Orotic Excretion Research Articles

Regulation of the pyrimidine biosynthetic pathway by lysine acetylation of E. coli OPRTase.

The de novo pyrimidine biosynthesis pathway is an important route due to the relevance of its products, its implications in health and its conservation among organisms. Here, we investigated the regulation by lysine acetylation of this pathway. To this aim, intracellular and extracellular metabolites of the route were quantified, revealing a possible blockage of the pathway by acetylation of the OPRTase enzyme (orotate phosphoribosyltransferase). Chemical acetylation of OPRTase by acetyl-P involved a decrease in enzymatic activity. To test the effect of acetylation in this enzyme, K26 and K103 residues were selected to generate site-specific acetylated proteins. Several differences were observed in kinetic parameters, emphasizing that the kcat of these mutants showed a strong decrease of 300 and 150-fold for OPRTase-103AcK and 19 and 6.3-fold for OPRTase-26AcK, for forward and reverse reactions. In vivo studies suggested acetylation of this enzyme by a nonenzymatic acetyl-P-dependent mechanism and a reversion of this process by the CobB deacetylase. A complementation assay of a deficient strain in the pyrE gene with OPRTase-26AcK and OPRTase-103AcK was performed, and curli formation, stoichiometric parameters and orotate excretion were measured. Complementation with acetylated enzymes entailed a profile very similar to that of the ∆pyrE strain, especially in the case of complementation with OPRTase-103AcK. These results suggest regulation of the de novo pyrimidine biosynthesis pathway by lysine acetylation of OPRTase in Escherichia coli. This finding is of great relevance due to the essential role of this route and the OPRTase enzyme as a target for antimicrobial, antiviral and cancer treatments.

Carbamoylphosphate synthetase 1 (CPS1) deficiency: clinical, biochemical, and molecular characterization in Malaysian patients.

Carbamoyl phosphate synthetase 1 (CPS1) deficiency is a rare autosomal recessive disorder of ureagenesis presenting as life-threatening hyperammonemia. In this study, we present the main clinical features and biochemical and molecular data of six Malaysian patients with CPS1 deficiency. All the patients have neonatal-onset symptoms, initially diagnosed as infections before hyperammonemia was recognized. They have typical biochemical findings of hyperglutaminemia, hypocitrullinemia, and low to normal urinary excretion of orotate. One neonate succumbed to the first hyperammonemic decompensation. Five neonatal survivors received long-term treatment consisting of dietary protein restriction and ammonia-scavenging drugs. They have delayed neurocognitive development of varying severity. Genetic analysis revealed eight mutations in CPS1 gene, five of which were not previously reported. Five mutations were missense changes while another three were predicted to create premature stop codons. In silico analyses showed that these new mutations affected different CPS1 enzyme domains and were predicted to interrupt interactions at enzyme active sites, disturb local enzyme conformation, and destabilize assembly of intact enzyme complex. All mutations are private except one mutation; p.Ile1254Phe was found in three unrelated families. Identification of a recurrent p.Ile1254Phe mutation suggests the presence of a common and unique mutation in our population. Our study also expands the mutational spectrum of the CPS1 gene.

Effects of arginine treatment on nutrition, growth and urea cycle function in seven Japanese boys with late-onset ornithine transcarbamylase deficiency

The aim of this study was to investigate the effects of arginine on nutrition, growth and urea cycle function in boys with late-onset ornithine transcarbamylase deficiency (OTCD). Seven Japanese boys with late-onset OTCD enrolled in this study resumed arginine treatment after the cessation of this therapy for a few years. Clinical presentations such as vomiting and unconsciousness, plasma amino acids and urinary orotate excretion were followed chronologically to evaluate urea cycle function and protein synthesis with and without this therapy. In addition to height and body weight, blood levels of proteins, lipids, growth hormone (GH), insulin-like growth factor-I (IGF-I) and IGF-binding protein -3 (IGFBP-3) were monitored. The frequency of hyperammonemic attacks and urinary orotate excretion decreased significantly following the resumption of arginine treatment. Despite showing no marked change in body weight, height increased gradually. Extremely low plasma arginine increased to normal levels, while plasma glutamine and alanine levels decreased considerably. Except for a slight increase in high-density lipoprotein cholesterol level, blood levels of markers for nutrition did not change. In contrast, low serum IGF-I and IGFBP-3 levels increased to age-matched control levels, and normal urinary GH secretion became greater than the level observed in the controls. Arginine treatment is able to reduces attacks of hyperammonemia in boys with late-onset OTCD and to increase their growth.

