Cyclic AMP-mediated desensitization of D1 dopamine receptor-coupled adenylyl cyclase in NS20Y neuroblastoma cells.
Cyclic AMP-mediated desensitization of D1 dopamine receptor-coupled adenylyl cyclase was investigated using NS20Y neuroblastoma cells. Pretreatment of the cells for 24 h with 8-(4-chlorophenylthio)-adenosine-3':5'-cyclic monophosphate (CPT-cAMP), a membrane-permeable analog of cAMP, resulted in an approximately 90% reduction of the maximum dopamine-stimulated adenylyl cyclase activity. In addition, there was a twofold reduction in the potency of dopamine for stimulating cAMP production that was not dependent on the concentration of Mg2+ in the assay. These effects of CPT-cAMP pretreatment were time dependent, showing a t1/2 of about 3 h and a maximum reduction after about 8 h. Receptor-binding activity, as measured using the D1-selective antagonist [3H]SCH-23390, also declined following CPT-cAMP pretreatment with a t1/2 of about 5 h and a maximum reduction of about 70% after 20 h. Saturation analysis indicated that the loss in radioligand binding was due to a reduction in maximum binding capacity (Bmax) with no alteration in receptor affinity (KD). The EC50 of CPT-cAMP for producing enzyme desensitization and D1 receptor downregulation was determined to be about 30 microM with a maximal response occurring at 1 mM. These regulatory effects of CPT-cAMP were pharmacologically specific as other analogs of cAMP, such as dibutryl-cAMP, 8-bromo-cAMP, and Sp-cAMPS, were capable of inducing D1 receptor desensitization and downregulation, whereas treatment of the cells with the cAMP antagonist Rp-cAMPS had no effect. Conversely, Rp-cAMPS was capable of blocking the regulatory effects of CPT-cAMP but was apparently without effect in blocking dopamine-induced desensitization.(ABSTRACT TRUNCATED AT 250 WORDS)
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Molecular Neuropharmacology Section, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland 20892.
Smyk-Randall
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Sibley
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D000262:Adenylyl Cyclases / Q000378:metabolism*; D001552:Benzazepines / Q000378:metabolism; D001667:Binding, Competitive; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000494:pharmacology / Q000502:physiology*; D004298:Dopamine / Q000378:metabolism; D015536:Down-Regulation; D009447:Neuroblastoma / Q000378:metabolism* / Q000473:pathology; D017447:Receptors, Dopamine D1 / Q000378:metabolism*; D013873:Thionucleotides / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D001552:Benzazepines; D017447:Receptors, Dopamine D1; D013873:Thionucleotides; C015929:8-((4-chlorophenyl)thio)cyclic-3',5'-AMP; D000242:Cyclic AMP; D000262:Adenylyl Cyclases; D004298:Dopamine