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Chronic Exposure of NG 108‐15 Cells to Opiate Agonists Does Not Alter the Amount of the Guanine Nucleotide‐Binding Proteins G i and G O
Author(s) -
Lang J.,
Costa T.
Publication year - 1989
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1989.tb08544.x
Subject(s) - pertussis toxin , g protein , dadle , guanine , (+) naloxone , chemistry , receptor , agonist , adp ribosylation , nucleotide , biochemistry , enkephalin , gs alpha subunit , microbiology and biotechnology , biology , opioid , nad+ kinase , enzyme , gene
We have characterized the pertussis toxin substrate in NG 108–15 cell membranes using site‐specific antisera and ADP‐ribosylation. Cell membranes contain two pertussis toxin‐sensitive guanine nucleotide‐binding protein α‐subunits (G α ) whose R f values in gel electrophoresis coincide with those of G α o and Gα i2 . The total quantity of G i and G o im‐munoreactivity amounted to 24.3 ± 2.8 pmol/mg, whereas only 1.5 ± 0.2 pmol/mg are capable of undergoing ADP‐ribosylation catalyzed by pertussis toxin. Pretreatment of cells with the agonist [D‐Ala 2 ,D‐Leu 2 ]‐enkephalin (DADLE) for 24 h and DADLE or morphine for 72 h did not alter the incorporation of ADP‐ribose or the immunoreactive amount of G i and G o subunits. However, pretreatment for 72 h with naloxone increased the incorporation of ADP‐ribose without an apparent change in affinity or in the immunochemically determined protein levels of G i and G o . This indicates that the process of down‐regulation and desensitization of the γ‐opioid receptor neither requires quantitative alterations in the levels of G i and G o nor changes in the degree of coupling among their subunits. In contrast, chronic exposure to antagonists seems to alter the degree of precoupling between α ‐and β ‐subunits of G i and/or G o .