Antibodies to the cloned mu-opioid receptor detect various molecular weight forms in areas of mouse brain.
Polyclonal antibodies directed against the amino-terminal portion of the cloned rat mu-opioid receptor (muOR) were raised in rabbits. The antibodies diminished the specific binding of 125I-Tyr27-beta-endorphin-(1-31) (human) and [3H][D-Ala2,N-MePhe4,Gly-ol5]enkephalin, but not that of the delta OR-selective ligand [3H][D-Pen2,5]enkephalin, to mouse brain membranes. The intracerebroventricular administration to mice of affinity-purified anti-muOR IgGs impaired the antinociception produced by the muOR agonists [D-Ala2,N-MePhe4,Gly-ol5]enkephalin and morphine and the mu/delta OR agonists beta-endorphin-(1-31) and [D-Ala2,D-Leu5]enkephalin, when studied 24 hr later in the tail-immersion test. Antinociception produced by the delta OR-selective agonists [D-Pen2,5]enkephalin and [D-Ala2]deltorphin II was fully displayed in these mice. Immunoblots of sodium dodecyl sulfate-solubilized membranes from mouse central nervous system regions revealed protein bands of M(r) 43,000, 51,000, and 58,000. Also detected were bands of higher molecular weights, 100,000 and 114,000, which probably corresponded to dimeric forms, because they disappeared after sonication of the solubilized tissues. This immunoreactivity was present in regions of mouse central nervous system and was barely detected in NG108-15 cells. After treatment of the solubilized material with endoglycosidase F, the antibodies labeled a band of M(r) 43,000, coincident with the weight of the cloned muOR. These results confirm the existence of several molecular forms of the muOR due to glycosylation.
Garzon
J
J
Instituto Cajal, Consejo Superior de Investigaciones Científicas, Madrid, Spain.
Juarros
J L
JL
Castro
M A
MA
Sanchez-Blazquez
P
P
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology*; D001923:Brain Chemistry; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; D004745:Enkephalins / Q000378:metabolism; D006168:Guinea Pigs; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D017450:Receptors, Opioid, mu / Q000032:analysis*; D001615:beta-Endorphin / Q000378:metabolism
D000906:Antibodies; D004745:Enkephalins; D010446:Peptide Fragments; D017450:Receptors, Opioid, mu; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; D001615:beta-Endorphin