guanosine-triphosphate and carbonyl-cyanide-phenylhydrazone

ATP-sensitive K+ (KATP) channels are present at high density in membranes of cardiac cells, where they regulate cardiac function during metabolic impairment. The present study analyzes the effects of extracellular ATP (ATPc), a P2-purinergic agonist that can be released under various conditions in the myocardial cell bed, on KATP current (IK-ATP) in rat ventricular myocytes. Under the whole-cell patch-clamp configuration at a physiological level of intracellular ATP, applying ATPc in the micromolar range did not activate IK-ATP. However, dialyzing the cell with a low-ATP (100 mumol/L) pipette solution elicited a slowly, quasilinearly increasing IK-ATP that was markedly enhanced by applying ATPe in the presence of a Purinergic antagonist. The effect was reversible on washing out the agonist. The IK-ATP enhancement was inhibited by cholera toxin treatment of the myocytes, suggesting that a Gs protein was involved to mediate the effect. Experiments on excised patches allowed us to exclude a membrane-delimited G protein-dependent pathway. Rather, the results suggested that ATPe activates the adenylyl cyclase, since its inhibition by 2'-deoxyadenosine 3'-monophosphate and SQ-22536, which respectively interact with the purine and catalytic site of the cyclase, strongly reduced the ATPe-induced IK-ATP enhancement, whereas neither compound affected IK-ATP in inside-out patches. Inhibition of cAMP-dependent protein kinase by protein kinase inhibitor peptide 5-24 did not alter the purinergic effect. The findings suggests that ATPe triggers the activation of adenylyl cyclase, which causes a subsarcolemmal ATP depletion sufficient to enhance IK-ATP as it develops during low-ATP dialysis of rat ventricular myocytes.

Topics: Adenine; Adenosine Triphosphate; Adenylyl Cyclase Inhibitors; Animals; Benzopyrans; Carbonyl Cyanide m-Chlorophenyl Hydrazone; Colforsin; Cromakalim; Deoxyadenine Nucleotides; Dose-Response Relationship, Drug; Enzyme Inhibitors; Extracellular Space; GTP-Binding Proteins; Guanidines; Guanosine Triphosphate; Heart Ventricles; Male; Patch-Clamp Techniques; Peptide Fragments; Pinacidil; Potassium Channels; Pyrroles; Rats; Rats, Wistar; Uncoupling Agents