Stable GDP analog-induced inactivation of G(i) proteins promotes cardiac adenylyl cyclase inhibition by guanosine 5'-(beta gamma-imino)triphosphate and muscarinic acetylcholine receptor.
Low concentrations of GDP and its stable analog guanosine 5'-O-(2-thio)diphosphate (GDP beta S) have been shown to stimulate adenylyl cyclase activity in canine cardiac sarcolemmal membranes independent from a phosphate transfer reaction. The mechanism of this stimulation was further examined. The stable GTP analog guanosine 5'-(beta gamma-imino)triphosphate (Gpp(NH)p) increased basal adenylyl cyclase activity and inhibited forskolin-stimulated activity with EC50 (half-maximal effective concentration) values of 0.7 mumol/l and 10 nmol/l, respectively. In the presence of GDP beta S (5 mumol/l), which increased basal activity by about 150%, addition of Gpp(NH)p inhibited adenylyl cyclase activity by up to 50% with an EC50 value of 40 nmol/l. Activation of cardiac muscarinic acetylcholine receptors by carbachol amplified this Gpp(NH)p-induced inhibition of GDP beta S-stimulated adenylyl cyclase activity. The stimulatory effect of GDP beta S and the inhibitory effect of Gpp(NH)p on GDP beta S-stimulated adenylyl cyclase activity were both attenuated by increasing the Mg2+ concentration or substituting Mn2+ for Mg2+ in the assay. Furthermore, both effects were strongly reduced or abolished upon pretreatment of the sarcolemmal membranes with a low concentration of the SH reagent N-ethylmaleimide (10 mumol/l). These results suggest that the stimulatory effect of GDP beta S on basal adenylyl cyclase activity in canine cardiac sarcolemmal membranes is caused by inactivation of G(i) proteins, which are then rendered susceptible to activation by Gpp(NH)p and inhibitory receptors.
Piacentini
L
L
Universität Heidelberg, Germany.
Mura
R
R
Jakobs
K H
KH
Niroomand
F
F
D000067956:Adenylyl Cyclase Inhibitors; D000818:Animals; D002217:Carbachol / Q000494:pharmacology; D004285:Dogs; D005033:Ethylmaleimide / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000037:antagonists & inhibitors* / Q000502:physiology; D006153:Guanosine Diphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology*; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D009206:Myocardium / Q000201:enzymology*; D011976:Receptors, Muscarinic / Q000187:drug effects / Q000502:physiology*; D012508:Sarcolemma / Q000201:enzymology; D013873:Thionucleotides / Q000494:pharmacology*
D000067956:Adenylyl Cyclase Inhibitors; D011976:Receptors, Muscarinic; D013873:Thionucleotides; D006153:Guanosine Diphosphate; D006165:Guanylyl Imidodiphosphate; D008345:Manganese; C023427:guanosine 5'-O-(2-thiodiphosphate); D002217:Carbachol; D019204:GTP-Binding Proteins; D008274:Magnesium; D005033:Ethylmaleimide