guanylyl-imidodiphosphate and Myocardial-Ischemia

To examine responses of the beta-adrenoceptor guanine nucleotide protein (G protein)/adenylyl cyclase complex to acute myocardial ischemia, we measured adenylyl cyclase activity stimulated at the beta-adrenoceptor and postreceptor levels and compared crude homogenates prepared from ischemic and nonischemic rabbit myocardium obtained after 30 minutes of coronary artery occlusion. Basal adenylyl cyclase activity was unchanged, but enzyme activity stimulated by the guanosine triphosphate analog guanyl-5'-imidodiphosphate (GppNHp) at 10 mumol/L was depressed 63% by ischemia (n = 16, p = 0.001). In contrast, adenylyl cyclase activity stimulated by 1 mumol/L (-)-isoproterenol in the presence of 10 mumol/L GppNHp was not significantly reduced (n = 10), a finding that indicates relative preservation of beta-adrenoceptor-mediated adenylyl cyclase activity in ischemia. The ratio of (-)-isoproterenol-stimulated to GppNHp-stimulated adenylyl cyclase activity increased fourfold in ischemic myocardium (n = 6, p = 0.001), consistent with more efficient beta-adrenergic signal transduction via less functional stimulatory G protein (Gs). These data could not be explained by augmented beta-adrenoceptor density or agonist affinity or by a reduction in inhibitory G protein-mediated inhibition of adenylyl cyclase. Forskolin (1 mmol/L) and Mn2+ (1 mmol/L), agents that directly stimulate the catalytic subunit of adenylyl cyclase, each increased enzyme activity significantly more in ischemic than in nonischemic myocardium. We conclude that preservation of (-)-isoproterenol-mediated adenylyl cyclase activity during acute myocardial ischemia in the rabbit results at least in part from enhanced function of the catalytic subunit of adenylyl cyclase.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenylyl Cyclases; Animals; Female; GTP-Binding Proteins; Guanylyl Imidodiphosphate; Isoproterenol; Male; Myocardial Ischemia; Myocardium; Rabbits; Receptors, Adrenergic, beta; Signal Transduction; Sodium-Potassium-Exchanging ATPase

It has been reported that the A1-adenosine receptor mediates the cardioprotective effect of ischemic preconditioning. This receptor couples inhibitory guanine nucleotide-binding protein (Gi) and inhibits adenylate cyclase activity. However, the role of adenylate cyclase in preconditioning is unknown.. We compared the effects of ischemia on the sarcolemmal beta-adrenergic receptor density (Bmax), the stimulatory guanine nucleotide-binding protein (Gs) activity as determined by reconstitution with S49 lymphoma cyc- membranes, and baseline and maximally stimulated adenylate cyclase activities (ACAs) in control and preconditioned rabbit hearts. The control population (n = 28) received 0, 10, 20, and 60 minutes of coronary occlusion (n = 6 to 8 per stage), and preconditioned rabbits (n = 24) received two cycles of alternating 5-minute occlusion and reperfusion before sustained ischemia (n = 6 per stage). In control hearts, occlusion induced rapid and progressive reductions in the Bmax, Gs, and ACAs after 10 to 60 minutes of ischemia. Preconditioning did not affect the reduction in Bmax, but it preserved reductions in Gs activity and ACAs after 10 to 20 but not 60 minutes of sustained ischemia. In another study, 18 rabbits were treated with pertussis toxin 48 hours before surgery to block Gi. During treatment, no significant difference was observed in the ischemia-induced reduction in ACAs in the ischemic region between control (n = 8) and preconditioned (n = 10) animals after 20 minutes of ischemia.. Preconditioning delays ischemia-induced reductions in beta-adrenergic signal transduction. Inhibition of ACA is not the target effect of the A1-adenosine receptor-Gi pathway responsible for the cardioprotective role of preconditioning.

Topics: Adenylate Cyclase Toxin; Adenylyl Cyclase Inhibitors; Adenylyl Cyclases; Animals; Colforsin; Female; GTP-Binding Proteins; Guanylyl Imidodiphosphate; Isoproterenol; Kinetics; Myocardial Ischemia; Myocardial Reperfusion Injury; Myocardium; Pertussis Toxin; Rabbits; Receptors, Adrenergic, beta; Receptors, Purinergic P1; Sarcolemma; Signal Transduction; Sodium Fluoride; Sodium-Potassium-Exchanging ATPase; Virulence Factors, Bordetella

The aim was to study the Gi protein mediated muscarinic signalling system in the myocardium of rats with chronic ischaemic heart failure.. Chronic ischaemic heart failure was induced by myocardial ischaemia (four weeks after coronary artery ligation) in rats. The densities and agonist affinities of muscarinic receptors, and the functional activity and concentration of Gi proteins were studied.. In failing hearts, the activity of adenylyl cyclase stimulated by guanyliminodiphosphate (Gpp(NH)p) was decreased by 46%. Stimulated activities of adenylyl cyclase by both sodium fluoride and forskolin, however, remained unchanged. Carbachol depressed forskolin stimulated adenylyl cyclase more in membranes from failing hearts than those from normal hearts. The functional level of Gs protein as measured by a reconstitution assay in sarcolemmal membrane did not differ between the two groups. Furthermore, muscarinic receptors exhibited superhigh and low affinities for agonist in failing hearts whereas those in control hearts displayed only high and low affinities. No significant difference in the peptide equivalent amount of membrane bound Gi protein was found in either group.. The experimental chronic failing heart due to myocardial ischaemia showed a depressed myocardial adenylyl cyclase signalling system. This may be due to the hypersensitivity of the Gi protein mediated muscarinic receptor-adenylyl cyclase system as shown by the increased inhibition of Gpp(NH)p mediated adenylyl cyclase, more potent inhibition of stimulated adenylyl cyclase by carbachol, and the superhigh affinity of the muscarinic receptors for carbachol.

Topics: Adenylyl Cyclases; Animals; Carbachol; Cell Membrane; Colforsin; Enzyme-Linked Immunosorbent Assay; GTP-Binding Proteins; Guanylyl Imidodiphosphate; Immunoblotting; Male; Myocardial Ischemia; Myocardium; Rats; Rats, Sprague-Dawley; Receptors, Muscarinic; Sarcolemma