Extremely Short Small Bowel Induces Focal Tubulointerstitial Fibrosis

ABSTRACTBackgroundArginine becomes an essential amino acid after massive resection of the small bowel as a result of decreased biosynthesis of citrulline in the remaining small bowel. It is also reported that nitric oxide (NO) is synthesized from l ‐arginine by NO synthase (NOS), and NO is involved in the regulation of blood flow in the kidney. The authors observed a patient with an extremely short small bowel, showing focal tubulointerstitial fibrosis. The experiment was designed to clarify whether massive small bowel resection (SBR) produces focal tubulointerstitial fibrosis in the kidney.MethodsAn experimental study was performed using 4‐week‐old rats with 90% proximal SBR either with or without arginine supplementation for 6 weeks after surgery.ResultsIn rats without arginine supplementation, low plasma levels of citrulline and arginine increased urinary excretion of orotate, and focal tubulointerstitial fibrosis was observed 6 weeks after 90% SBR. The data from plasma amino acid chromatography and increased excretion of urinary orotate suggested the presence of arginine deficiency. The kidney pathology was similar to that of our patient. Rats with arginine supplementation after 90% SBR and pair‐fed control rats without 90% SBR showed almost normal glomeruli and tubulointerstitium.ConclusionsExperimental study strongly suggests that arginine deficiency causes focal tubulointerstitial fibrosis in the kidney after massive SBR.

Extremely short small bowel induces focal tubulointerstitial fibrosis.

Arginine becomes an essential amino acid after massive resection of the small bowel as a result of decreased biosynthesis of citrulline in the remaining small bowel. It is also reported that nitric oxide (NO) is synthesized from l-arginine by NO synthase (NOS), and NO is involved in the regulation of blood flow in the kidney. The authors observed a patient with an extremely short small bowel, showing focal tubulointerstitial fibrosis. The experiment was designed to clarify whether massive small bowel resection (SBR) produces focal tubulointerstitial fibrosis in the kidney. An experimental study was performed using 4-week-old rats with 90% proximal SBR either with or without arginine supplementation for 6 weeks after surgery. In rats without arginine supplementation, low plasma levels of citrulline and arginine increased urinary excretion of orotate, and focal tubulointerstitial fibrosis was observed 6 weeks after 90% SBR. The data from plasma amino acid chromatography and increased excretion of urinary orotate suggested the presence of arginine deficiency. The kidney pathology was similar to that of our patient. Rats with arginine supplementation after 90% SBR and pair-fed control rats without 90% SBR showed almost normal glomeruli and tubulointerstitium. Experimental study strongly suggests that arginine deficiency causes focal tubulointerstitial fibrosis in the kidney after massive SBR.

Influence of dose and age on the response of the allopurinol test for ornithine carbamoyltransferase deficiency in control infants.

Measurement of urinary orotidine and orotic acid after an oral allopurinol challenge is an important diagnostic test for ornithine carbamoyltransferase deficiency that is sometimes used in infants (< 1 year of age), although there is little information on normal test results in this age group. We found higher orotidine excretion in normal infants than in older children given the test, whereas orotate excretion was similar in both groups. The increased orotidine excretion appears to be due to the use in the infants of higher allopurinol doses per kilogram of body weight than in the children. The normalized-dose dependency of the orotidine response extends even to adult age. Thus, dose-normalized responses should be used in the test and there is no need for careful age-matching of the controls.

The allopurinol load test lacks specificity for primary urea cycle defects but may indicate unrecognized mitochondrial disease.

