Stimulatory effect of vanadate on 3',5'-cyclic guanosine monophosphate-inhibited low Michaelis-Menten constant 3',5'-cyclic adenosine monophosphate phosphodiesterase activity in isolated rat fat pads.
When isolated rat fat pads were incubated with vanadate, the low Michaelis-Menten constant (Km) cAMP phosphodiesterase (PDE) activity in the microsomal fraction was increased in a time- and dose-dependent manner with vanadate. 3',5'-Cyclic GMP inhibited the vanadate-stimulated PDE activity with similar profile to the insulin-stimulated one. The stimulatory effect of vanadate was inhibited by inhibitors of tyrosine kinases such as amiloride, biochanin A, and genistein to various extents. Vanadate and insulin both showed the full effect in the absence of either K+, N+, or Ca2+ in the medium, while preincubation of the fat pads with a chelator of intracellular Ca2+ inhibited the vanadate action in a dose-dependent manner. The insulin action was not inhibited by it at tested concentrations. These results suggest that the vanadate action, in contrast to the insulin one, is dependent on the intracellular level of Ca2+. Preincubation of the fat pads with inhibitors of protein kinase C such as 1-(5-isoquinoline sulfonyl)-2-methylpiperazine (H-7) and staurosporine inhibited, in part, the vanadate action but did not inhibit the insulin one. Furthermore, vanadate increased the protein kinase C activity in fat pads but insulin did not increase. H-7 and amiloride showed a significant inhibition of stimulation of protein kinase C activity by vanadate. These results suggest that vanadate stimulates, in part, the 3',5'-cyclic GMP-inhibited low Km cAMP PDE activity in the microsomal fraction of fat pads through the activation of tyrosine kinase and protein kinase C-mediated processes.
Ueki
H
H
Department of Biochemistry, Faculty of Pharmacy and Pharmaceutical Sciences, Fukuyama University, Hiroshima, Japan.
Okuhama
R
R
Sera
M
M
Inoue
T
T
Tominaga
N
N
Morita
T
T
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000378:metabolism*; D000273:Adipose Tissue / Q000201:enzymology*; D000470:Alkaloids / Q000494:pharmacology; D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D006152:Cyclic GMP / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D007328:Insulin / Q000494:pharmacology; D007546:Isoquinolines / Q000494:pharmacology; D007700:Kinetics; D008297:Male; D008861:Microsomes / Q000201:enzymology; D010879:Piperazines / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D011972:Receptor, Insulin / Q000187:drug effects / Q000502:physiology; D019311:Staurosporine; D014638:Vanadates / Q000008:administration & dosage / Q000494:pharmacology*
D000470:Alkaloids; D007328:Insulin; D007546:Isoquinolines; D010879:Piperazines; D014638:Vanadates; D000584:Amiloride; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin; D011493:Protein Kinase C; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D006152:Cyclic GMP; D019311:Staurosporine