Plasma membrane-bound cyclic AMP phosphodiesterase activity in 3T3-L1 adipocytes.
Plasma membranes were isolated from 3T3-L1 adipocytes. Plasma membrane phosphodiesterase (PM-PDE) was measured in the presence of 5 microM cilostamide. Time course and cAMP dose response ranging from 0 to 2 microM were measured. PM-PDE remained linear up to 20 min. Non-linear curve fitting analysis showed that the low Km cAMP dose data fit a two component curve significantly better than a one component curve, indicating that there are two iso-forms of PDE in the plasma membrane of 3T3-L1 adipocytes, similar to swine adipocytes. The Km and Vmax values for this two component curve were Km1=0.12 microM, Vmax1=3.08 pmol min(-1) mg(-1) protein, and Km2=3.67 microM, Vmax2=83.8 pmol min(-1) mg(-1) protein. Inhibitors of PDE1, PDE2 and PDE5 failed to inhibit PM-PDE, as observed in swine adipocyte plasma membranes. However, PDE4 inhibitors were three-fold more effective at inhibiting PDE in 3T3-L1 PM compared to swine adipocyte PM. One mM 1, 3-dipropyl-8-p-sulfophenylxanthine (DPSPX) inhibited PM-PDE by approximately 75% in both preparations. These data demonstrate that PM-PDE is distinct from microsomal membrane PDE and may be responsible for extracellular cAMP metabolism to AMP in 3T3-L1 adipocytes.
Zhang
Xiujun
X
Department of Animal and Nutritional Sciences, University of New Hampshire, 403 Kendall Hall, Durham, NH 03824, USA.
Carey
Gale B
GB
D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D041721:3T3-L1 Cells; D017667:Adipocytes / Q000201:enzymology*; D000818:Animals; D000242:Cyclic AMP / Q000494:pharmacology; D054677:Cyclic Nucleotide Phosphodiesterases, Type 1; D004305:Dose-Response Relationship, Drug; D007700:Kinetics; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051379:Mice; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D020033:Protein Isoforms / Q000037:antagonists & inhibitors / Q000378:metabolism; D015363:Quinolones / Q000494:pharmacology
D008565:Membrane Proteins; D010726:Phosphodiesterase Inhibitors; D020033:Protein Isoforms; D015363:Quinolones; C021294:cilostamide; D000242:Cyclic AMP; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D054677:Cyclic Nucleotide Phosphodiesterases, Type 1