Differential expression of components of the CGRP-receptor family in human coronary and human middle meningeal arteries: functional implications

Abstract

Background: Different responses in human coronary arteries (HCA) and human middle meningeal arteries (HMMA) were observed for some of the novel CGRP receptor antagonists, the gepants, for inhibiting CGRP-induced relaxation. These differences could be explained by the presence of different receptor populations in the two vascular beds. Here, we aim to elucidate which receptors are involved in the relaxation to calcitonin gene-related peptide (CGRP), adrenomedullin (AM) and adrenomedullin 2 (AM2) in HCA and HMMA.

Methods: RNA was isolated from homogenized human arteries (23 HCAs; 12 F, 11 M, age 50 ± 3 years and 26 HMMAs; 14 F, 12 M, age 51 ± 3 years) and qPCR was performed for different receptor subunits. Additionally, relaxation responses to CGRP, AM or AM2 of the human arteries were quantified using a Mulvany myograph system, in the presence or absence of the adrenomedullin 1 receptor antagonist AM_22-52 and/or olcegepant.

Results: Calcitonin-like receptor (CLR) mRNA was expressed equally in both vascular beds, while calcitonin receptor (CTR) and receptor activity-modifying protein 3 (RAMP3) expression was low and could not be detected in all samples. RAMP1 expression was similar in HCA and HMMA, while RAMP2 expression was higher in HMMA. Moreover, receptor component protein (RCP) expression was higher in HMMA than in HCA. Functional experiments showed that olcegepant inhibits relaxation to all three agonists in both vascular beds. In HCA, antagonist AM_22-52 did not inhibit relaxation to any of the agonists, while a trend for blocking relaxation to AM and AM2 could be observed in HMMA.

Conclusion: Based on the combined results from receptor subunit mRNA expression and the functional responses in both vascular tissues, relaxation of HCA is mainly mediated via the canonical CGRP receptor (CLR-RAMP1), while relaxation of HMMA can be mediated via both the canonical CGRP receptor and the adrenomedullin 1 receptor (CLR-RAMP2). Future research should investigate whether RAMP2 predominance over RAMP1 in the meningeal vasculature results in altered migraine susceptibility or in a different response to anti-migraine medication in these patients. Moreover, the exact role of RCP in CGRP receptor signalling should be elucidated in future research.

Keywords: AM22-52; Adrenomedullin; CGRP; Human arteries; Olcegepant; Receptor expression; Vasodilation; qPCR.

Fig. 1

mRNA expression of receptor subunits in the human coronary artery and human middle meningeal artery. A) Overview of the different receptors within the calcitonin gene-related peptide family and the transmembrane proteins that constitute the receptor. B) Expression of CLR, CTR, RAMP1, RAMP2, RAMP3 and RCP in human coronary artery (n = 23) and human middle meningeal artery (n = 26). Data are expressed as mean ± SD. *p < 0.05, **p < 0.01

Fig. 2

Concentration-response curve to CGRP, adrenomedullin or adrenomedullin 2 in the human coronary artery. Experiments were performed in the presence or absence of CGRP receptor antagonist olcegepant (1 µM) and adrenomedullin receptor antagonist AM_{22 − 52} (1 µM). (A) Relaxation to CGRP (n = 8). (B) Relaxation to adrenomedullin (n = 6). (C) Relaxation to adrenomedullin 2 (n = 6)

Fig. 3

Concentration-response curve to CGRP, adrenomedullin or adrenomedullin 2 in the human middle meningeal artery. Experiments were performed in the presence or absence of CGRP receptor antagonist olcegepant (1 µM) and adrenomedullin receptor antagonist AM_22-52 (1 µM). (A) Relaxation to CGRP (n = 6–11). (B) Relaxation to adrenomedullin (n = 6–9). (C) Relaxation to adrenomedullin 2 (n = 8–10)