Elsevier

Peptides

Volume 25, Issue 3, March 2004, Pages 365-391
Peptides

Review
Receptor autoradiography as mean to explore the possible functional relevance of neuropeptides: focus on new agonists and antagonists to study natriuretic peptides, neuropeptide Y and calcitonin gene-related peptides

https://doi.org/10.1016/j.peptides.2004.01.013Get rights and content

Abstract

Over the past 20 years, receptor autoradiography has proven most useful to provide clues as to the role of various families of peptides expressed in the brain. Early on, we used this method to investigate the possible roles of various brain peptides. Natriuretic peptide (NP), neuropeptide Y (NPY) and calcitonin (CT) peptide families are widely distributed in the peripheral and central nervous system and induced multiple biological effects by activating plasma membrane receptor proteins. The NP family includes atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP) and C-type natriuretic peptide (CNP). The NPY family is composed of at least three peptides NPY, peptide YY (PYY) and the pancreatic polypeptides (PPs). The CT family includes CT, calcitonin gene-related peptide (CGRP), amylin (AMY), adrenomedullin (AM) and two newly isolated peptides, intermedin and calcitonin receptor-stimulating peptide (CRSP). Using quantitative receptor autoradiography as well as selective agonists and antagonists for each peptide family, in vivo and in vitro assays revealed complex pharmacological responses and radioligand binding profile. The existence of heterogeneous populations of NP, NPY and CT/CGRP receptors has been confirmed by cloning. Three NP receptors have been cloned. One is a single-transmembrane clearance receptor (NPR-C) while the other two known as CG-A (or NPR-A) and CG-B (or NPR-B) are coupled to guanylate cyclase. Five NPY receptors have been cloned designated as Y1, Y2, Y4, Y5 and y6. All NPY receptors belong to the seven-transmembrane G-protein coupled receptors family (GPCRs; subfamily type I). CGRP, AMY and AM receptors are complexes which include a GPCR (the CT receptor or CTR and calcitonin receptor-like receptor or CRLR) and a single-transmembrane domain protein known as receptor-activity-modifying-proteins (RAMPs) as well as an intracellular protein named receptor-component-protein (RCP). We review here tools that are currently available in order to target each NP, NPY and CT/CGRP receptor subtype and establish their respective pathophysiological relevance.

Introduction

Early on, receptor autoradiography proven most useful as a mean to explore the possible pathophysiological relevance of various family of peptides in the central and peripheral nervous systems (for review [46]). We were first to report on the discrete distribution of neuropeptide Y (NPY) receptor binding sites in the rat brain in Peptides in 1986 [250]. It was also in Peptides that we were first to demonstrate the discrete distribution of specific atria natriuretic peptide (ANP) binding sites in the rat brain [308]. These data were highly suggestive of the existence of brain peptides related to the ANP family. This hypothesis was subsequently confirmed with the isolation of brain natriuretic peptides (BNPs) [360]. Finally, data on the discrete localization of calcitonin gene-related peptide (CGRP) receptors in the brain were first reported in PNAS [377] and demonstrated that CGRP receptors were distinct from calcitonin (CT) receptors. This review will summarize recent findings about these three families of peptides.

The natriuretic peptide family is composed of three peptides namely ANP, BNP and C-type natriuretic peptides (CNPs) [330]. ANP was the first member of this peptide family to be isolated from the atria [77]. Two other natriuretic peptides, BNP and CNP, were isolated from porcine brain [360], [362]. Although BNP was first isolated from porcine brain, it was later found to be predominantly produced by the heart and vascular endothelial cells [285]. These peptides share a common core structure (a 17 amino acids disulfide ring) which is essential for their biological actions. Most research on these peptides has focused on their peripheral effects in the heart, blood vessels and kidney while much less is currently known about their role in the central nervous system [223], [330]. ANP, BNP and CNP bind to two single-transmembrane proteins which belong to a family of guanylyl cyclase (GC)-coupled receptors termed GC-A and GC-B [223], [266], [374]. They are also known as NPR-A or NPR-1 and NPR-B receptors [330]. Additionally, a third receptor designated as NPR-C is a clearance or silent receptor [125], [244]. The pharmacological and molecular profiles of each receptor subtype are summarized in Table 1.

