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RECEPTORS AND SIGNAL TRANSDUCTION

Immature osteoblastic MG63 cells possess two calcitonin gene-related peptide receptor subtypes that respond differently to [Cys(Acm)^2,7] calcitonin gene-related peptide and CGRP_8–37

¹Division of Cellular Pharmacology, Department of Signal Transduction Research, and ²Division of Periodontology, Department of Oral Biological Science, Graduate School of Medical and Dental Sciences, Niigata University, Niigata, Japan; and ³Medical Research Service, Kansas City Department of Veterans Affairs Medical Center, and Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Missouri

Submitted 14 October 2004 ; accepted in final form 4 May 2005

Calcitonin gene-related peptide (CGRP) is clearly an anabolic factor in skeletal tissue, but the distribution of CGRP receptor (CGRPR) subtypes in osteoblastic cells is poorly understood. We previously demonstrated that the CGRPR expressed in osteoblastic MG63 cells does not match exactly the known characteristics of the classic subtype 1 receptor (CGRPR₁). The aim of the present study was to further characterize the MG63 CGRPR using a selective agonist of the putative CGRPR₂, [Cys(Acm)^2,7]CGRP, and a relatively specific antagonist of CGRPR₁, CGRP_8–37. [Cys(Acm)^2,7]CGRP acted as a significant agonist only upon ERK dephosphorylation, whereas this analog effectively antagonized CGRP-induced cAMP production and phosphorylation of cAMP response element-binding protein (CREB) and p38 MAPK. Although it had no agonistic action when used alone, CGRP_8–37 potently blocked CGRP actions on cAMP, CREB, and p38 MAPK but had less of an effect on ERK. Schild plot analysis of the latter data revealed that the apparent pA₂ value for ERK is clearly distinguishable from those of the other three plots as judged using the 95% confidence intervals. Additional assays using 3-isobutyl-1-methylxanthine or the PKA inhibitor N-(2-[p-bromocinnamylamino]ethyl)-5-isoquinolinesulfonamide hydrochloride (H-89) indicated that the cAMP-dependent pathway was predominantly responsible for CREB phosphorylation, partially involved in ERK dephosphorylation, and not involved in p38 MAPK phosphorylation. Considering previous data from Scatchard analysis of [¹²⁵I]CGRP binding in connection with these results, these findings suggest that MG63 cells possess two functionally distinct CGRPR subtypes that show almost identical affinity for CGRP but different sensitivity to CGRP analogs: one is best characterized as a variation of CGRPR₁, and the second may be a novel variant of CGRPR₂.

cAMP; MAP kinase; preosteoblasts; Schild plot

This article has been cited by other articles:

				D. L. Hay, D. R. Poyner, and R. Quirion International Union of Pharmacology. LXIX. Status of the Calcitonin Gene-Related Peptide Subtype 2 Receptor Pharmacol. Rev., June 1, 2008; 60(2): 143 - 145. [Abstract] [Full Text] [PDF]