Hepatic calcium efflux during cytochrome P-450-dependent drug oxidations at the endoplasmic reticulum in intact liver.

During metabolism of (type I) drugs by cytochrome P-450-dependent monooxygenase of the endoplasmic reticulum, the NADPH/NADP+ ratio in rat liver selectively decreases to approximately one-half of the control values, whereas the NADH/NAD+ ratio remains practically unaffected [Sies, H. & Brauser, B. (1970) Eur. J. Biochem. 15, 521-540]. In view of the observations with isolated mitochondria [Lehninger, A. L., Vercesi, A. & Bababunmi, E. A. (1978) Proc. Natl. Acad. Sci. USA 75, 1690-1694] of stimulated Ca2+ efflux upon nicotinamide nucleotide oxidation, the selective oxidation of NADPH in cytosol and mitochondria during drug oxidations was considered a useful experimental tool for the determination of whether the oxidation of NADPH or of NADH is responsible for Ca2+ efflux. With perfused livers from phenobarbital-treated rats, Ca2+ efflux was demonstrated, amounting to 8 nmol/min per gram of liver (wet weight), with aminopyrine, ethylmorphine, or hexobarbital as drug substrates. Drug-associated Ca2+ release was diminished when the inhibitor metyrapone was also present, or when drug oxidation was suppressed during N2 anoxia or in the presence of antimycin A in livers from fasted rats. Ca2+ efflux was elicited also by infusion of the thiol oxidant diamide, and by t-butyl hydroperoxide. However whereas Ca2+ efflux elicited by these compounds was restricted upon addition of the thiol dithioerythritol, there was little, if any, sensitivity of the drug-associated Ca2+ efflux to the thiol. Further mitochondrial oxidation of NADPH by addition of ammonium chloride had no effect on drug-associated Ca2+ efflux. Prior addition of the alpha-agonist phenylephrine suppressed the Ca2+ release by drug addition. While the molecular mechanism involved in Ca2+ efflux from liver mitochondria and from hepatocytes as well as the regulatory significance are not yet known, it is concluded from the present experiments that in case of nicotinamide nucleotide-linked Ca2+ efflux the oxidation of NADPH may suffice, with oxidation of NADH not being a requirement.