Identification of three new alternatively spliced variants of the rat mu opioid receptor gene: dissociation of affinity and efficacy.
Mu opioid receptors mediate the pharmacological actions of morphine and morphine-like drugs, such as heroin. The mouse and human Oprm genes undergo splicing. In these present studies, we have identified and characterized three new MOR-1 splice variants from the rat Oprm gene. Using an RT-PCR approach, we isolated the new exons 7, 8 and 9 downstream of exon 3. The rat exons 7 and 9 were homologous to the mouse exons 7 and 9 while the rat exon 8 was not. Northern blot analysis with the new exon probes showed distinctive and abundant transcripts of the variants in the rat brain. Full-length cDNA clones containing the new exons, rMOR-1C1, rMOR-1C2 and rMOR-1D were obtained using an RT-PCR approach. Each contained the same exons 1, 2 and 3 as the original rMOR-1, followed by different combinations of the new exons in place of exon 4. In addition, we also isolated another new variant, rMOR-1A, which contains only exons 1, 2 and 3, and is homologous to the human variant MOR-1A previously identified. All the variants were highly mu-selective in binding studies with little difference in affinities for the mu ligands among the variants. However, functional evaluation of assessments of the variants using agonist stimulated [(35)S]GTPgammaS binding assays revealed marked differences among the variants, both in terms of potency and efficacy of the drugs. The relative efficacy of a series of mu opioids to each other varied depending upon the variant studied. Efficacy in the [(35)S]GTPgammaS assay did not correlate with either receptor binding affinity or with potency. Thus, selectivity of opioid action might be achieved by designing compounds with varying efficacies at different MOR-1 variants.
Pasternak
David A
DA
Laboratory of Molecular Neuropharmacology, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
Pan
Ling
L
Xu
Jin
J
Yu
Rui
R
Xu
Ming-Ming
MM
Pasternak
Gavril W
GW
Pan
Ying-Xian
YX
D017398:Alternative Splicing / Q000235:genetics*; D000818:Animals; D001667:Binding, Competitive / Q000235:genetics; D015152:Blotting, Northern; D001921:Brain / Q000378:metabolism; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D005091:Exons / Q000235:genetics; D018014:Gene Transfer Techniques; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000493:pharmacokinetics; D009294:Narcotics / Q000493:pharmacokinetics; D051381:Rats; D017450:Receptors, Opioid, mu / Q000096:biosynthesis / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017385:Sequence Homology; D013379:Substrate Specificity
D009294:Narcotics; D017450:Receptors, Opioid, mu; D016244:Guanosine 5'-O-(3-Thiotriphosphate)