Guanine nucleotide binding proteins and the regulation of cyclic AMP synthesis in NS20Y neuroblastoma cells: role of D1 dopamine and muscarinic receptors.
D1 dopamine receptors on NS20Y neuroblastoma cells stimulate adenylate cyclase activity, whereas muscarinic receptors on the same cells negatively regulate adenylate cyclase. To determine the mechanisms which underlie these processes, cyclic AMP accumulation was measured in intact cells following either cholera or pertussis toxin treatment. Pretreatment with pertussis toxin (100 ng/ml), which ribosylated greater than 95% of inhibitory quinine nucleotide binding protein (Gi), caused the complete loss of muscarinic induced inhibition. Conversely, pertussis toxin did not affect the ability of dihydrexidine (1 microM, a full efficacy D1 agonist), PGE1 (100 nM), or forskolin (1 microM, a direct activator) to stimulate cAMP accumulation. Both the dihydrexidine-induced stimulation and the carbachol-induced inhibition of cyclic AMP accumulation were unaffected by either removal of extracellular calcium, or increased intracellular calcium caused by the addition of the calcium ionophore A23187. Cholera toxin dose- and time-dependently induced large accumulations of cAMP. At low cholera toxin concentrations, the effects of dihydrexidine (300 nM) were additive with those of cholera toxin. At cholera toxin concentrations greater than 100 ng/ml, dihydrexidine became ineffective in stimulating further cAMP synthesis. Conversely, forskolin (1 microM) still caused marked increases in cAMP accumulation after all cholera toxin treatments. Dihydrexidine-stimulated cAMP accumulation was additive with forskolin-stimulated cAMP accumulation at low forskolin concentrations (10 nM-3 microM), but synergistic at high concentrations (3-100 microM). Additionally, forskolin was much more potent after cholera toxin treatment, suggesting that an activated stimulatory guanine nucleotide binding protein (Gs) may be required for full activation of adenylate cyclase by forskolin in this cell type.(ABSTRACT TRUNCATED AT 250 WORDS)
Lovenberg
T W
TW
Department of Pharmacology, University of North Carolina School of Medicine, Chapel Hill 27599.
Nichols
D E
DE
Nestler
E J
EJ
Roth
R H
RH
Mailman
R B
RB
D037361:Adenylate Cyclase Toxin; D000527:Alprostadil / Q000494:pharmacology; D000818:Animals; D002217:Carbachol / Q000494:pharmacology*; D002460:Cell Line; D002772:Cholera Toxin / Q000494:pharmacology*; D005576:Colforsin / Q000494:pharmacology*; D000242:Cyclic AMP / Q000096:biosynthesis*; D015259:Dopamine Agents / Q000494:pharmacology*; D019204:GTP-Binding Proteins / Q000502:physiology*; D007700:Kinetics; D009447:Neuroblastoma; D037342:Pertussis Toxin; D010617:Phenanthridines / Q000494:pharmacology*; D011954:Receptors, Dopamine / Q000187:drug effects / Q000502:physiology*; D017447:Receptors, Dopamine D1; D011976:Receptors, Muscarinic / Q000187:drug effects / Q000502:physiology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*
D037361:Adenylate Cyclase Toxin; D015259:Dopamine Agents; D010617:Phenanthridines; D011954:Receptors, Dopamine; D017447:Receptors, Dopamine D1; D011976:Receptors, Muscarinic; D010566:Virulence Factors, Bordetella; D005576:Colforsin; C061532:dihydrexidine; D002217:Carbachol; D002772:Cholera Toxin; D000242:Cyclic AMP; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins; D000527:Alprostadil