Effect of calcium on the binding in vitro of [3H]cAMP to synaptosomal membranes from rat brain.
1. The binding of [3H]cAMP in vitro to synaptosomal membranes from rat brain was resolved in two components; one saturable at 20 nM cAMP with dissociation constant (KD) of 4.7 nM, and another nonsaturable within the 5-133 nM cAMP concentration range with an estimated KD value of 0.26 microM. 2. MgATP at concentration of 0.4 mM effected complete inhibition of the binding of [3H]cAMP to synaptosomal membranes throughout the used concentration range. This and the above finding indicate that the studied binding was focused on to the cAMP kinase on the membrane. 3. Calcium at concentrations of 0.1 and 10 mM stimulated a transient 20-30% increase of [3H]cAMP binding to the membranes which was influenced, as regards its time of appearance, by the concentration of cAMP. 4. The stimulation by calcium of the binding of [3H]cAMP to the membranes was inversely related to the phosphorylation of an Mr = 80,000 membrane protein, indicating stimulation of a negative effector function of cAMP--through cAMP-mediated phosphorylation--in the phosphorylation by calcium of this substrate. Moreover, the temporal displacement by cAMP of the peak of [3H]cAMP binding, produced similar temporal displacement of the inhibitory effect of cAMP on the Mr = 80,000 substrate phosphorylation. 5. These results suggest interaction in vitro of calcium and cAMP in modulation of the activity of cAMP kinase on the synaptosomal membranes.
Papaphilis
A D
AD
Department of Experimental Physiology, University of Athens, School of Medicine, Greece.
Kotsifaki
H E
HE
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D001921:Brain / Q000378:metabolism*; D002118:Calcium / Q000494:pharmacology*; D000242:Cyclic AMP / Q000378:metabolism*; D005260:Female; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains; D011953:Receptors, Cyclic AMP / Q000187:drug effects / Q000378:metabolism*; D013574:Synaptosomes / Q000378:metabolism*
D011953:Receptors, Cyclic AMP; D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D002118:Calcium