EFFECT OF ISCHEMIA AND REPERFUSION ON CARDIAC RYANODINE RECEPTORS - SARCOPLASMIC-RETICULUM CA2+ CHANNELS

We investigated the effect of ischemia and reperfusion on the cardiac ryanodine receptor, which corresponds to the sarcoplasmic reticulum Ca 2+ channel. Isolated working rat hearts were subjected to 10 to 30 min utes of global ischemia, followed or not by reperfusion. Ischemia prod uced significant reduction in the density of high-affinity H-3-ryanodi ne binding sites, determined either in whole-heart homogenate (B-max 2 20+/-22, 203+/-12, and 228+/-14 fmol/mg protein after 10, 20, and 30 m inutes of ischemia versus 298+/-18 fmol/mg protein in the control cond ition; P<.01) or in a fraction enriched in sarcoplasmic reticulum (B-m ax, 1.08+/-0.15 pmol/mg protein after 20 minutes of ischemia versus 1. 69+/-0.08 pmol/mg protein in the control condition; P<.01). The K-d (1 .5+/-0.1 nmol/L) and the Ca2+ dependence of high-affinity H-3-ryanodin e binding were not affected by ischemia. The density of low-affinity H -3-ryanodine binding sites was also reduced after 20 minutes of ischem ia (14.0+/-2.3 versus 34.0+/-8.2 pmol/mg protein in the sarcoplasmic r eticulum fraction, P<.05), without significant changes in K-d (4.7+/-1 .2 versus 2.4+/-1.0 mu mol/L). All these changes persisted after 20 mi nutes of reperfusion. Analysis of tissue fractions showed that 55% of the ryanodine binding sites were retained in the pellet of a low-speed centrifugation (''nuclear pellet'') and that the effects of ischemia concerned only the receptors released in the supernatant (''postnuclea r supernatant''). In parallel experiments, we evaluated the effect of ryanodine on oxalate-supported Ca2+ uptake, which represents sarcoplas mic reticulum Ca2+ uptake. As expected, we found that high concentrati ons of ryanodine stimulated Ca2+ uptake, owing to channel blockade. Th e response to 900 mu mol/L ryanodine was slightly reduced in crude hom ogenate and significantly reduced in postnuclear supernatant obtained from ischemic hearts. In conclusion, the number of ryanodine receptors is reduced after ischemia; this effect concerns a subpopulation of th e receptors, persists after reperfusion, and might contribute to modif y sarcoplasmic reticulum function.