Hepatic glutathione plays a key role in detoxification, protecting against toxins and carcinogens. A poorly understood factor in the homeostasis of hepatic glutathione is its efflux from intracellular to extracellular across both the sinusoidal and canalicular surfaces. This proposal is aimed at a better understanding of the mechanism of hepatic glutathione efflux and its importance. Using the in situ perfused rat liver model, the following studies are proposed: 1) To determine the contribution of efflux to the turnover of hepatic glutathione and the pool(s) from which biliary and perfusate glutathione arise; 2) To determine the relative maturation of sinusoidal and canalicular glutathione efflux with development from young to adult rats; 3) To determine the differential effect of inducing agents, such as phenobarbital, on sinusoidal and canalicular efflux and glutathione turnover; 4) To determine the relationship between the efflux of oxidized and reduced glutathione; 5) To determine the role of Alpha-glutamyltranspeptidase in hepatic glutathione efflux. The evidence available suggests that glutathione efflux into bile may be carrier-mediated whereas this is less certain for sinusoidal efflux. To further understand the mechanism of biliary efflux, the kinetics of glutathione binding to canalicular enriched liver plasma membrane will be determined. The binding interaction of reduced and oxidized glutathione and the influence of inducing agents and the maturational process on the binding kinetics will be determined. The work proposed will define the contribution of glutathione efflux to hepatic glutathione turnover, examine the relationship between canalicular and sinusoidal efflux through studies of the maturation and induction of efflux, assess the relationship between the efflux of reduced and oxidized glutathione, and through binding studies, identify the putative canalicular carrier(s) for reduced and oxidized glutathione.

Project Start
1982-02-01
Project End
1987-01-31
Budget Start
1985-02-01
Budget End
1986-01-31
Support Year
4
Fiscal Year
1985
Total Cost
Indirect Cost
Name
University of California Los Angeles
Department
Type
Schools of Medicine
DUNS #
119132785
City
Los Angeles
State
CA
Country
United States
Zip Code
90095
Takikawa, H; Kaplowitz, N (1988) Comparison of the binding sites of GSH S-transferases of the Ya- and Yb-subunit classes: effect of glutathione on the binding of bile acids. J Lipid Res 29:279-86
Fernandez-Checa, J C; Ren, C; Aw, T Y et al. (1988) Effect of membrane potential and cellular ATP on glutathione efflux from isolated rat hepatocytes. Am J Physiol 255:G403-8
Takikawa, H; Sugiyama, Y; Kaplowitz, N (1988) Comparison of the effects of bile acids and GSH on the fluorescence of bound 1-anilino-8-naphthalene sulfonate and the enzymatic activity of cationic and neutral human hepatic GSH S-transferases. Biochim Biophys Acta 954:37-43
Sugimoto, M; Takikawa, H; Stolz, A et al. (1987) Subunit heterogeneity of cationic human hepatic glutathione S-transferases. Pharmacology 35:65-78
Takikawa, H; Ookhtens, M; Stolz, A et al. (1987) Cyclical oxidation-reduction of the C3 position on bile acids catalyzed by 3 alpha-hydroxysteroid dehydrogenase. II. Studies in the prograde and retrograde single-pass, perfused rat liver and inhibition by indomethacin. J Clin Invest 80:861-6
Aw, T Y; Ookhtens, M; Kuhlenkamp, J F et al. (1987) Trans-stimulation and driving forces for GSH transport in sinusoidal membrane vesicles from rat liver. Biochem Biophys Res Commun 143:377-82
Ookhtens, M; Lyon, I; Kaplowitz, N (1987) Effect of age on the sinusoidal release of hepatic glutathione from the perfused rat liver. Biochem Pharmacol 36:4015-7
Takikawa, H; Stolz, A; Kaplowitz, N (1987) Cyclical oxidation-reduction of the C3 position on bile acids catalyzed by rat hepatic 3 alpha-hydroxysteroid dehydrogenase. I. Studies with the purified enzyme, isolated rat hepatocytes, and inhibition by indomethacin. J Clin Invest 80:852-60
Takikawa, H; Sugiyama, Y; Kaplowitz, N (1986) Binding of bile acids by glutathione S-transferases from rat liver. J Lipid Res 27:955-66
Aw, T Y; Ookhtens, M; Kaplowitz, N (1986) Mechanism of inhibition of glutathione efflux by methionine from isolated rat hepatocytes. Am J Physiol 251:G354-61

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