Journal article
Characterization and Regulation of High Affinity Calcitonin Gene-Related Peptide Receptors in Cultured Neonatal Rat Cardiac Myocytes
Endocrinology (Philadelphia), Vol.128(6), pp.2731-2738
06/1991
DOI: 10.1210/endo-128-6-2731
PMID: 1645249
Abstract
Previous studies have shown that stimulation of cultured beating cardiac myocytes with calcitonin gene-related peptide (CGRP) produces increased beating frequency, increased cellular cAMP concentration, and a homologous desensitization of the cAMP-elevating action of CGRP. In the present study, the characteristics and regulation of [125I]CGRP binding sites in cultured cardiac myocytes were investigated. Binding of [125I] CGRP to membranes prepared from these cells was selective, saturable, and of high affinity. Scatchard transformation of the saturation isotherm generated a linear plot suggesting the existence of a homogeneous population of binding sites with an equilibrium binding constant of 41 +/- 7 pM and maximum binding capacity of 31 +/- 5 fmol/mg protein. Binding of [125I]CGRP to membranes was inhibited completely by guanosine 5'-(3-O-thio)triphosphate (250 microM), suggesting association of the binding sites with a G protein. Consistent with the saturation binding data, association kinetic studies indicated that [125I]CGRP associated with a single population of binding sites. Dissociation kinetic data, in contrast, indicated that [125I]CGRP dissociated from two affinity component sites on membranes, suggesting the existence of multiple affinity states of the G protein-coupled forms of the CGRP receptor. Nonequilibrium dissociation kinetic experiments revealed a time-dependent conversion of [125I] CGRP binding sites from a fast- to a slow-dissociating state. Desensitization of cells to CGRP by prior exposure to CGRP (10 nM) for 5 min reduced the maximal cAMP response of cells to further CGRP challenge and the number of [125I]CGRP binding sites in membranes prepared from these cells approximately 90% and 80%, respectively. These results demonstrate the existence of high affinity CGRP receptors in cardiac myocytes which appear coupled to G proteins and which undergo ligand-induced affinity alterations and desensitization-induced loss of receptor activity. The present findings also suggest the existence of multiple affinity states of the CGRP:receptor:G protein ternary complex.
Details
- Title: Subtitle
- Characterization and Regulation of High Affinity Calcitonin Gene-Related Peptide Receptors in Cultured Neonatal Rat Cardiac Myocytes
- Creators
- TAPAN K CHATTERJEEJADINE A MOYRORY A FISHER
- Resource Type
- Journal article
- Publication Details
- Endocrinology (Philadelphia), Vol.128(6), pp.2731-2738
- DOI
- 10.1210/endo-128-6-2731
- PMID
- 1645249
- ISSN
- 0013-7227
- eISSN
- 1945-7170
- Language
- English
- Date published
- 06/1991
- Academic Unit
- Iowa Neuroscience Institute; Neuroscience and Pharmacology; Internal Medicine
- Record Identifier
- 9984040452202771
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