The NPY family includes NPY and two other peptides known as peptide YY (PYY) and the pancreatic polypeptides (PPs) [367]. NPY is one of the most abundant peptide expressed in the mammalian brain [49], [50], [78], [79], while PYY and PPs are mostly found in endocrine cells of the intestinal tract [353]. It is now well established that NPY induced its effects via the activation of at least five receptor subtypes termed Y1, Y2, Y4, Y5 and y6 [261]. All these receptors have been cloned and belong to the type I GPCR receptor family [101]. Each NPY receptor subtype has distinctive pharmacological profile as defined using various agonists and antagonists. Most agonists can activate more than one receptor subtype while antagonists generally display higher selectivity (Table 2).

The calcitonin family includes CT, amylin (AMY), two CGRPs termed CGRPα and CGRPβ, adrenomedullin (AM) [301] and two most recently isolated peptides called calcitonin receptor-stimulating peptide (CRSP) [206] and intermedin [321]. Key features of this peptide family is the presence of a disulfide bridge in the N-terminus and the deletion of amino acid residues in this region generates C-terminal fragments [CGRP(8–37) and others] with antagonistic properties [81], [83]. The biological effects induced by this peptide family are mediated via binding to two closely related type II GPCRs, the calcitonin receptor (CTR) and calcitonin receptor-like receptor (CRLR). Moreover, and in contrast to most GPCRs, CTR and CRLR require the presence of chaperone proteins such as the receptor-activity-modifying-proteins (RAMPs) [254] and receptor-component-protein (RCP) [233] in order to provide functional responses to CGRP, AM and AMY. Furthermore, depending of the associated RAMPs distinctive pharmacological profiles can be generated [301]. These profiles are summarized in Table 3.

Section snippets

Natriuretic peptide family

Natriuretic peptides (NPs) are members of a family of vasoactive hormones that play important roles in the regulation of blood pressure, cardiovascular homeostasis and kidney functions [223], [330], [374]. ANP is a cardiac peptide derived from a 151 (rat) and 152 (human) amino acid residue preprohormone which by proteolytic cleavage (Ser99–Tyr126) results in an active peptide of 28 amino acids [ANP(1–28)] [77]. BNP is a 32 amino acid peptide that although first isolated from porcine brain has

The NPY family

NPY is a 36 amino acid residues polypeptide, first isolated from porcine brain [366]. This peptide shares not only high sequence homologies with PYY and the PPs but also structural elements which confer a hairpin-like structure to these molecules [4], [245]. NPY is one of most evolutionary conserved peptides [225], suggesting significant roles in basic physiological functions. In fact, this peptide family has demonstrated a broad range of biological effects including increased food and water

Calcitonin and calcitonin gene-related peptide family

The first peptide of this family was isolated from the thyroid gland and called CT. This peptide is implicated in calcium homeostasis and bone resorption [68]. Many years later, molecular cloning of DNA complementary to rat CT mRNA revealed that the CT gene could generate a second distinct transcript coding for a 37 amino acid polypeptide named CGRP [6], [324]. The possible existence of other gene products related to CGRP was investigated by screening library and resulted in the identification

General conclusion

The development of highly selective agonists and antagonists is critical in order to clearly establish the functional significance of a given receptor subtype. Without such tools it is very difficult to obtain definitive answers. Additionally, the development of radiolabeled molecules highly selective for a receptor subtype is important to precisely access discrete receptor distribution and to characterize receptor binding properties. Finally, receptor localization has clearly proven to be most

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    DOI of original article: 10.1016/0196-9781(84)90183-9.

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