Identification of NPB, NPW and Their Receptor in the Rat Heart

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Abstract

Members of neuropeptide B/W signaling system have been predominantly detected and mapped within the CNS. In the rat, this system includes neuropeptide B (NPB), neuropeptide W (NPW) and their specific receptor NPBWR1. This signaling system has a wide spectrum of functions including a role in modulation of inflammatory pain and neuroendocrine functions. Expression of NPB, NPW and NPBWR1 in separate heart compartments, dorsal root ganglia (DRG) and stellate ganglia was proven by RT-qPCR, Western blot (WB) and immunofluorescence. Presence of mRNA for all tested genes was detected within all heart compartments and ganglia. The presence of proteins preproNPB, preproNPW and NPBWR1 was confirmed in all the chambers of heart by WB. Expression of preproNPW and preproNPB was proven in cardiac ganglionic cells obtained by laser capture microdissection. In immunofluorescence analysis, NPB immunoreactivity was detected in nerve fibers, some nerve cell bodies and smooth muscle within heart and both ganglia. NPW immunoreactivity was present in the nerve cell bodies and nerve fibers of heart ganglia. Weak nonhomogenous staining of cardiomyocytes was present within heart ventricles. NPBWR1 immunoreactivity was detected on cardiomyocytes and some nerve fibers. We confirmed the presence of NPB/W signaling system in heart, DRG and stellate ganglia by proteomic and genomic analyses.

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Determination of cell types and numbers during cardiac development in the neonatal and adult rat and mouse.

Indroneal Banerjee, John Fuseler, Robert L Price … (2007)

Cardiac fibroblasts, myocytes, endothelial cells, and vascular smooth muscle cells are the major cellular constituents of the heart. The aim of this study was to observe alterations in myocardial cell populations during early neonatal development in the adult animal and to observe any variations of the cardiac cell populations in different species, specifically, the rat and mouse. Whole hearts were isolated from either mice or rats during the neonatal and adult stages of development, and single cell suspensions were prepared via sequential collagenase digestion. Heterogeneous cell populations were immunolabeled for specific cell types and analyzed using fluorescence-activated cell sorting (FACS). In addition, the left ventricle, right ventricle, and septa were isolated, fixed, and sectioned for morphometric analyses. These same cardiac regions were also analyzed using FACS. We observed that the adult murine myocardium is composed of approximately 56% myocytes, 27% fibroblasts, 7% endothelial cells, and 10% vascular smooth muscle cells. Moreover, our morphometric and FACS data demonstrated similar percentages in the three regions examined. During murine neonatal cardiac development, we observed a marked increase in numbers of cardiac fibroblasts and a resultant decrease in percentages of myocytes in late neonatal development (day 15). Finally, FACS analyses of the rat heart during development displayed similar results in relation to increases in cardiac fibroblasts during development; however, cell populations in the rat differed markedly from those observed in the mouse. Taken together, these data enabled us to establish a homeostatic model for the myocardium that can be compared with genetic and cardiac disease models.

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Identification of neuropeptide W as the endogenous ligand for orphan G-protein-coupled receptors GPR7 and GPR8.

M Harada, Masaaki Mori, Yoshio Matsumoto … (2002)

The structurally related orphan G-protein-coupled receptors GPR7 and GPR8 are expressed in the central nervous system, and their ligands have not been identified. Here, we report the identification of the endogenous ligand for both of these receptors. We purified the peptide ligand from porcine hypothalamus using stable Chinese hamster ovary cell lines expressing human GPR8 and cloned the cDNA encoding its precursor protein. The cDNA encodes two forms of the peptide ligand with lengths of 23 and 30 amino acid residues as mature peptides. We designated the two ligands neuropeptide W-23 (NPW23) and neuropeptide W-30 (NPW30). The amino acid sequence of NPW23 is completely identical to that of the N-terminal 23 residues of NPW30. Synthetic NPW23 and NPW30 activated and bound to both GPR7 and GPR8 at similar effective doses. Intracerebroventricular administration of NPW23 in rats increased food intake and stimulated prolactin release. These findings indicate that neuropeptide W is the endogenous ligand for both GPR7 and GPR8 and acts as a mediator of the central control of feeding and the neuroendocrine system.

