Molecular cloning, tissue distribution and chromosomal localization of a novel member of the opioid receptor gene family (original) (raw)
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Molecular pharmacology of the opioid receptors
Pharmacology & Therapeutics, 1995
Opioid receptors are the primary sites of actions of opiates and endogenous opioid peptides, which have a wide variety of pharmacological and physiological effects. The opioid receptors are classified into at least three subtypes, p, 6, and x, and their cDNAs have been cloned. In this review, we describe the molecular cloning of opioid receptor gene family and studies of the structure-function relationships, modes of coupling to second messenger systems, pharmacological effects of antisense oligonucleotides, and anatomical distribution of opioid receptor mRNAs.
Tissue Distribution of Opioid Receptor Gene Expression in the Rat
Biochemical and Biophysical Research Communications, 1996
The endogenous opioid peptides have multiple physiological actions at both a central and peripheral level which are mediated by 3 main classes of opioid receptors, , ␦ and. The rat , ␦ and opioid receptors have recently been cloned and their distribution of expression in the central nervous system has been mapped. In these studies we have used the reverse transcriptase polymerase chain reaction and Southern blotting to determine the distribution of expression of the , ␦ and opioid receptors in the peripheral tissues of the rat. All 3 opioid receptors were found to be widely expressed in several peripheral tissues including the small intestine, large intestine, adrenal, kidney, lung, spleen, testis, ovary and uterus. In the stomach, ␦ and but not opioid receptor transcripts were detected. Predominantly ␦ transcripts were detected in the heart, with no and only a weak signal for the receptor. In the liver and ␦ but not transcripts were present. Opioid receptor expression was not detected in endothelium or synovium. There is therefore a broad, but tissue specific distribution of opioid receptor expression in the periphery of the rat, suggesting that the endogenous opioid peptides play an endocrine, paracrine, or autocrine role in the regulation of physiology at a peripheral as well as central level.
Cloning and Pharmacological Characterization of a Rat kappa Opioid Receptor
Proceedings of The National Academy of Sciences, 1993
A full-length cDNA was isolated from a rat striatal library by using low-stringency screening with two PCR fragments, one spanning transmembrane domains 3-6 of the mouse δ opioid receptor and the other unidentified but homologous to the mouse δ receptor from rat brain. The novel cDNA had a long open reading frame encoding a protein of 380 residues with 59% identity to the mouse δ receptor and topography consistent with a seven-helix guanine nucleotide-binding protein-coupled receptor. COS-1 cells transfected with the coding region of this clone showed high-affinity binding to kappa opioid receptor-selective ligands such as dynorphin A and U-50,488 and also nonselective opioid ligands such as bremazocine, ethylketocyclazocine, and naloxone. Not bound at all (or bound with low affinity) were dynorphin A-(2-13), enantiomers of naloxone and levophanol [i.e., (+)-naloxone and dextrorphan], and selective μ and δ opioid receptor ligands. Activation of the expressed receptor by kappa receptor agonists led to inhibition of cAMP. Finally, in situ hybridization revealed a mRNA distribution in rat brain that corresponded well to the distribution of binding sites labeled with kappa-selective ligands. These observations indicate that we have cloned a cDNA encoding a rat kappa receptor of the kappa_1 subtype.
Cloning, Characterization, and Distribution of a μ-Opioid Receptor in Rat Brain
Journal of Neurochemistry, 2008
Abstract: We report the isolation and characterization of a rat cDNA clone encoding a μ-opioid receptor. This receptor, a 398 amino acid protein, shares 59% overall identity with the mouse Δ-and K-opioid receptors. Transient expression of the receptor in COS cells revealed high-affinity binding of μ-selective opioid antagonists and agonists, with a KD for naloxone ∼1.5 nM, and for [D-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin (DAMGO) and morphine at the high-affinity site of 2–4 nM, confirming a μ-opioid pharmacological profile. Northern blotting and in situ hybridization histoohemistry revealed that the μ-opioid receptor mRNA was expressed in many brain regions, including cerebral cortex, caudate putamen, nucleus accumbens, olfactory tubercle, septal nuclei, thalamus, hippocampus, and medial habenular nucleus, in keeping with the known distribution of the μ-opioid receptor.
Mu, delta, and kappa opioid receptor mRNA expression in the rat CNS: An in situ hybridization study
The Journal of Comparative Neurology, 1994
The p, 6, and K opioid receptors are the three main types of opioid receptors found in the central nervous system (CNS) and periphery. These receptors and the peptides with which they interact are important in a number of physiological functions, including analgesia, respiration, and hormonal regulation. This study examines the expression of p, 6, and K receptor mRNAs in the rat brain and spinal cord using in situ hybridization techniques. Tissue sections were hybridized with 35S-labeled cRNA probes to the rat IJ. (744-1,064 b), 6 (304-1,287 b), and K (1,351-2,124 b) receptors. Each mRNA demonstrates a distinct anatomical distribution that corresponds well to known receptor binding distributions. Cells expressing p receptor mRNA are localized in such regions as the olfactory bulb, caudate-putamen, nucleus accumbens, lateral and medial septum, diagonal band of Broca, bed nucleus of the stria terminalis, most thalamic nuclei, hippocampus, amygdala, medial preoptic area, superior and inferior colliculi, central gray, dorsal and median raphe, raphe magnus, locus coeruleus, parabrachial nucleus, pontine and medullary reticular nuclei, nucleus ambiguus, nucleus of the solitary tract, nucleus gracilis and cuneatus, dorsal motor nucleus of vagus, spinal cord, and dorsal root ganglia. Cellular localization of 6 receptor mRNA varied from p or K, with expression in such regions as the olfactory bulb, allo-and neocortex, caudate-putamen, nucleus accumbens, olfactory tubercle, ventromedial hypothalamus, hippocampus, amygdala, red nucleus, pontine nuclei, reticulotegmental nucleus, motor and spinal trigeminal, linear nucleus of the medulla, lateral reticular nucleus, spinal cord, and dorsal root ganglia. Cells expressing K receptor mRNA demonstrate a third pattern of expression, with cells localized in regions such as the claustrum, endopiriform nucleus, nucleus accumbens, olfactory tubercle, medial preoptic area, bed nucleus of the stria terminalis, amygdala, most hypothalamic nuclei, median eminence, infundibulum, substantia nigra, ventral tegmental area, raphe nuclei, paratrigeminal and spinal trigeminal, nucleus of the solitary tract, spinal cord, and dorsal root ganglia. These findings are discussed in relation to the physiological functions associated with the opioid receptors. D 1% Wiley-Liss, Inc.
Molecular Brain Research, 1994
A rat brain cDNA library was screened for clones homologous to the recently cloned mouse 8-opioid receptor (DOR-1). Among the clones isolated was Hyp 8-1, a clone with a unique nucleotide sequence capable of encoding a putative protein which is 57-58% identical to the amino acid sequences of the cloned 3,/~ and r opioid receptors, indicating a close relationship of Hyp 8-1 with the opioid receptor family. Several cDNAs representing possible splice variants of Hyp 8-1 were also isolated. Binding studies of COS-7 cells transfected with clone Hyp 8-1 failed to demonstrate specific binding with several 3H-opioid ligands. In situ hybridization studies indicate that the mRNA for Hyp 8-1 is distributed discretely throughout the rat brain, in an overall pattern which is different from that of several other G-protein-coupled seven transmembrane receptors. Thus, it is likely that the Hyp 8-1 cDNA encodes a novel peptide receptor.