The Metabolism of Citrate-C14 in Normal and in Fluoroinhibitor-Poisoned Rats*

Abstract

A cyclical exchange of citrate has been shown to exist in the intact organism, with the blood plasma serving as the transporting medium. In vivo studies have indicated a considerable uptake of citrate by the liver (1) and kidney (2), and a release of citrate into the blood by the peripheral tissues (1, 3) and intestine (4,5). Interest in the blood citrate flux has centered primarily around the importance of this intermediate in carbohydrate metabolism, in its relationship to calcium mobilization and deposition in bone (6), and in its role as a chelating agent for calcium in the urine (7, 8).The studies reported here have been concerned with an attempt to elucidate the metabolic fate of the blood citrate in rats with the aid of radioactive citrate. The data, obtained by following the pattern of respiratory C140, and the urinary excretion of radioactive carbon after the intravenous administration of sodium citrate-1,5-C'4, indicated a rapid turnover of the blood citrate. Three hours after injection of the labeled citrate, approximately 90 per cent of the radioactivity was recovered in the respiratory CO., and in the urinary citrate. Evidence for the utilization of blood citrate as a precursor of tissue components was provided by assaying the C14 content of protein and lipid of kidneys and liver after intravenous citrate-C14 administration.In an effort to define further the metabolic fate of the blood citrate, inhibitors of citrate oxidation were administered prior to citrate-C14 administration. Peters (9) Fed, female Sprague-Dawley rats, weighing between 185 and 235 g, were used. Citric acid-1,5-C"' was obtained from Tracerlab, Inc., and sodium acetate--C1'4 from Volk Radiochemical Co. Sodium fluoroacetate 1 was purchased from Bios Laboratories, Inc. Barium fluorocitrate, synthesized by the method of Rivett (10), was obtained from the Sonbert Chemical Co., Brooklyn, N. Y.; this preparation contained less than 1 per cent barium citrate by specific analysis. Before use, the barium salt was dissolved in dilute HCl, the barium was precipitated with saturated sodium sulfate and the supernatant fluid was neutralized with dilute NaOH. Solutions of the fluoroinhibitors were freshly prepared prior to use.In the in vivo experiments, sodium citrate-1,5-C"4 in isotonic saline was given intravenously in a volume of 0.3 ml. Collection of respiratory CO2 was accomplished by drawing CO2-free air through a glass metabolism jar containing the rat, and the respiratory CO2 was trapped in NaOH. Barium carbonate precipitates were prepared from aliquots of the NaOH solutions. After the BaCO3 was washed with water and then alcohol, it was plated and counted in a Nuclear-Chicago gas-flow counter with a Micromil window. Urine collections were made up to standard volume, and aliquots were taken for direct plating and for conversion of the carboxyl groups of the urinary citrate to CO2 with ceric sulfate (11); the CO2

Effect of Acute Metabolic Acidosis and Alkalosis on Acetate and Citrate Metabolism in the Rat*

“…The hypothesis that citrate metabolism in the renal tubular cell is closely related to urinary citrate excretion is particularly attractive because of the known avidity of the kidney for the blood citrate (19,20) and because of the demonstrated rapid metabolism of citrate by kidney tissue (13). The present experiments conclusively demonstrate an effect of acidosis and alkalosis on renal clearance of plasma citrate.…”

“…Although hypercitraturia has been shown to occur in association with an increased citrate concentration in kidney tissue in such diverse conditions as alkalosis (22), after sodium malate administration (23), and in fluorocitrate-poisoned rats (13), the increased urinary excretion need not be a consequence of the increased tissue concentration (13,24). The data obtained in the present study suggest that tubular synthesis of citrate is not a major determinant of citrate excretion in the urine.…”

“…The second possibility, diminished metabolism of blood citrate in the alkalotic rat, is supported by experiments with citrate-C14. In the normal animal, approximately 55 per cent of an administered dose of citrate-C14 is found in the respiratory CO2 in 3 hours (13). In the alkalotic animal, only 35 per cent at most could be metabolized, since 65 per cent of the administered citrate-C14 is excreted in the urine.…”

“…The radioactivity of the urine and of the urinary citrate was determined by methods previously described with appropriate corrections for self-absorption and conversion of values to infinite thickness (13). Quantitative analysis of blood and urinary citrate was performed as previously described, by the technique of Elliott (14).…”

“…It has previously been shown in this laboratory that the urinary citrate is at least in part derived from the blood citrate, since an appreciable portion of intravenously administered citrate-C14 appears unchanged in the urine (13). In the present investigations, a similar experimental approach employing radioactive tracer substances was used to study the mechanism by which acute alterations of acid-base balance influence urinary citrate excretion.…”