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The cloning and chromosomal mapping of two novel human opioid-somatostatin-like receptor genes, GPR7 and GPR8, expressed in discrete areas of the brain.

B O'Dowd, X. Shi, A Marchese … (1995)

Following the cloning of the opioid receptors mu, kappa, and delta, we conducted a search for related receptors. Using oligonucleotides based on the opioid and also the structurally related somatostatin receptors, we amplified genomic DNA using the polymerase chain reaction and isolated fragments of novel G protein-coupled receptor genes. Two of these gene fragments designated clones 12 and 11 were used to isolate the full-length genes. The intronless coding sequences of these genes, named GPR7 and GPR8, shared 70% identity with each other, and each shared significant similarity with the sequences encoding transmembrane regions of the opioid and somatostatin receptors. GPR7 was mapped to chromosome 10q11.2-q21.1 and GPR8 to chromosome 20q13.3. Northern blot analysis using human mRNA demonstrated expression of GPR7 mainly in cerebellum and frontal cortex, while GPR8 was located mainly in the frontal cortex. In situ hybridization revealed expression of GPR7 in the human pituitary. A partial sequence of the mouse orthologue of GPR7 was obtained, and in situ hybridization demonstrated expression in discrete nuclei of brain, namely suprachiasmatic, arcuate, and ventromedial nuclei of hypothalamus. A stable cell line expressing the GPR7 gene was created, but expression levels of the receptor were low. The available pharmacology indicated binding to several opioid drugs such as bremazocine, levorphanol, and beta-FNA, but not to the opioid receptor subtype-selective mu, delta, or kappa agonists.

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Author and article information

Journal

Journal ID (nlm-ta): Int J Mol Sci

Journal ID (iso-abbrev): Int J Mol Sci

Journal ID (publisher-id): ijms

Title: International Journal of Molecular Sciences

Publisher: MDPI

ISSN (Electronic): 1422-0067

Publication date (Electronic): 22 October 2020

Publication date Collection: November 2020

Volume: 21

Issue: 21

Electronic Location Identifier: 7827

Affiliations

[1 ]Department of Pharmacology and Toxicology, Faculty of Medicine in Pilsen, Charles University, 323 00 Pilsen, Czech Republic; shashank.pandey@ 123456lfp.cuni.cz

[2 ]Biomedical Center, Faculty of Medicine in Pilsen, Charles University, 323 00 Pilsen, Czech Republic; zdenek.tuma@ 123456lfp.cuni.cz

[3 ]PeptLab@UCP Platform and CNRS BioCIS, CY Cergy Paris Université, 950 31 Cergy-Pontoise CEDEX, France; elisa.peroni@ 123456u-cergy.fr (E.P.); olivier.monasson@ 123456u-cergy.fr (O.M.); annamaria.papini@ 123456unifi.it (A.M.P.)

[4 ]Interdepartmental Research Unit of Peptide and Protein Chemistry and Biology, Department of Chemistry “Ugo Schiff”, University of Florence, 500 19 Sesto Fiorentino, Italy

[5 ]Department of Physiology, Faculty of Medicine in Pilsen, Charles University, 323 00 Pilsen, Czech Republic

Author notes

[* ]Correspondence: Magdalena.dvorakova@ 123456lfp.cuni.cz ; Tel.: +420-377-593-343

Author information

Zdenek Tuma https://orcid.org/0000-0002-2375-4169

Olivier Monasson https://orcid.org/0000-0002-0886-1687

Anna Maria Papini https://orcid.org/0000-0002-2947-7107

Magdalena Chottova Dvorakova https://orcid.org/0000-0002-6078-1959

Article

Publisher ID: ijms-21-07827

DOI: 10.3390/ijms21217827

PMC ID: 7659951

PubMed ID: 33105700

SO-VID: f83b2ac6-361e-4d90-8ab2-3dc628bb6315

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Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

Identification of NPB, NPW and Their Receptor in the Rat Heart

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Abstract

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Higher order chromatin architecture

Most cited references 24

Determination of cell types and numbers during cardiac development in the neonatal and adult rat and mouse.

Identification of neuropeptide W as the endogenous ligand for orphan G-protein-coupled receptors GPR7 and GPR8.

The cloning and chromosomal mapping of two novel human opioid-somatostatin-like receptor genes, GPR7 and GPR8, expressed in discrete areas of the brain.

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