title: Protein kinase-dependent oxidative regulation of the cardiac Na+-K+ pump: evidence from in vivo and in vitro modulation of cell signalling.
abstract: The widely reported stimulation of the cardiac Na(+)-K(+) pump by protein kinase A (PKA) should oppose other effects of PKA to increase contractility of the normal heart. It should also reduce harmful raised myocyte Na(+) levels in heart failure, yet blockade of the β1 adrenergic receptor (AR), coupled to PKA signalling, is beneficial. We treated rabbits with the β1 AR antagonist metoprolol to modulate PKA activity and studied cardiac myocytes ex vivo. Metoprolol increased electrogenic pump current (Ip) in voltage clamped myocytes and reduced glutathionylation of the β1 pump subunit, an oxidative modification causally related to pump inhibition. Activation of adenylyl cyclase with forskolin to enhance cAMP synthesis or inclusion of the catalytic subunit of PKA in patch pipette solutions abolished the increase in Ip in voltage clamped myocytes induced by treatment with metoprolol, supporting cAMP/PKA-mediated pump inhibition. Metoprolol reduced myocardial PKA and protein kinase C (PKC) activities, reduced coimmunoprecipitation of cytosolic p47(phox) and membranous p22(phox) NADPH oxidase subunits and reduced myocardial O2(•-)-sensitive dihydroethidium fluorescence. Treatment also enhanced coimmunoprecipitation of the β1 pump subunit with glutaredoxin 1 that catalyses de-glutathionylation. Since angiotensin II induces PKC-dependent activation of NADPH oxidase, we examined the effects of angiotensin-converting enzyme inhibition with captopril. This treatment had no effect on PKA activity but reduced the activity of PKC, reduced β1 subunit glutathionylation and increased Ip. The PKA-induced Na(+)-K(+) pump inhibition we report should act with other mechanisms that enhance contractility of the normal heart but accentuate the harmful effects of raised cytosolic Na(+) in the failing heart. This scheme is consistent with the efficacy of β1 AR blockade in the treatment of heart failure.
authors.affiliation: North Shore Heart Research Group, Kolling Institute, University of Sydney, Australia.
authors.forename: Keyvan Karimi
authors.initials: KK
authors.lastname: Galougahi
authors.forename: Chia-Chi
authors.initials: CC
authors.lastname: Liu
authors.forename: Alvaro
authors.initials: A
authors.lastname: Garcia
authors.forename: Natasha A S
authors.initials: NA
authors.lastname: Fry
authors.forename: Elisha J
authors.initials: EJ
authors.lastname: Hamilton
authors.forename: Helge H
authors.initials: HH
authors.lastname: Rasmussen
authors.forename: Gemma A
authors.initials: GA
authors.lastname: Figtree
mesh_terms: D000200:Action Potentials / Q000187:drug effects; D000262:Adenylyl Cyclases / Q000378:metabolism; D000319:Adrenergic beta-Antagonists / Q000494:pharmacology; D000806:Angiotensin-Converting Enzyme Inhibitors / Q000494:pharmacology; D000818:Animals; D002216:Captopril / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000819:agonists / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D054477:Glutaredoxins / Q000378:metabolism; D008297:Male; D008790:Metoprolol / Q000494:pharmacology; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D019255:NADPH Oxidases / Q000378:metabolism; D010084:Oxidation-Reduction; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D021122:Protein Subunits / Q000378:metabolism; D011817:Rabbits; D015398:Signal Transduction; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*
chemical_list: D000319:Adrenergic beta-Antagonists; D000806:Angiotensin-Converting Enzyme Inhibitors; D054477:Glutaredoxins; D047428:Protein Kinase Inhibitors; D021122:Protein Subunits; D005576:Colforsin; D002216:Captopril; D000242:Cyclic AMP; D019255:NADPH Oxidases; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D000262:Adenylyl Cyclases; D000254:Sodium-Potassium-Exchanging ATPase; D008790:Metoprolol