Calcitonin gene-related peptide effect on adenylate cyclase activity in the pregnant rat

From this study, we predicted that the human calcitonin gene-related peptide (hCGRP) fragment hCGRP-(8-37) would be a selective antagonist for CGRP receptors but an agonist for calcitonin (CT) receptors. In rat liver plasma membrane, where CGRP receptors predominate and CT appears to act through these receptors, hCGRP-(8-37) dose dependently displaced 125I-[Tyr0]rat CGRP binding. However, hCGRP-(8-37) had no effect on adenylate cyclase activity in liver plasma membrane. Furthermore, hCGRP-(8-37) inhibited adenylate cyclase activation induced not only by hCGRP but also by hCT. On the other hand, in LLC-PK1 cells, where calcitonin receptors are abundant and CGRP appears to act via these receptors, the bindings of 125I-[Tyr0]rat CGRP and 125I-hCT were both inhibited by hCGRP-(8-37). In contrast to liver membranes, interaction of hCGRP-(8-37) with these receptors led to stimulation of adenosine 3',5'-cyclic monophosphate (cAMP) production in LLC-PK1 cells, and moreover, this fragment did not inhibit the increased production of cAMP induced not only by hCT but also by hCGRP. Thus hCGRP-(8-37) appears to be a useful tool for determining whether the action of CGRP as well as that of CT is mediated via specific CGRP receptors or CT receptors.

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Effect of denervation and reserpine treatment on adenylate cyclase stimulated by calcitonin gene-related peptide and isoproterenol in rat diaphragm

The Japanese Journal of Pharmacology ◽

10.1016/s0021-5198(19)55939-x ◽

1990 ◽

Vol 52 ◽

pp. 372

Author(s):

Naoko Matsumoto ◽

Wang Xiao-Bing ◽

Shuji Uchida

Keyword(s):

Adenylate Cyclase ◽

Calcitonin Gene Related Peptide ◽

Rat Diaphragm ◽

Related Peptide ◽

Reserpine Treatment ◽

Calcitonin Gene

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A rat skeletal muscle cell line (L6) expresses specific adrenomedullin binding sites but activates adenylate cyclase via calcitonin gene-related peptide receptors

Biochemical Journal ◽

10.1042/bj3180241 ◽

1996 ◽

Vol 318 (1) ◽

pp. 241-245 ◽

Cited By ~ 36

Author(s):

Hedley A COPPOCK ◽

Ali A OWJI ◽

Stephen R BLOOM ◽

David M SMITH

Keyword(s):

Protein Kinase ◽

Adenylate Cyclase ◽

Binding Site ◽

Binding Sites ◽

Calcitonin Gene Related Peptide ◽

Dependent Protein Kinase ◽

Related Peptide ◽

Camp Dependent Protein Kinase ◽

Calcitonin Gene ◽

Dependent Protein

We have previously demonstrated specific binding sites for adrenomedullin, a novel member of the calcitonin family of peptides, in rat muscles. It is unclear whether these receptors are vascular or muscular. Receptors for the structurally similar calcitonin gene-related peptide (CGRP) are present on myocytes and might be involved in the regulation of myocyte glucose metabolism and control by motor neurons. We investigated whether adrenomedullin binding sites were present on L6 myocytes. Specific [125I]adrenomedullin binding sites were demonstrated where adrenomedullin competed with an IC50 of 0.22±0.04 nM (mean±S.E.M.) and a concentration of binding sites (Bmax) of 0.95±0.19 pmol/mg of protein (mean±S.E.M.). CGRP and the specific CGRP receptor antagonist CGRP(8–37) competed weakly at this site (IC50 > 10 and 601±298 nM respectively). Binding studies with [125I]CGRP revealed a binding site for CGRP (IC50 = 0.13±0.01 nM; Bmax = 0.83±0.10 pmol/mg of protein) where both CGRP(8–37) and adrenomedullin competed with [125I]CGRP with IC50 values of 1.15±0.12 and 8.68±0.98 nM respectively. Chemical cross-linking showed the CGRP and adrenomedullin binding site–ligand complexes to have approximate molecular masses of 82 and 76 kDa respectively. Both CGRP and adrenomedullin increased adenylate cyclase activity with similar potencies. In both cases adenylate cyclase activation was blocked by CGRP(8–37). Stimulation with 10 nM adrenomedullin or CGRP caused an increase in the percentage of total activated cellular cAMP-dependent protein kinase from 38% in resting cells to 100% and 98% respectively. Therefore in L6 cells adrenomedullin can bind to CGRP receptors, activating adenylate cyclase and cAMP-dependent protein kinase.

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Characterization of specific calcitonin gene related peptide receptors present in hamster pancreatic β cells

Bioscience Reports ◽

10.1007/bf01123504 ◽

1993 ◽

Vol 13 (4) ◽

pp. 221-231 ◽

Cited By ~ 9

Author(s):

A. Barakat ◽

G. Rosselin ◽

J.-C. Marie

Keyword(s):

Adenylate Cyclase ◽

Cell Membranes ◽

Specific Binding ◽

Calcitonin Gene Related Peptide ◽

Receptor Subtypes ◽

Cgrp Receptor ◽

Β Cells ◽

Related Peptide ◽

Insulinoma Cell ◽

Calcitonin Gene

Calcitonin gene-related peptide (CGRP) shares about 46% and 20% amino acid sequence homology with islet amyloid polypeptide (IAPP) and salmon calcitonin (sCT). We investigated whether these related peptides could cross-react with the specific binding of 125I-[His]hCGRP I to the CGRP receptor in hamster insulinoma cell membranes. A rapid dissociation of membrane bound 125I-[His]hCGRP I could be induced in the presence of 1 μM chicken CGRP (cCGRP). The specific 125I-[His]hCGRP I binding was inhibited by the related peptides and their half-maximal inhibitory concentrations (IC50) were: cCGRP (0.1 nM), rat CGRP I and human CGRP I and II (1.0–2.0 nM), fragment of hCGRP I (8–37) (150 nM), human IAPP (440 nM). The non-amidated form of hIAPP; human diabetes-associated peptide (hDAP) did not inhibit the binding of 125I-[His]hCGRP I and sCT was only effective at a high concentration (1 μM). Binding of 125I-[His]hCGRP I was dose dependently inhibited by guanosine-5′-O-(3-thiotriphosphate) or (GTPγS) and a 70% reduction of binding was obtained with 0.1 mM GTPγS. The IC50 value of cCGRP (0.1 nM) was increased 100-fold in the presence of 0.1 mM GTPγS. Human CGRP I and cCGRP at 2.5 μM did not stimulate the activity of hamster insulinoma cell membranes adenylate cyclase, while glucagon (1 μM) induced a 2-fold increase. Thus, specific CGRP receptors present in hamster β cells are associated with G protein (s) and IAPP can interact with these receptors. These results and the observation that cCGRP and hCGRP I did not influence adenylate cyclase activity provide further evidence for CGRP receptor subtypes.

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