Effects of peroxide and superoxide on coronary artery: angiotensin response and sarcoplasmic reticulum ca2+ pump. Grover, A. K., S. E. Samson, V. P. Fomin & E. S. Werstiuk. Department of Biomedical Sciences, McMaster University, Hamilton, Ontario, Canada L8N3Z5
APStracts 2:0140C, 1995.
In isolated membranes, the SR (sarcoplasmic reticulum) Ca2+ pump is damaged by the reactive oxygen species (ROS) peroxide or superoxide. Hence, we hypothesize that ROS treatment of endothelium denuded pig coronary artery rings, or smooth muscle cells (SMC) cultured from them, diminishes their responses to angiotensin II (Ang II) which contracts the arterial smooth muscle by mobilizing intracellular Ca2+. We treated the artery rings or SMC with peroxide and superoxide, removed the added ROS and then assayed them for various activities. This experimental design eliminated direct ROS interference in the assay solutions and thus monitored irreversible effects. Treating the arteries with peroxide inhibited the Ang II induced contractions with K0.5 = 74+/-5 _M. 250 _M peroxide inhibited the contractions to Ang II and CPA (SR Ca2+-pump inhibitor cyclopiazonic acid) by 78.3+/-5.1 and 67.4+/-6.3%, respectively but did not significantly affect the contractions by 60 mM KCl. Treating SMC with peroxide inhibited the Ang II induced increase in [Ca2+i] with K0.5 = 24+/-3 _M for peroxide. 100 _M peroxide 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 acylphosphates. Treating SMC with 100 _M peroxide inhibited the acylphosphate levels by 36.3+/-3.2%. Peroxide (100 _M) treatment of SMC did not significantly alter their binding to 125I-sar1-ile8-angiotensin or inhibition of this binding by Ang II. Superoxide was generated using xanthine oxidase (X.O.)+0.3 mM xanthine. Xanthine+X.O. preferentially inhibited the arterial contractions to Ang II and CPA over the KCl contractions. However, a part of the effect of xanthine+X.O. was sensitive to catalase. In SMC, xanthine+X.O. inhibited the Ang II induced increase in [Ca2+i]. The X.O. concentration which inhibited about 80% of this Ang II response, inhibited less than 50% of the SR Ca2+-pump activity and had no effect on Ang II binding. Thus, ROS damaged the SR Ca2+-pump, thereby diminishing the SR Ca2+-pool and decreasing the smooth muscle response to Ang II. However, the inhibition of the Ang II induced [Ca2+i] increase was more severe than the damage to the Ca2+-pump. These results do not prove but are consistent with the concept that the SR Ca2+-pool is heterogenous in its sensitivity to ROS. 

Received 18 April 1994; accepted in final form 15 February 1995.
APS Manuscript Number C206-4.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 21 March 1995.