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 t he cloned rat mu-opioid receptor (mu OR) were raised in rabbits. The a ntibodies diminished the specific binding of I-125-Tyr(27)-beta-endorp hin-(1-31) (human) and [H-3][D-Ala(2),N-MePhe(4),Gly-ol(5)]enkephalin, but not that of the delta OR-selective ligand [H-3][D-Pen(2,5)]enkeph alin, to mouse brain membranes. The intracerebroventricular administra tion to mice of affinity-purified anti-mu OR IgGs impaired the antinoc iception produced by the mu OR agonists [D-Ala(2),N-MePhe(4),Gly-ol(5) ]enkephalin and morphine and the mu/delta OR agonists beta-endorphin-( 1-31) and [D-Ala(2),D-Leu(5)]enkephalin, when studied 24 hr later in t he tail-immersion test. Antinociception produced by the delta OR-selec tive agonists [D-Pen(2,5)]enkephalin and [D-Ala(2)]deltorphin II was f ully displayed in these mice. Immunoblots of sodium dodecyl sulfate-so lubilized 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 sonicat ion of the solubilized tissues, This immunoreactivity was present in r egions of mouse central nervous system and was barely detected in NG10 8-15 cells. After treatment of the solubilized material with endoglyco sidase F, the antibodies labeled a band of M(r) 43,000, coincident wit h the weight of the cloned mu OR. These results confirm the existence of several molecular forms of the mu OR due to glycosylation.