Desensitisation of adrenomedullin and CGRP receptors Debbie L. Hay a, * , David R. Poyner b , David M. Smith a a Department of Metabolic Medicine, Imperial College School of Medicine, Hammersmith Hospital, Du Cane Road, London W12 0NN, UK b School of Life Sciences, Aston University, Aston Triangle, Birmingham B4 7ET, UK Received 26 April 2002; accepted 24 October 2002 Abstract Adrenomedullin (AM), a potent vasoactive peptide, is elevated in certain disease states such as sepsis. Its role as a physiologically relevant peptide has been confirmed with the advent of the homozygous lethal AM peptide knockout mouse. So far, there have been few and conflicting studies which examine the regulatory role of AM at the receptor level. In this article, we discuss the few studies that have been presented on the desensitisation of AM receptors and also present novel data on the desensitisation of endogenous AM receptors in Rat-2 fibroblasts. D 2003 Elsevier Science B.V. All rights reserved. Keywords: Adrenomedullin; CGRP; RAMP; CRLR; Rat-2 fibroblast; Desensitisation 1. Introduction Adrenomedullin (AM) and calcitonin gene-related pep- tide (CGRP) are related members of the calcitonin family of peptides, sharing many biological properties [1]. Probably their most characterised action is that of potent vasodilation [1,2]. The similarities of these peptides stretch beyond their biological actions in that they both activate the same cell surface family B G-protein-coupled receptor (GPCR), the calcitonin receptor-like receptor (CRLR) [3]. Specificity for these peptides at this receptor is conferred by a family of single transmembrane proteins termed receptor activity- modifying proteins (RAMPs). There are three known RAMPs, with low amino acid homology but stronger struc- tural homology. CRLR is nonfunctional in the absence of RAMP proteins and is differentially responsive to AM and CGRP according to the RAMP with which it is associated. CRLR, coexpressed in a cell line with RAMP1, yields a receptor preferentially responsive to CGRP. CRLR coex- pressed with either RAMP2 or RAMP3 generates AM- preferring receptors [3]. To date, there appears to be little difference between the AM receptors generated by RAMP2 or RAMP3. In all cases, CRLR signalling is predominantly via the G s , adenylate cyclase-linked pathway [3–5] although AM and CGRP receptors in a variety of cell lines have additionally been shown to signal via different pathways (reviewed in Refs. [1,6]). CRLR, when transfected into HEK293 cells with any RAMP, was able to elevate intra- cellular calcium levels [7,8]. GPCRs constitute the largest family of cell surface receptors and mediate physiological responses to a wide array of stimuli. These receptors are tightly regulated and undergo the phenomenon of desensitisation to prevent the detrimental effects of overstimulation. This type of receptor regulation is likely to be extremely important in conditions where agonist (e.g., AM) levels are elevated, leading to excessive receptor stimulation, and where other hormones may indirectly affect the receptor in question. It is also important to dissect these mechanisms to fully understand the impact of drug treatment on receptors. Desensitisation is classically divided into homologous or heterologous mechanisms. Homologous desensitisation is the result of agonist-dependent activation of the receptor and involves a family of serine/threonine kinases termed GPCR kinases (GRKs). Alternatively, heterologous desensitisation involves second messenger-dependent kinases, such as the cAMP-dependent protein kinase (PKA) or protein kinase C (PKC), and may be the result of activation of a different receptor. Following phosphorylation events, receptor endo- cytosis depletes the plasma membrane of phosphorylated receptors and receptor downregulation occurs over a longer 0167-0115/03/$ - see front matter D 2003 Elsevier Science B.V. All rights reserved. doi:10.1016/S0167-0115(03)00032-6 * Corresponding author. Current address: The Liggins Institute, University of Auckland, Auckland, New Zealand. Tel.: +64-9-373-7599; fax: +64-9-373-7497. E-mail address: dl.hay@auckland.ac.nz (D.L. Hay). www.elsevier.com/locate/regpep Regulatory Peptides 112 (2003) 139 – 145