P1: KWW/KKL P2: KWW/HCN QC: KWW/FLX T1: KWW GRBT050-16 Olesen- 2057G GRBT050-Olesen-v6.cls July 9, 2005 4:30 ••Chapter 16 ◗ Calcitonin Gene-Related Peptide and Other Peptides Susan Brain and Lars Edvinsson Vasoactive peptides can be either stored or synthesized de THE CGRP FAMILY OF PEPTIDES novo before release from a range of tissues in the brain or from the walls of intracranial vasculature. In this chapter, The expression of mRNA from the calcitonin gene is tissue we concentrate on neuropeptides that are released from specific in that CGRP mRNA is predominantly expressed perivascular nerves. These include calcitonin gene-related in nerves and calcitonin mRNA in the thyroid (5). The 37 peptide (CGRP), substance P, neurokinin A, nociceptin, amino acid peptide CGRP belongs to a family that include somatostatin, and opioids (Table 16-1). The endothelium the more recently discovered peptides adrenomedullin produces the potent vasoconstrictors endothelin and an- that is primarily produced by non-neuronal tissues, espe- giotensin, and dilators such as nitric oxide, prostacyclin, cially vascular tissues and amylin that is mainly produced and endothelium-derived hyperpolarizing factors. In ad- in the pancreas. They share some structural homology (ap- dition there are circulating agents; among these the most proximately 25–40%) and also some, but not total, similar- potent is 5-hydroxytryptamine. The neuronal messengers ities in biological activities (see Brain and Grant [11] for stored in the intracranial vessels have been reviewed recent review). CGRP is abundant in the body and has a (32) and it was revealed that sympathetic nerves store wide distribution throughout the central and peripheral noradrenaline, neuropeptide Y, and ATP, the parasympa- nervous systems. It has a number of biological activities, thetic fibers nitric oxide, acetylcholine, vasoactive intesti- but the most relevant to migraine are its activities within nal peptide (VIP), peptide histidine isoleucine, pituitary the nervous and cardiovascular systems. CGRP is an ex- adenylate cyclase-activating peptide (PACAP), and helo- tremely potent and long-lasting vasodilator, that is active dermine. Although VIP is of interest in certain forms of at all levels of the cardiovascular system with good evi- primary headaches, it is mainly the peptides contained in dence for exquisite activity in the cerebral circulation (see the sensory system, and this has attracted the most ex- Edvinsson [23]). citement during the last few years. A growing body of CGRP is most usually found in unmyelinated sensory studies indicate that the distribution and biological ac- C-fibers and myelinated Aδ-fibers that are commonly as- tivity of these neuropeptides to the areas of the cerebral sociated with blood vessels, where perivascular nerves vasculature and nervous system that are involved in mi- terminate in close association with the vessels. Specific la- graine is highly relevant. Research into the precise role beling of the C- and Aδ-fibers with wheat germ agglutinin– and importance of these peptides is often lacking because conjugated horse radish peroxidase and cholera toxin sub- of the paucity of selective agents that either enhance or unit b revealed that the C-fibers are located in lamina 1 and block activity or synthesis. However, there have been some 2, and the Aδ-fibers from mechanoreceptors lay located in major clinical trials that have focused on these peptides lamina 3 and 4 (48). Labeling experiments have revealed in recent years that have, for instance, ruled out a role that the temporal artery and the superior sagittal sinus for substance P, but provided evidence for the involve- differ in their somatotopic organization. CGRP is the most ment of CGRP. This chapter is written in a comparative prevalent of the neuropeptides in the sensory fibers. CGRP manner to allow an appreciation of our relative knowl- is commonly colocalized with other peptides in C-fibers, edge of the biology of these peptides and their role in which include substance P (49). There are two forms of migraine. CGRP and αCGRP (or CGRPI), encoded by the calcitonin 159 P1: KWW/KKL P2: KWW/HCN QC: KWW/FLX T1: KWW GRBT050-16 Olesen- 2057G GRBT050-Olesen-v6.cls July 9, 2005 4:30 160 Basic Science Aspects of the Headaches ◗ TABLE 16-1 Characteristics of Some Peptides Thought to be Involved in the Pathogenesis of Migraine Activity of Modulates Possible Animal Ligand Amino Relevance Receptor Model of Effective Neuropeptide Acids to Migraine Location Family Migraine in Migraine CGRP 37 Vasodilator Sensory nerves CGRP Yes, antagonist BIBN4096BS effective (CL/RAMP1) (either CGRP8−37 in phase II clinical or BIBN4096BS) trials decreases dural blood flow Substance P 11 Vasodilator; pain Sensory nerves Tachykinin Yes, NK1 NK1 receptor transmission; plasma family antagonist blocks antagonists not extravasation (NK1) dural plasma effective in acute extravasation migraine Somatostatin 14 (or 28) Inhibits sensory nerve Sensory nerves Somatostatin Yes octa peptide activity and other cells (sst1–sst5) benefits migraine pain Opioids Varies Inhibits sensory nerve Sensory nerves Opioid Yes, inhibits dural Yes, but use limited by (enkephalins, activity and other cells (µδκ and plasma abusive nature etc.) ORL-1) extravasation VIP 28 Vasodilator Found in range of VIP (VPAC) Not known Not known nerves Neuropeptide Y 36 Vasoconstrictor Sympathetic Y1–Y5 Not known Not known nerves gene, that are relevant to the cerebral vasculature. βCGRP nating in the trigeminal ganglia and innervates cerebral (or CGRPII), which has a high structural similarity (90%), blood vessels (30). CGRP,when given intravenously, acts in is primarily found in the gut and formed from a distinct a hypotensive manner, but it is generally considered that gene (57). the major activity of CGRP is local to site of release (12). The distribution of CGRP-containing nerves has been Plasma levels of CGRP are in the low picomolar level in nor- evaluated in detail in the cerebral circulation (23). CGRP mal volunteers (26) and migraineurs, but have been shown is contained in and released from sensory nerves origi- to increase in blood samples taken from the jugular vein ipsilateral to the attack (27). The release of CGRP occurs in response to nerve stim- CL ulation, and this has been studied in tissues in response CGRP receptor to chemical, physical, and mechanical stimulation in the CL/RAMP1 laboratory. It is important to note that the TRPV (or VR-1) Agonists: CGRP>AM receptor for capsaicin, the hot extract of chili peppers, ex- Antagonists: CGRP8-37 RAMP1 ists on the majority of CGRP-containing sensory nerves & BIBN4096BS (13) both in animals and in human trigeminal ganglion CL (39). The endogenous stimuli for this receptor are un- Adrenomedullin (AM1) receptor der study and include protons, noxious heat, and a range CL/RAMP2 of endogenous mediators, although their relative impor- Agonists: AM>>>CGRP tance in pathology is unclear (2,3,4). Furthermore, pre- RAMP2 Antagonist: AM22-52 synaptic/prejunctional receptors on the sensory nerves can modulate CGRP release. These receptors include CL those for opioids, 5-hydroxytryptamine (5-HT1 receptor), γ -aminobutyric acid (GABAB receptor), histamine (H3 re- Adrenomedullin (AM2) receptor ceptor), neuropeptide Y, somatostatin, VIP, purines, and CL/RAMP3 galanin (see below and Maggi [51]). Agonists: AM>CGRP RAMP3 CGRP receptors were classed as CGRP1 and CGRP2 in Antagonists: CGRP & AM 8-37 22-52 the late 1980s, as a consequence of pharmacologic studies. FIGURE 16-1. Summary of the characteristics of the CGRP re- The CGRP1 receptor is considered the important cardio- ceptor family. vascular receptor. The 8-37 amino acid fragment of CGRP, P1: KWW/KKL P2: KWW/HCN QC: KWW/FLX T1: KWW GRBT050-16 Olesen- 2057G GRBT050-Olesen-v6.cls July 9, 2005 4:30 Calcitonin Gene-Related Peptide and Other Peptides 161 CGRP8−37,isaselective antagonist for this receptor (14). There are multiple mechanisms by which CGRP pro- By comparison, the CGRP2 receptor remains poorly de- duces vasodilation that have been previously detailed fined (60). It is now realized that the CGRP family of re- (9,11). It is accepted that vascular relaxation is medi- ceptors consists of the seven transmembrane G protein- ated via the CGRP1 receptor (CL/RAMP1). This can occur coupled calcitonin receptor-like receptor CL with one of via nitric oxide–dependent endothelium-dependent mech- three single membrane-spanning receptor activity mod- anisms, as has been shown in mouse pial vessels (61), ifying proteins (RAMPs; 53). This complex is associated or cAMP endothelium–independent pathways. The latter with a CGRP-receptor component that is suggested to en- mechanism is observed in the majority of cerebral ves- hance receptor coupling and activation (25). The RAMP sels studied to date. CGRP acts directly on cerebral vessels molecule is important for localization of the functional to stimulate adenylate cyclase (40). The resulting rise in receptor to the cell surface and receptor phenotype, be- [cAMP]i activates protein kinase A, leading to phosphor- cause it influences ligand specificity (65). CL is a G protein- ylation, opening of potassium channels, and relaxation. In coupled receptor that is important for ligand binding. rat pial arterioles, vasodilatation to CGRP was inhibited in Three RAMPs (RAMP1, RAMP2, and RAMP3) are known. the presence of glibenclamide or charybdotoxin (a large- CL, when presented as a heterodimer with RAMP1 at the conductance Ca2+–activated K+ channel blocker) (37). cell surface, functions as a CGRP receptor that is antago- Exogenous CGRP can induce a migraine-like headache nized by CGRP8−37 and the nonpeptide CGRP antagonist (45), but CGRP is not involved in tension-type headaches BIBN4096BS (see below). CL with RAMP2 produces an (6). By comparison, CGRP levels are increased in samples adrenomedullin (AM) receptor that is blocked by the weak taken from the draining jugular vein during the painful AM antagonist ADM22−52. CL with RAMP3 leads to an AM phases of both migraine and cluster headaches (27,28,30).