Functional Calcitonin Gene-Related Peptide Subtype 2 Receptors in Porcine Coronary Arteries Are Identified as Calcitonin Gene-Related Peptide Subtype 1 Receptors by Radioligand Binding and Reverse Transcription-Polymerase Chain Reaction

摘要

Calcitonin gene-related peptide (CGRP) receptors are classified into CGRP subtype 1 9CGRP_1) and CGRP subtype 2 (CGRP_2) based on the affinity of the antagonist, human #alpha# (h#alpha#)-CGRP_(8-37). h#alpha#-CGRP_(8-37) antagonizes CGRP_1 receptor-mediated responses with high affinity (K_B < 100 nM) and antagonizes CRGP_2 receoptor-mediated resoponses with low affinity (K_B > 1 #mu#M). CGRP-2 receptors have been previously reported to mediate relaxation of large oporcine coronary arteries because this action is antagonized with low affinity by h#alpha#-CGRP_(8-37). In the present study, we used reverse transcription-polymerase chain reaction, isolated tissue experiments to compare the CGRP receptor in porcine coronary arteries with the procine CGRP_1 receptor stably expressed in human embryonic kidney (HEK) 293 cells. We identified calcitonin receptor-like receptor and receptor activity modifying protein 1 mRNA in coronary arteries. We also found that the ligand binding characteristics of the CGRP receptor in coronary arteries and the cloned CGRP_1 receptor were highly similar. K_I values for h#alpha#-CGRP_(8-37) were 6.6 and 5.7 nM in porcine coronary arteries and the cloned CGRP_1 receptor, respectively. The affinities (K_B) of h#alpha#-CGRP_(8-37) and five other antagonists were 22-to 707-fold lower in functional experiments measuring relaxation of coronary arteries than in radioligand binding experiments. Despite this difference in absolute affinity values, there was a high correlated of the rank order of affinity for the antagonists determined by the two methods. Thus h#alpha#-CGRP_(8-37) antagonizes CGRP-induced relaxation of porcine coronary arteries with low affinity at the CGROP, receptor. Taken together, these data do not support the existence of the CGRP_2 receptor.