TY - JOUR
T1 - Characterization and Distribution of a Cloned Rat μ‐Opioid Receptor
AU - Bunzow, James R.
AU - Zhang, Ge
AU - Bouvier, Claudia
AU - Saez, Carmen
AU - Ronnekleiv, Oline K.
AU - Kelly, Martin J.
AU - Grandy, David K.
PY - 1995/1
Y1 - 1995/1
N2 - Abstract: We have cloned and expressed a rat brain cDNA, TS11, that encodes a μ‐opioid receptor based on pharmacological, physiological, and anatomical criteria. Membranes were prepared from COS‐7 cells transiently expressing TS11 bound [3H]diprenorphine with high affinity (KD = 0.23 ± 0.04 nM). The rank order potency of drugs competing with [3H]diprenorphine was as follows: levorphanol (Ki = 0.6 ± 0.2 nM) ≈β‐endorphin (Ki = 0.7 ± 0.5 nM) ≈ morphine (Ki = 0.8 ± 0.5 nM) ≈ [d‐Ala2, N‐Me‐Phe4,Gly‐ol5]‐enkephalin (DAMGO; Ki = 1.6 ± 0.5 nM) ⋙ U50,488 (Ki = 910 ± 0.78 nM) > [d‐Pen2,5]‐enkephalin (Ki = 3,170 ± 98 nM) > dextrorphan (Ki = 4,100 ± 68 nM). The rank order potencies of these ligands, the stereospecificity of levorphanol, and morphine's subnanomolar Ki are consistent with a μ‐opioid binding site. Two additional experiments provided evidence that this opioid‐binding site is functionally coupled to G proteins: (a) In COS‐7 cells 50 µM 5′‐guanylylimidodiphosphate shifted a fraction of receptors with high affinity for DAMGO (IC50 = 3.4 ± 0.5 nM) to a lower‐affinity state (IC50 = 89.0 ± 19.0 nM), and (b) exposure of Chinese hamster ovary cells stably expressing the cloned μ‐opioid receptor to DAMGO resulted in a dose‐dependent, naloxone‐sensitive inhibition of forskolin‐stimulated cyclic AMP production. The distribution of mRNA corresponding to the μ‐opioid receptor encoded by TS11 was determined by in situ hybridization to brain sections prepared from adult female rats. The highest levels of μ‐receptor mRNA were detected in the thalamus, medial habenula, and the caudate putamen; however, significant hybridization was also observed in many other brain regions, including the hypothalamus.
AB - Abstract: We have cloned and expressed a rat brain cDNA, TS11, that encodes a μ‐opioid receptor based on pharmacological, physiological, and anatomical criteria. Membranes were prepared from COS‐7 cells transiently expressing TS11 bound [3H]diprenorphine with high affinity (KD = 0.23 ± 0.04 nM). The rank order potency of drugs competing with [3H]diprenorphine was as follows: levorphanol (Ki = 0.6 ± 0.2 nM) ≈β‐endorphin (Ki = 0.7 ± 0.5 nM) ≈ morphine (Ki = 0.8 ± 0.5 nM) ≈ [d‐Ala2, N‐Me‐Phe4,Gly‐ol5]‐enkephalin (DAMGO; Ki = 1.6 ± 0.5 nM) ⋙ U50,488 (Ki = 910 ± 0.78 nM) > [d‐Pen2,5]‐enkephalin (Ki = 3,170 ± 98 nM) > dextrorphan (Ki = 4,100 ± 68 nM). The rank order potencies of these ligands, the stereospecificity of levorphanol, and morphine's subnanomolar Ki are consistent with a μ‐opioid binding site. Two additional experiments provided evidence that this opioid‐binding site is functionally coupled to G proteins: (a) In COS‐7 cells 50 µM 5′‐guanylylimidodiphosphate shifted a fraction of receptors with high affinity for DAMGO (IC50 = 3.4 ± 0.5 nM) to a lower‐affinity state (IC50 = 89.0 ± 19.0 nM), and (b) exposure of Chinese hamster ovary cells stably expressing the cloned μ‐opioid receptor to DAMGO resulted in a dose‐dependent, naloxone‐sensitive inhibition of forskolin‐stimulated cyclic AMP production. The distribution of mRNA corresponding to the μ‐opioid receptor encoded by TS11 was determined by in situ hybridization to brain sections prepared from adult female rats. The highest levels of μ‐receptor mRNA were detected in the thalamus, medial habenula, and the caudate putamen; however, significant hybridization was also observed in many other brain regions, including the hypothalamus.
KW - Cyclic AMP
KW - G protein coupling
KW - In situ hybridization
KW - Receptor binding
KW - μ‐Opioid receptors
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U2 - 10.1046/j.1471-4159.1995.64010014.x
DO - 10.1046/j.1471-4159.1995.64010014.x
M3 - Article
C2 - 7798908
AN - SCOPUS:0028888556
VL - 64
SP - 14
EP - 24
JO - Journal of Neurochemistry
JF - Journal of Neurochemistry
SN - 0022-3042
IS - 1
ER -