Thirty-three children ranging from 2 weeks to 12 years of age were selected for allopurinol loading, 16 on the basis of an increased urinary ourotate excretion detected by routine organic acid analysis (group A), and 17 for clinical reasons suggesting a urea cycle defect (group B). The allopurinol load test proved positive in 13 of 16 patients from group A, mean peak orotate 64.0 mumol/mmol creatinine (upper limit of reference range, 13.2) and 11 of 17 patients from group B, mean peak orotate 41.0 mumol/mmol creatinine (upper limit of reference range, 13.2). Thorough investigation of these patients including urinary and plasma amino acid analysis and, in 17 cases, liver biopsy for histology and measurement of ornithine carbamyltransferase (OCT) and carbamyl-phosphate synthetase (CPS) activity failed to identify any evidence of a urea cycle disorder. However, muscle biopsies performed in 11 patients showed some evidence of mitochondrial disease in four cases, two defined on the basis of reduced respiratory chain enzyme activity and two on the basis of mtDNA abnormalities. These findings indicate that an increased excretion of orotate in sick children may not be uncommon and that a positive allopurinol load test result may not indicate a specific inherited urea cycle defect. In addition, these results raise the interesting possibility that defective ureagenesis may be a feature of mitochondrial disease in some individuals.

The effects of various inhibitors on the regulation of orotic acid excretion in sparse-fur mutant mice (spf/Y) deficient in ornithine transcarbamylase

Experiments were conducted to determine whether the excessive orotic aciduria, induced in sparse-fur male mice (spf/Y) deficient in ornithine transcarbamylase (OTC), may be regulated by some inhibitors, such as acivicin (0.014 mmol/100 g body weight, i.p.), N- (phosphonoacetyl)- l-aspartate (PALA, 2.5 mg/100 g body weight, i.p.), adenine (3 g/kg diet) and cycloheximide (0.35 mmol/kg body weight, i.p.). We also administered ornithine (1 mmol/100 g body weight, i.p.), a substrate of the urea cycle, to alleviate the metabolic deficiency of arginine in spf/Y mice which may also be responsible for excessive orotic aciduria. The orotic aciduria remained insensitive to acivicin, indicating mitochondria as the source of carbamyl phosphate. However, orotate excretion was significantly decreased by PALA ( P < 0.01), due to its effect on the aspartate transcarbamylase activity. The ingestion of adenine resulted in an increase ( P < 0.05) of urinary orotate, suggesting the blockage of the utilization of orotate for nucleotide biosynthesis. Ornithine administration led to a reduction ( P < 0.01) of the excretion of orotate induced by the OTC deficiency in these mice, indicating that one of the regulatory steps in its synthesis may be the availability of ornithine. There were no changes in urinary orotate excretion in spf/Y mice when treated with cycloheximide. On the other hand, pretreatment with cycloheximide in an artificial model of OTC deficiency (Swiss-ICR normal mice on an arginine-deficient diet treated thereafter with norvaline, an inhibitor of OTC), caused a significant decrease in urinary orotate. These results suggest that spf/Y mice are unique in that the increased synthesis of orotate is not sensitive to cycloheximide. Perhaps this may reflect an adaptive phenomenon developed by the mutant mice to handle excess mitochondrial carbamyl phosphate and orotic acid.

Regulation of orotic acid biosynthesis and excretion induced by oral glutamine administration in mice.

