Expression of isoproterenol-induced activation of protein kinase A in coronary smooth muscle in dependence on homogenization buffer.
The activity of cyclic AMP-dependent protein kinase from the 30,000 x g supernatant fraction of isolated coronary arteries preincubated with either isoproterenol or L-thyroxine was concentration-dependently enhanced by the agonists if the quick-freezed vessels were homogenized at physiological ionic strength of 0.16 M in a buffer containing 20 mM MOPS or 20 mM imidazole. Substitution of phosphate for MOPS or imidazole in the homogenization medium completely abolished the expression of the effects of the hormones unless the ionic strength was increased by addition of 0.25 M NaCl. However, the increase in ionic strength markedly reduced the total activity of the enzyme measured in the presence of 2 x 10(-6) M cyclic AMP indicating a depressive effect of high salt solution. The use of imidazole as the buffer substance at physiological ionic strength likewise reduced the total enzyme activity, whereas the total activities measured after homogenization with MOPS or phosphate buffer at mu = 0.16 were not significantly different.
Institute of Pharmacology and Toxicology, Ernst-Moritz-Arndt-University, Greifswald.
K U
KU
Möritz
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R
Fermum
M
M
Giele
D000818:Animals; D002021:Buffers; D002417:Cattle; D003331:Coronary Vessels / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D066298:In Vitro Techniques; D007545:Isoproterenol / Q000494:pharmacology*; D009131:Muscle, Smooth, Vascular / Q000201:enzymology*; D009994:Osmolar Concentration; D011494:Protein Kinases / Q000378:metabolism*; D013974:Thyroxine / Q000494:pharmacology
D002021:Buffers; D011494:Protein Kinases; D007545:Isoproterenol; D013974:Thyroxine