Effect of mercurials on lipid peroxidation in rabbit renal cortical mitochondria.

It is well known that mercury is ac cumulated in the kidney and causes renal failure. Mercury has been a subject of extensive study, but a definite mechanism for its biochemical toxicity remains to be elucidated. In a previous paper (1), we reported that mersalyl (mercurial diuretic) had a more powerful stimulatory effect on the lipid peroxidation in renal cortical mito chondria than any other diuretics. In the present study, mercuric chloride, mersalyl and methyl mercury chloride were compared for their ability to stimulate mitochondrial lipid peroxidation in vitro. Male rabbits fed on standard laboratory chow (Oriental Yeast, Co., Tokyo) ad libitum, weighing 2-2.5 kg, were used for the experiment. The kidneys were removed from anesthetized (sodium pentobarbital, 30 mg/kg i.v.) rabbits and rapidly chilled in ice-cold saline. The cortex was homogenized in freshly prepared 0.15 M KCI/0.02 M Tris-HCI buffer (pH 7.4) to make a 10% (w/v) homogenate, and then mitochondria were prepared according to the method of Hoge boom (2). Mitochondria (ca. 5 mg protein) were incubated in 5 ml of the Tris-HCI buffer at 37'C for 60 min. Mercuric chloride (0.1 mM), mersalyl (0.5 mM) and methyl mercury chloride (1 mM) were added to the medium. In preliminary experiments, we found that the maximally effective stimulatory concen tration of mercuric chloride, mersalyl and methyl mercury chloride was 0.1 mM, 0.5 mM and 1 mM, respectively (data not shown). The reaction was stopped after 10, 20, 30 and 60 min by the addition of 1.5 ml of 40% trichloroacetic acid, and the lipid peroxides were assayed by the thiobarbituric acid reaction, as described previously (3). Malondialdehyde was expressed as the thio barbituric acid values (absorbance at 532 nm/mg protein). Protein was determined by the method of Lowry et al. with bovine serum albumin as a standard (4). The values were expressed as the mean± standard error (S.E.). Student's t-test was employed to evalute the differences among the mean values. Figure 1 shows the time course of the lipid peroxidation of rabbit renal cortical mitochondria during the incubation with or without mercurials. The lipid peroxidation in the presence of mercuric chloride (0.1 mM) reached the maximum in about 10 min (2.6 fold), and this was maintained until the end of the 60 min incubation. Mersalyl (0.5 mM) and methyl mercury chloride (1 mM) stimu lated the lipid peroxidation at a low level as compared with mercuric chloride during the …