY100635) to 5-HT(1A) receptors in rat hippocampal membranes has revealed complex regulation mechanism for this radioligand. Saturation binding experiments revealed that [(3)H]WAY100635 binds to a single class of receptors with very high apparent affinity (K (D) = 87 +/- 4 pM, B (max) = 15.1 +/- 0.2 fmol/mg protein). The binding was almost irreversible, as the dissociation rate constant obtained k (off) = (7.8 +/- 1.1) x 10(-3) min(-1), means that equilibrium with this radioligand cannot be achieved before 7.5 h incubation at 25 degrees C. Systematic association kinetic studies of [(3)H]WAY100635 binding revealed sharp reaction acceleration at higher radioligand concentration, proposing mechanism of positive cooperativity. The affinities of antagonists determined from competition with [(3)H]WAY100635 did not coincide with their abilities to inhibit 5-HT-dependent activation of [(35)S]GTPgammaS binding probably due to the ligand's kinetic peculiarities. Thus, [(3)H]WAY100635 appears to be an excellent tool for determining receptor binding sites, but its applicability in equilibrium studies is strongly limited.

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Section: Resultsmentioning

confidence: 97%

Characteristics of Binding of [3H]WAY100635 to Rat Hippocampal Membranes

Parkel

Rinken

2006

Neurochem Res

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“…3,13 For endogenous peptide α-MSH, the affinities were in the sub-nanomolar range (pK i = 9.3-9.9), whereas synthetic agonist NDP-α-MSH had a slightly higher affinity (pK i = 9.9-10.1), and MS-05 had a lower affinity (pK i = 9.1). 3,13 The pEC 50 values obtained from functional assays were 8 to 8.6 for α-MSH and 7.9 for NDP-α-MSH. 14,15 Functional responses in these studies have been determined using [ and requires the use of radioactively labeled precursors.…”

Section: Resultsmentioning

confidence: 99%

“…It enabled making the receptor sample and assay environment "cleaner" for the binding studies, so acceptable data of MCR ligand-binding affinities became available. After revealing allosteric interactions between binding sites of MCR, 3 models of tandemly arranged ligandbinding sites 4 and ago-allosteric modulation 5 were proposed. Implementation of a fluorescence anisotropy/intensity assay with a dye-labeled NDP-α-MSH for MC 4 R allowed monitoring of ligand binding in real time 6 and revealed complexities in the binding processes, which have a big impact on the apparent affinities.…”

Section: Introductionmentioning

confidence: 99%

BacMam System for FRET-Based cAMP Sensor Expression in Studies of Melanocortin MC1 Receptor Activation

et al. 2012

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“…For the most part, the animal stroke models utilized in previous ␣ -MSH and NDP-␣ MSH studies were characterized by brief periods of ischemia, and when histologic assessments were conducted they focused on numbers of viable cells in very discrete brain regions. With particular regard to NDP-␣ MSH, studies of the binding kinetics of 125 I-NDP-␣ MSH to expressed human MCRs have demonstrated a significant fraction of ligand (20-50% of specific bound) remaining firmly bound and dissociating only very slowly or negligibly from MC1R, MC3R and MC4R subtypes [31] . This observation suggests the possibility that NDP-␣ MSH may exhibit enhanced efficacy in vivo by virtue of prolonged occupancy and activation of one or more MCR subtypes, as compared to other MCR ligands or small molecule probes with more traditional complete dissociation from receptor.…”

Section: Discussionmentioning

confidence: 99%

Lack of Protection with a Novel, Selective Melanocortin Receptor Subtype-4 Agonist RY767 in a Rat Transient Middle Cerebral Artery Occlusion Stroke Model

Regan

Shepherd²,

Strack³

et al. 2008

Pharmacology

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Previous studies utilizing α-melanocyte-stimulating hormone (α-MSH) or the synthetic analog [Nle⁴, D-Phe⁷] α-MSH have reported beneficial effects in animal models of ischemic stroke, with the latter studies suggesting melanocortin receptor subtype-4 (MC4R) activation as a protective mechanism. The present study directly addresses the hypothesis that MC4R activation may ameliorate ischemic brain injury by assessing the efficacy of a novel small molecule MC4R agonist RY767, administered in a pharmacokinetically guided and pharmacologically validated dosing regimen, in a rat stroke model of transient middle cerebral artery occlusion (tMCAO). Male Wistar rats were subjected to 90-min tMCAO followed by 72 h of reperfusion. Treatments were i.p. pretreatment with MK-801 (15 min prior to occlusion, positive control), or combined i.v. and p.o. daily administrations of vehicle, dextrose (negative control) or RY767 in blinded fashion initiated 2 h after occlusion. Infarct volume in MK-801-treated rats (158.7 ± 22.3 mm³) was reduced significantly compared to vehicle infarct volume (243.4 ± 12.5 mm³), whereas infarct volumes in dextrose- (224.3 ± 16.5 mm³) and RY767- (262.1 ± 19.2 mm³) treated rats did not differ from vehicle infarct volume. These results indicate that selective MC4R activation provides no significant neuroprotection, as reflected by infarct volume, in a rat stroke model utilizing a 90-min ischemic insult.

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Co-operative regulation of ligand binding to melanocortin receptor subtypes: Evidence for interacting binding sites

Cited by 32 publications

References 37 publications

Characteristics of Binding of [3H]WAY100635 to Rat Hippocampal Membranes

Characteristics of Binding of [3H]WAY100635 to Rat Hippocampal Membranes

BacMam System for FRET-Based cAMP Sensor Expression in Studies of Melanocortin MC1 Receptor Activation

Lack of Protection with a Novel, Selective Melanocortin Receptor Subtype-4 Agonist RY767 in a Rat Transient Middle Cerebral Artery Occlusion Stroke Model

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