Identification and Characterization of Three New Alternatively Spliced μ-Opioid Receptor Isoforms

Abstract

We have identified four new mu-opiod receptor (MOR)-1 exons, indicating that the gene now contains at least nine exons spanning more than 200 kilobases. Replacement of exon 4 by combinations of the new exons yields three new receptors. When expressed in Chinese hamster ovary cells, all three variants displayed high affinity for mu-opioid ligands, but kappa and delta drugs were inactive. However, there were subtle, but significant, differences in the binding profiles of the three variants among themselves and from MOR-1. Immunohistochemically, the major variant, MOR-1C, displayed a regional distribution quite distinct from that of MOR-1. Region-specific processing also was seen at the mRNA level. Antisense mapping revealed that the four new exons were all involved in morphine analgesia. Together with two other variants generated from alternative splicing of exon 4, there are now six distinct MOR-1 receptors.

Morphine‐6β‐glucuronide has a higher efficacy than morphine as a mu‐opioid receptor agonist in the rat locus coeruleus

“…M6G is also more potent than morphine (146 : 1) at producing rewarding e ects measured with place preference testing following intracerebral administration (Abbott & Franklin, 1991). Binding studies show that M6G has an equal or lower a nity for the muopioid receptor than morphine (Abbott & Palmour, 1988;Chen et al, 1991;Hucks et al, 1992;Lambert et al, 1993;Mignat et al, 1995;LoÈ ser et al 1996;Brown et al, 1997b;Pan et al, 1999). In order to explain these di erences in the potencies of M6G and morphine, it has recently been hypothesized that M6G has selective antinociceptive and other behavioural agonist e ects at a receptor that is not activated by morphine (reviewed by Pasternak & Standifer, 1995).…”

Alternatively spliced mu opioid receptor C termini impact the diverse actions of morphine

“…The truncations had little effect on ED 50 values, but significantly reduced DAMGO-and morphine-induced maxi- ]DAMGO), a mu agonist, with high affinity and with only subtle differences in selectivity for the endogenous opioid peptides dynorphin A and β endorphin (18,25,26). However, in [ 35 S]GTPγS-binding assays, these C-terminal splice variants displayed marked differences in mu opioid-induced G protein coupling in both potency (EC 50 ) and efficacy (% of maximal stimulation) (26,27), suggesting that the distal carboxyl terminal sequences influence mu agonist-induced receptor-G protein coupling and signal transduction.…”

Involvement of exon 11-associated variants of the mu opioid receptor MOR-1 in heroin, but not morphine, actions

“…Total RNA was extracted from mouse brain with the guanidinium thiocyanate/phenol/chloroform extraction method (26), treated with DNase I by using TURBO DNA-free reagents (Ambion) to remove potentially contaminating genomic DNA, and reversed transcribed with random hexamer or oligo(dT) and SuperScript II reverse transcriptase (Invitrogen) as previously described (9). The reverse transcriptase product was treated with RNase H to remove RNA complementary to the first-strand cDNA.…”

“…This inability to reconcile the diverse pharmacology of -opioids with their binding selectivity led to the proposal of multiple -opioid-receptors (7), a concept initially proposed based on binding and pharmacological studies in animal models and subsequently confirmed with the identification of a large number of splice variants of the cloned -opioid-receptor MOR-1 in mice, rats, and humans ( Fig. 1A) (8)(9)(10)(11)(12)(13)(14)(15).…”