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Synthesis and binding characteristics of [H-3]neuromedin N, a NTS2 receptor ligand

Tóth, Fanni and Mallareddy, Jayapelreddy and Tourwé, Dirk and Lipkowski, Andrzej W. and Bujalska-Zadrozny, Magdalena and Benyhe, Sándor and Tóth, Géza (2016) Synthesis and binding characteristics of [H-3]neuromedin N, a NTS2 receptor ligand. NEUROPEPTIDES, 57. pp. 15-20. ISSN 0143-4179

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Abstract

Neurotensin (NT) and its analog neuromedin N (NN) are formed by the processing of a common precursor in mammalian brain tissue and intestines. The biological effects mediated by NT and NN (e.g. analgesia, hypothermia) result from the interaction with G protein-coupled receptors. The goal of this study consisted of the synthesis and radiolabeling of NN, as well as the determination of the binding characteristics of [H-3]NN and G protein activation by the cold ligand. In homologous displacement studies a weak affinity was determined for NN, with IC50 values of 454 nM in rat brain and 425 nM in rat spinal cord membranes. In saturation binding experiments the Kd value proved to be 264.8 +/- 30.18 nM, while the B-max value corresponded to 3.8 02 pmol/mg protein in rat brain membranes. The specific binding of [H-3]NN was saturable, interacting with a single set of homogenous binding sites. In sodium sensitivity experiments, a very weak inhibitory effect of Na+ ions was observed on the binding of resulting in an IC50 of 150.6 inM. In [S-35]GTP gamma S binding experiments the E-max value was 112.3 +/- 1.4% in rat brain and 112.9 +/- 2.4% in rat spinal cord membranes and EC50 values of 0.7 nM and 0.79 nM were determined, respectively. NN showed moderate agonist activities in stimulating G proteins. The stimulatory effect of NN could be maximally inhibited via use of the NTS2 receptor antagonist levocabastine, but not by the opioid receptor specific antagonist naloxone, nor by the NTS1 antagonist SR48692. These observations allow us to conclude that [H-3]NN labels NTS2 receptors in rat brain membranes. (C) 2015 Elsevier Ltd. All rights reserved.

Item Type: Article
Uncontrolled Keywords: SENSITIVITY; POTENTIATION; INACTIVATION; SITES; CLONING; ANTAGONIST; RAT-BRAIN; aspartate residue; NEUROMEDIN-N; NEUROTENSIN-LIKE PEPTIDE; RAT; NTS2; Receptor binding assays; Neuromedin N LEITH WALK, EDINBURGH EH1 3AF, MIDLOTHIAN, SCOTLAND
Subjects: Q Science / természettudomány > QD Chemistry / kémia > QD04 Organic chemistry / szerves kémia
Q Science / természettudomány > QH Natural history / természetrajz > QH301 Biology / biológia > QH3011 Biochemistry / biokémia
R Medicine / orvostudomány > RC Internal medicine / belgyógyászat > RC0321 Neuroscience. Biological psychiatry. Neuropsychiatry / idegkórtan, neurológia, pszichiátria
SWORD Depositor: MTMT SWORD
Depositing User: MTMT SWORD
Date Deposited: 01 Mar 2017 15:01
Last Modified: 01 Mar 2017 15:01
URI: http://real.mtak.hu/id/eprint/49914

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