Induced Association of μ Opioid (MOP) and Type 2 Cholecystokinin (CCK₂) Receptors by Novel Bivalent Ligands

Abstract

Both mu opioid (MOP) † and type 2 cholecystokinin (CCK 2 ) receptors are present in areas of the central nervous system that are involved in modulation of pain processing. We conducted bioluminescence resonance energy transfer (BRET) studies on COS cells coexpressing MOP and CCK 2 receptors to determine whether receptor heterodimerization is involved in such modulation. These studies revealed the absence of constitutive or monovalent ligand-induced heterodimerization. Heterodimerization of MOP and CCK 2 receptors therefore is unlikely to be responsible for the opposing effects between morphine and CCK in the CNS. However, association was induced, as indicated by a positive BRET signal, on exposure of the cells to bivalent ligands containing muopioid agonist and CCK 2 receptor antagonist pharmacophores linked through spacers containing 16 to 22 atoms, but not with a shorter (9-atom) spacer. These studies demonstrate for the first time that an appropriately designed bivalent ligand is capable of inducing association of G protein-coupled receptors. The finding that opioid tolerance studies with these ligands in mice showed no correlation with the BRET data is consistent with the absence of association of MOP and CCK 2 receptors in vivo.

Expression of Cholecystokinin, Gastrin, and Their Receptors in the Mouse Cornea

“…57 On another hand, peripheral nerve injury causes a marked increase of CCK levels in DRG neurons. 24 Considering the ''anti-opioid'' role described for CCK, 37,41,58,59 the possibility exists that in injured tissues, including the cornea, increased CCK levels antagonize peripheral opioid receptors, thus counteracting the inhibitory effects of peripheral opioids on nociceptor sensitization during inflammation.…”

N -naphthoyl-β-naltrexamine (NNTA), a highly selective and potent activator of μ/κ-opioid heteromers

“…Moreover, it has been shown that μ-opioid and CCK2 receptors do not form constitutive heteromers when overexpressed in Chinese hamster ovary (CHO) cells, but can be induced to form heteromers in the presence of a bivalent ligand containing μ-agonist and CCK antagonist pharmacophores (25). Thus, heteromerization does not appear to be an artifact related to receptor overexpression in cultured cells.…”

Twin and Triplet Drugs in Opioid Research

“…Recently, to investigate the possible existence of heterodimeric m opioid/ CCK 2 receptors, which may be responsible for the pharmacological observations mentioned above, the twin drugs 71 and 72 possessing the m agonist oxymorphone (45) [71] and the CCK 2 antagonist L-365,260 (70) [150] with spacers of various lengths were designed (Fig. 16) [151]. From the bioluminescence resonance energy transfer (BRET) studies [152], it is suggested that the m opioid receptor and the CCK 2 receptor would form homodimers in the COS cells.…”