HALOTHANE AND ISOFLURANE EFFECTS ON CA2+ FLUXES OF ISOLATED MYOCARDIAL SARCOPLASMIC-RETICULUM

To elucidate better the differential myocardial depressant actions of halogenated volatile anesthetics, anesthetic-induced changes in Ca2+ accumulation, release, and Ca-ATPase (Ca2+ pump) activity of isolated canine cardiac sarcoplasmic reticulum (SR) vesicles were examined. An initial crude microsomal fraction of homogenized canine ventricle was subfractionated on a discontinuous sucrose gradient after Ca2+ loading in the presence of phosphate. Junctional SR (JSR) enriched with terminal cisternae was identified by its content of an electrophoretically verified approximately 450-kDa protein, the Ca2+-release channel (CaRC). When the CaRC of JSR was blocked by 1-mu-M ruthenium red (RR), the rate of Ca2+ uptake increased 47% as measured spectrophotometrically using the Ca-sensitive dye anti-pyrylazo III. A second fraction was identified as primarily longitudinal SR (LSR) based on its trace content of 450-kDa protein and 11% increase of Ca2+ uptake with RR. Halothane (0.75-2.5%) or isoflurane (2.5-4%) decreased net Ca2+ accumulation rate by either LSR or JSR, and the decrease in uptake rate of JSR was only partially reversed by addition of 1-mu-M RR (27% increase for isoflurane, 7% increase for halothane). Both halothane and isoflurane increased JSR ATP consumption as measured by a coupled-enzyme assay. Ca-45(2+) efflux from passively loaded SR vesicles was then determined to verify that the decreased net uptake rate was due to enhanced Ca2+ efflux from vesicles. Both anesthetics increased passive Ca2+ efflux from SR vesicles in which the CaRC was blocked by 10-mu-M RR as well as those in which Ca2+ release via the CaRC was activated by 10-mu-M Ca2+. By enhancing nonspecific Ca2+ efflux, halothane and isoflurane decrease Ca2+ retention of isolated cardiac SR vesicles, an effect that may mask specific actions on Ca2+ flux through the CaRC.