EFFECTS OF PEROXIDE AND SUPEROXIDE ON CORONARY-ARTERY - ANG-II RESPONSE AND SARCOPLASMIC-RETICULUM CA2+ PUMP

The sarcoplasmic reticulum (SR) Ca2+ pump in membranes isolated from arterial smooth muscle is damaged by reactive oxygen species (ROS). Because angiotensin II (ANG II) contracts arterial smooth muscle by mobilizing intracellular Ca2+ concentrations ([Ca2+](i)), we determined the effects of ROS pretreatment on ANG II-induced contractions in coronary artery rings and [Ca2+](i) transients in smooth muscle cells (SMC) cultured from them. This experimental design eliminates direct BOS interference in assay solutions, thus monitoring only the tissue damage. Pretreating the arteries with peroxide inhibited the ANG II contractions with the concentration for half-maximal activation (K-0.5) = 74 +/- 5 mu M Peroxide (250 mu M) inhibited the contractions to ANG II and cyclopiazonic acid (CPA, SR Ca2+-pump inhibitor) by 78.3 +/- 5.1 and 67.4 +/- 6.3%, respectively, but did not significantly affect the contractions by 60 mM KCl. Pretreating SMC with peroxide inhibited the ANG II-induced increase in [Ca2+](i) with K-0.5 = 24 +/- 3 mu M for peroxide. Peroxide (100 mu M) inhibited the increase in [Ca2+](i) in response to ANG II and CPA by 78.9 +/- 5.1 and 38.3 +/- 4.9%, respectively. The SR Ca2+-pump activity was also measured as the Ca2+-dependent formation of 115-kDa acylphosphate. Pretreating SMC with 100 mu M peroxide inhibited the acylphosphate levels by 36.3 +/- 3.2%. Peroxide (100 mu M) pretreatment of SMC did not significantly affect their ANG II binding. The results with superoxide were consistent with those obtained with peroxide, indicating that ROS damaged the SR Ca2+ pump, thereby diminishing the SR Ca2+ pool and decreasing the smooth muscle response to ANG II, but the inhibition of the ANG II-induced [Ca2+](i) increase was more severe than the damage to the Ca2+ pump. Therefore, we propose that the SR Ca2+ pool is heterogeneous in its sensitivity to ROS.