Opiate-dependent changes in the sensitivity of adenylate cyclase to stimulatory agonists and 5'-guanylylimidodiphosphate are independent of G protein abundance and eukaryotic ADP-ribosyltransferase activity in NG108-15 cells.
NG108-15 cells were exposed in culture to 1 microM [D-Ala2,D-Leu5]enkaphalin (DADLE) for 17 h. This treatment increased the maximum iloprost- and 5'-(N-ethylcarboxamido)adenosine-dependent activation of adenylate cyclase, as well as basal enzyme activity. In addition, there was an increase in the capacity of 5'-guanylylimidodiphosphate [Gpp(NH)p] to inhibit adenylate cyclase activity by direct interaction with the alpha-subunit of the Gi regulatory protein. A similar effect was observed if the cells were exposed to 10 microM carbachol. These treatments of NG108-15 cells did not alter the capacity of NaF to activate adenylate cyclase by direct interaction with Gs alpha. Exposure of NG108-15 cells to DADLE alone or DADLE plus carbachol had no effect on the capacity of pertussis toxin to ADP-ribosylate membrane proteins in these cells; neither was there any change in the activity of eukaryotic ADP-ribosyltransferase expressed in these cells. Under these conditions, the endogenous enzyme did not label any protein with a molecular mass similar to Gi alpha, 41 kDa. Treatment of the cells with DADLE or carbachol had no effect on the abundance of Gs alpha, Gi alpha, or G beta. The underlying mechanism for the changes in agonist-dependent stimulatory responses or Gpp(NH)p-dependent inhibition of adenylate cyclase remains obscure, but appears not to be mediated by eukaryotic ADP-ribosyltransferase activity or a change in the abundance of G proteins known to regulate adenylate cyclase.
Boyd
R S
RS
Department of Clinical Pharmacology, Royal Postgraduate Medical School, London, England.
Donnelly
L E
LE
MacDermot
J
J
D000067956:Adenylyl Cyclase Inhibitors; D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D002217:Carbachol / Q000494:pharmacology; D004338:Drug Combinations; D016308:Enkephalin, Leucine-2-Alanine / Q000494:pharmacology*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology*; D015151:Immunoblotting; D009536:Niacinamide / Q000494:pharmacology; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D000067956:Adenylyl Cyclase Inhibitors; D004338:Drug Combinations; D009536:Niacinamide; D006165:Guanylyl Imidodiphosphate; D016308:Enkephalin, Leucine-2-Alanine; D002217:Carbachol; D011065:Poly(ADP-ribose) Polymerases; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases