Decreased Tissue Distribution of L-Carnitine in Juvenile Visceral Steatosis Mice1

We kinetically analyzed the disposition of L-carnitine of juvenile visceral steatosis (JVS) mice compared with that of normal mice to elucidate the mechanism of the systemic L-carnitine deficiency of JVS mice. There were significant differences in the plasma concentration-time course of total radioactive carnitine (L-[H]carnitine, [acetyl-H]carnitine, and other [acyl-H]carnitines) between normal and JVS mice after a single i.v. or p.o. administration of L-[H]carnitine (250 ng/kg). The oral bioavailability of L-[H]carnitine in JVS mice (0.341) was about half of that in normal mice (0.675). The cumulative urinary excretion of total radioactive carnitine in JVS mice was about 10-fold more than that in normal mice, and the total clearance of unchanged L-[H]carnitine for JVS mice (6.70 ml/min) was significantly higher than that for normal mice (2.45 ml/min). The distribution volume at the steady state of unchanged L-[H]carnitine in JVS mice (1.10 liters/kg) was significantly smaller than that in normal mice (8.16 liters/kg). At 4 h after an i.v. administration, the apparent tissue-to-plasma concentration ratios of unchanged L-[H]carnitine for various tissues of JVS mice, except for brain, were about one half to one 20th of those in normal mice. In conclusion, this in vivo disposition kinetic study of L-carnitine supports the previous in vitro finding that the L-carnitine transporter is absent or functionally deficient in JVS mice because the renal reabsorption, the intestinal absorption, and the apparent tissue-to-plasma concentration ratios in JVS mice are significantly lower than those in normal mice. The JVS mouse should be a useful experimental model for studying carnitine deficiency diseases. It is well known that L-carnitine plays an important role in the transport of long-chain fatty acids across the mitochondrial inner membrane for b-oxidation and energy metabolism (Bremer, 1962; Fritz and Yue, 1964). Primary chronic Lcarnitine deficiency may cause encephalopathy through hypoketonemia and hyperammonemia, and cardiomyopathy or hepatic encephalopathy in combination with skeletal myopathy (Breningstall, 1990; Scholte et al., 1990). Furthermore, L-carnitine and acetylcarnitine may have a role in the clinical treatment of acute myocardial infarction (Iliceto et al., 1995) and Alzheimer’s disease (Parnetti, 1995). In 1988, we found that homozygous mutant mice, named juvenile visceral steatosis (JVS) mice, have systemic L-carnitine deficiency and develop fatty liver, hyperammonemia, and hypoglycemia (Koizumi et al., 1988). There have been some in vitro studies on the mechanism of L-carnitine deficiency in JVS mice. Horiuchi et al. (1994) examined kidney slice preparations and concluded that the primary deficiency of JVS mice is most probably related to a reduction in reabsorption of L-carnitine. Kuwajima et al. (1996) reported that at the endogenous L-carnitine concentration (50 mM), the L-carnitine transport activity of fibroblasts obtained from the heart of JVS mice was only 18% of that of normal mice. Our kinetic analysis using embryonic fibroblasts derived from normal and JVS mice suggested that JVS mice lack the high-affinity carnitine transporter, which has Na and temperature dependence (Hashimoto et al., 1998). These in vitro results suggest that the JVS mouse would be a useful animal model of primary L-carnitine transporter deficiency. Therefore, the next step should be to elucidate the precise mechanism of the systemic L-carnitine deficiency by means of kinetic studies of the tissue distribution and elimination of L-carnitine in the whole animal. Such studies should also help to confirm the relationship between the development of disease and the effect of L-carnitine and to establish the role of transcellular transport in determining the characteristic tissue distribution of L-carnitine in JVS mice. In the present study, we investigated the disposition kinetics of L-carnitine Received for publication July 27, 1998. 1 This work was supported in part by a grant in-aid for Scientific Research from the Ministry of Education, Science Sports and Culture, Japan. ABBREVIATIONS: JVS, juvenile visceral steatosis; TLC, thin-layer chromatography; Vdss, distribution volume at the steady state; CLtot, plasma total clearance; Kp, app, apparent tissue-to-plasma concentration ratios. 0022-3565/99/2891-0224$03.00/0 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 289, No. 1 Copyright © 1999 by The American Society for Pharmacology and Experimental Therapeutics Printed in U.S.A. JPET 289:224–230, 1999 224 at A PE T Jornals on July 7, 2017 jpet.asjournals.org D ow nladed from after i.v. and p.o. administration in normal and JVS mice and confirmed the existence of an L-carnitine transporter in vivo. Experimental Procedures Materials. L-[methyl-H]Carnitine hydrochloride (L-[H]carnitine, 79 Ci/mmol, radiochemical purity, 99.6%) was purchased from Amersham International Ltd. (Buckinghamshire, UK). Acetyl-L[methyl-H]carnitine hydrochloride ([acetyl-H]carnitine, 65 Ci/ mmol, radiochemical purity, 97.5%) was purchased from Moravek Biochemicals Inc. (Brea, CA). All other chemicals were of reagent grade and were used without further purification. Endogenous unchanged L-carnitine was determined by using a commercial kit (Free Carnitine Kit; Kainos Co., Tokyo, Japan). Animal Experiments. JVS mice were originally found among mice of the C3H.OH strain in our laboratory (Koizumi et al., 1988). The autosomal recessive mutant gene, jvs, was then backcrossed into C57BL/6 (CLEA, Tokyo, Japan), and this strain, C57BL/6-jvs, was