The influence of oxidative stress on various inotropic responses in isolated rat left atria.
The effects of free radicals, generated by electrolysis of a physiological salt solution, on various inotropic responses to drugs in isolated rat left atria were studied. Evidence for the generation of hydroxyl radicals was obtained from an appropriate fluorimetric assay. The amount of free radicals produced by electrolysis of the medium proved current-dependent. Exposure of isolated rat left atria to the medium which had been subjected to electrolysis caused a current-dependent decrease in contractile force. Oxidative stress, as a result of the electrolysis of the medium, caused altered inotropic responses to extra cellular Ca2+ (pD2 control group: 2.62 +/- 0.06 vs. 2.44 +/- 0.07 electrolysis group), sodium withdrawal (rise in contractile force control group: 1.73 +/- 0.19 mN vs. 0.48 +/- 0.21 mN electrolysis group) and lowering of stimulation frequency. The response to isoprenaline was diminished in atria subjected to oxidative stress and led to a rightward shift of the concentration response curves (pD2 control group: 7.56 +/- 0.10 vs. 6.77 +/- 0.11 electrolysis group). In addition, the inotropic responses to forskolin (pD2 control group: 6.17 +/- 0.12 vs. < 4.5 electrolysis group) and dibutyryl cAMP (rise in contractile force caused by 1 x 10(-5) M db-cAMP in control group: 2.15 +/- 0.01 mN vs. 1.21 +/- 0.10 mN electrolysis group) proved blunted as well. Measurement of the adenylyl cyclase activity revealed that free radicals attenuated the basal (by 11.1%) and forskolin stimulated (155.0 +/- 5.1 vs. 48.0 +/- 1.8 pmol cAMP/mg prot./min for control and electrolysis group respectively) activity of the adenylyl cyclase. DMSO, a well known hydroxyl radical scavenger, was able to abolish the free radical-induced decrease in the response to isoprenaline. Surprisingly, addition of alpha-adrenoceptor agonists to atria subjected to electrolysis-generated free radicals led to a rapid decrease in contractile force. DMSO was unable to counteract the negative intropic effect of methoxamine in atria subjected to oxidative stress. This negative inotropic response to alpha-adrenoceptor agonists in atria subjected to electrolysed medium is unlikely to be the direct result of phospholipase C or protein kinase C activation. Angiotensin II (which stimulates PLC, as well) did not reduce contractile force and chelerythrine (a PKC inhibitor) was unable to counteract the negative inotropic effect of the adrenoceptor agonists. In addition, the negative inotropic effect of methoxamine proved insensitive to 10(-6) M phentolamine and 10(-5) M doxazosin, which indicates an alpha-adrenoceptor independent mechanism. From this study we conclude that free radicals alter responses to various inotropic stimuli. These alterations may be the result of injured contractile elements, transporter molecules and molecules involved in signal transduction. Addition of alpha-adrenoceptor agonists after oxidative stress leads to a alpha-adrenoceptor. PLC and PKC independent decrease in contractile force.
Peters
S L
SL
University of Amsterdam, Department of Pharmacotherapy, The Netherlands.
Pfaffendorf
M
M
van Zwieten
P A
PA
D000262:Adenylyl Cyclases / Q000378:metabolism; D000316:Adrenergic alpha-Agonists / Q000494:pharmacology; D000818:Animals; D016279:Atrial Function, Left / Q000187:drug effects / Q000502:physiology*; D003994:Bucladesine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002316:Cardiotonic Agents / Q000494:pharmacology; D004572:Electrolysis; D005609:Free Radicals; D006325:Heart Atria / Q000201:enzymology; D066298:In Vitro Techniques; D007545:Isoproterenol / Q000494:pharmacology; D008297:Male; D009200:Myocardial Contraction / Q000187:drug effects / Q000502:physiology*; D018384:Oxidative Stress / Q000502:physiology*; D051381:Rats; D017208:Rats, Wistar; D015398:Signal Transduction / Q000187:drug effects
D000316:Adrenergic alpha-Agonists; D002316:Cardiotonic Agents; D005609:Free Radicals; D003994:Bucladesine; D000262:Adenylyl Cyclases; D007545:Isoproterenol; D002118:Calcium