Opioids and Their Complicated Receptor Complexes Bryen A. Jordan, Ph.D., Svetlana Cvejic, Ph.D., and Lakshmi A. Devi, Ph.D. No field more eagerly awaits a molecular clarification for that opioid receptors and a number of other GPCRs exist as G-protein coupled receptor (GPCR) dimerization than the dimers in biochemical, functional and pharmacological assays. opioid receptor field. Extensive evidence of pharmacological It is with these new tools that we seek to understand the and functional interactions between opioid receptor types has mechanisms and implications of dimerization. Initial results primed this field for such a resolution. In retrospect, much of of these studies have demonstrated that the dimerization of the data collected on synergy between different opioid receptor opioid receptors may help consolidate several pharmacological types may represent the functional correlate for the newly findings that have remained unanswered. In this review we found opioid receptor dimerization. While previous reports of present biochemical, pharmacological and functional evidence functional synergy have been, for the most part, consistent in for opioid receptor complexes and add evidence from our demonstrating cross-regulation between two receptor types, recent studies on opioid receptor dimerization. We believe a the lack of highly receptor-selective ligands allowed skeptics to thorough understanding of receptor dimerization is crucial in remain doubtful over the interpretations of these results. clarifying the mechanism of action of opioids and other drugs Today, two important developments in the opioid receptor and may serve a more practical purpose in aiding the field help reinvigorate the hypothesis of functional, cross- development of novel therapeutic drugs. modulating opioid receptor complexes: (1) The existence of [Neruopsychopharmacology 23:S5–S18, 2000] © 2000 highly selective ligands which eliminate any possibility of American College of Neuropsychopharmacology. Published cross-reactivity between receptor types, and (2) the discovery by Elsevier Science Inc. KEY WORDS: Opioids; Oligomerization; Signal transduction; that of pain control, where opioids are known to inhibit G-proteins; Dimers; Heterodimers; Complex; Subtype; neurotransmitter release from dorsal root ganglion pro- Synergy jections in the dorsal horn of the spinal cord (Mac- Ancient civilizations used opium for its medicinal use. Donald and Nelson 1978; Mudge et al. 1979; Yaksh Current research has shown that the administration of 1993). Opioids can modulate endocrine processes opioids results in a variety of biological effects. These (Genazzani and Petraglia 1989; Maggi et al. 1995; Scha- include analgesia, miosis, bradycardia, general seda- fer and Martin 1994) and can also affect the immune re- tion, hypothermia, insensitivity and depression of sponse (Brown et al. 1974; Roy and Loh 1996). All these flexor reflexes. However, the best studied effect is in effects are mediated by the interactions of opioids with members of the opioid receptor family. Opioid binding sites were first discovered in mam- malian brain (Pert and Snyder 1973; Simon et al. 1973; From the Department of Pharmacology, New York University School of Medicine, 550 First Avenue, New York, NY 10016. Terenius 1973). Subsequent pharmacological studies Address correspondence to: Lakshmi A. Devi, Ph.D., Department using different ligands uncovered three opioid receptor of Pharmacology, New York University School of Medicine, MSB types, kappa, delta and mu (Chang and Cuatrecasas 411, 550 First Avenue, New York, NY 10016. Tel.: (212) 263-7119; Fax: (212) 263-7133; E-mail: [email protected] 1979; Lord et al. 1977) (reviewed in Paterson et al. 1983) Received April 6, 2000; accepted May 1, 2000. which differed in their distribution, ligand binding and NEUROPSYCHOPHARMACOLOGY 2000–VOL. 23, NO. S4 © 2000 American College of Neuropsychopharmacology Published by Elsevier Science Inc. 0893-133X/00/$–see front matter 655 Avenue of the Americas, New York, NY 10010 PII S0893-133X(00)00143-3 S6 B.A. Jordan et al. NEUROPSYCHOPHARMACOLOGY 2000–VOL. 23, NO. S4 function. The endogenous ligands were found in the first opioid binding sites (Pert and Snyder 1973; Simon mid-1970s and were shown to be peptides (Bradbury et et al. 1973; Terenius 1973). Other agonists, ketocyclazo- al. 1976; Cox et al. 1976; Goldstein et al. 1981; Hughes et cine, morphine and DADLE, were subsequently shown al. 1975; Pasternak et al. 1976). These include enkepha- to compete against three different fractions of total opi- lins, dynorphins and endorphins which are derived oid receptor binding (see Table 1 for ligands and selec- from three larger precursors: proenkephalin A (Noda et tivities). This resulted in the classification of opioid re- al. 1982), prodynorphin (Kakidani et al. 1982) and pro- ceptors into three types: kappa (for ketocyclazocine), opioimelanocortin, respectively (Nakanishi et al. 1979). mu (for morphine) and delta (for DADLE) (Chang and Recently two new mu receptor selective peptides Cuatrecasas 1979; Lord et al. 1977). As the number of named endomorphin-1 and endomorphin-2 have been opioid specific ligands grew, so did the list of apparent reported although the presence of a precursor for these opioid receptor subtypes found in pharmacological as- has not yet been determined (Zadina et al. 1997). says. For example, the new delta receptor selective The search for novel opioids resulted in the identifi- compounds deltorphin II, DALCE, BNTX and naltrin- cation of a large number of opioid ligands which ex- dole isothiocyanante (NTII) could differentially com- hibit a variety of efficacies, potencies, affinities and se- pete with selective binding to delta receptors in spinal lectivities for the opioid receptor types. The use of these cord and brain, suggesting the presence of subpopula- led to the identification of similar yet distinct receptor tions within the delta receptor type. There are now sev- subtypes. The pharmacological differences observed eral opioid receptor subtypes and the list may grow as led to the proposal of receptor subtypes (kappa1, novel compounds are identified. However, a central kappa2, kappa3; delta1, delta2; mu1, mu2, mu3) (for re- concern pertaining to this subject remains the lack of view see Traynor 1989; Traynor and Elliot 1993). How- cDNAs corresponding to these subtypes. The cloning of ever to date, despite large-scale efforts to clone the opi- the opioid receptors revealed only three cDNAs corre- oid receptor subtypes, no cDNA corresponding to a sponding to the original mu, delta and kappa receptor receptor subtype has been found. types. A mu receptor isoform named MOR1b has also The cloning of the opioid receptors (Evans et al. 1992; been cloned and differs in the c-terminal tail (Zimprich Kieffer et al. 1992; Yasuda et al. 1993; Chen et al. 1993) et al. 1995). However, the pharmacological profile of revealed that they are members of the GPCR superfam- this isoform appears to be identical to that of the previ- ily. The amino acid sequence revealed no more than a ously cloned mu receptor. Therefore the pharmacologi- 30% homology to other GPCRs whereas a comparison cal entities may not have a genetic correlate and thus between kappa, delta and mu receptors revealed a 65– warrant other explanations. Along with the question of 70% homology between the three with highest homol- receptor subtypes and prior to the postulation of opioid ogy among the transmembrane domains, intracellular receptor subtypes, another pharmacological phenome- loops and a small portion of the C-terminal tail near the non had many researchers perplexed. The apparent in- 7th transmembrane region (TM7). The most divergent teraction of receptor type selective compounds in both areas are the second and third extracellular loops as pharmacological and functional assays led to one of the well as the N- and C-terminal tail which are extracellu- first discussions of receptor “complexing,” or more re- lar and intracellular, respectively. cently known as receptor “dimerization.” From phar- macological assays, functional complexes of mu and delta receptors were proposed to exist. Interestingly, OPIOID RECEPTOR COMPLEXES characterization of these “receptor complexes” yielded information linking them to certain opioid receptor One of the principal goals in opioid receptor research is subtypes and led some to propose that a mu-delta com- to dissociate the analgesic potential of opioid ligands plex may in fact represent one of the delta receptor sub- from their liability to be abused. While an understand- types. Our work with kappa/delta receptor het- ing of the molecular nature of receptor regulation is erodimers suggests that these may represent a kappa crucial for this undertaking, the discovery of ligands receptor subtype (Jordan and Devi 1999). The following with a decreased addicitive component has remained is the evidence collected from several labs to support the simplest approach. In spite of an enormous effort, the premise of opioid receptor dimerization. such a compound has yet to be found. However, this search has resulted in a large library of compounds that exhibit varying degrees of efficacy, affinities, potencies FUNCTION and selectivities for the three opioid receptor types. Standard ligand binding assays using these compounds The administration of morphine results in a powerful were crucial in the classification of opioid binding sites. analgesic response that is mediated primarily by mu The stereospecific