Glutamine, the most abundant amino acid in blood and tissues, is degraded by the renal and splanchnic tissues, especially the small intestinal mucosa. Due to the activity of glutaminase, it may be broken down in these tissues and contribute to ammoniagenicity. Glutamine, either directly or through ammonia production, may act as a nitrogenous source for pyrimidine biosynthesis. We have evaluated the effect of glutamine on orotate metabolism in mice, by gavaging (ig) L-glutamine, 1.0 to 4.0 mmol/100 g of body wt/day, during 6 weeks of experimentation. Glutamine at doses of 2.5 to 4.0 mmol/100 g of body wt caused a significant increase in plasma ammonia and urinary orotate. The regulation of the orotic acid biosynthesis and excretion was studied by testing the effects of various inhibitors in mice force-fed with glutamine (4 mmol/100 g of body wt, ig). The orotic aciduria was insensitive to acivicin (1 and 5 mg/100 g of body wt, ip), a specific inhibitor of the cytoplasmic carbamyl phosphate synthetase-II, thus pointing toward the mitochondrion as the principal source of carbamyl phosphate. Cycloheximide (15 and 100 mg/kg of body wt, ip) caused a significant decrease in urinary orotate indicating that the induction of orotate synthesis by glutamine may be associated with the translation of a specific protein. However, orotate excretion was significantly decreased by N-(phosphonoacetyl)-L-aspartate (PALA) (5 mg/100 g of body wt, ip) due to its inhibitory effect on the aspartate transcarbamylase activity. There was a significant increase of urinary orotate following ingestion of adenine supplemented diets (0.1% and 0.2%), suggesting the blockage of the utilization of orotate for nucleotide biosynthesis by glutamine. Since orotate synthesis may also be influenced by ornithine metabolism, we evaluated the effect of glutamine administration on various ornithine-metabolizing enzymes. There was a decrease in hepatic ornithine decarboxylase activity with no change in hepatic ornithine aminotransferase activity following the administration of glutamine. This observation indicates that an increased metabolic utilization of ornithine is not responsible for the increase in orotate excretion, which may be caused principally through an effect of glutamine on mitochondrial carbamyl phosphate synthesis.

The metabolism and urinary excretion of orotic acid in hyperargininaemic sheep.

The synthesis, blood plasma turnover and urinary excretion of the orotic acid in normo- and hyperargininaemic sheep was investigated. The whole-body orotate formation was evaluated indirectly by the measurement of urinary orotate excretion after blockage of pyrimidine pathway with 6-azauridine (4 hour i.v. infusion of 0.2 mg.kg-1.min-1). Simultaneous infusion of L-arginine (2.5 mumols.kg-1.min-1) significantly elevated the blood plasma arginine, ornithine and urea level, however, it did not significantly influence urinary orotate excretion. In normoargininaemia blood plasma turnover of exogenous orotic acid amounted to 4.9 min and 67% of this compound was eliminated through the kidneys. The renal clearance of orotic acid amounted to 21.7 ml.min-1.kg-0.75. Hyperargininaemia elevated blood plasma turnover to 8.2 min, and diminished the renal clearance of this metabolite to 13.7 ml.min-1.kg-0.75. These results indicate that hyperargininaemia and hyperornithinaemia do not change the whole body synthesis of orotic acid in sheep but they can affect renal excretion of this metabolite, particularly at the rate of tubular secretion close to saturation.

Arginine-Deficient Diets Alter Plasma and Tissue Amino Acids in Young and Aged Rats

Blood and urine metabolites were measured in two experiments for young (2-mo-old) and aged (20-mo-old) male Sprague-Dawley rats fed arginine-devoid diets made isonitrogenous to a control 1.12% arginine diet by adding alanine or glycine. Diet, fed for 7 or 13 d, had little effect on urinary or plasma ammonia and urea. Urinary orotate excretion was more than 40-fold higher in rats fed the arginine-deficient diets (P less than 0.01) in both experiments. Source of nonessential N (alanine or glycine) in the arginine-deficient diets did not alter orotic acid excretion or plasma or urine ammonia or urea. Changes in plasma arginine, alanine and glycine concentrations reflected the levels of these amino acids in the diet. Tissue ornithine levels reflected dietary arginine level, but tissue citrulline was unaffected by dietary arginine. Glutamate and glutamine were greater in the plasma and liver of rats fed arginine-deficient diets. Plasma concentrations of glutamate and glutamine were positively correlated with urinary orotic acid excretion (P less than 0.05) and ornithine and arginine were negatively correlated with orotic acid excretion (P less than 0.01). Increased tissue glutamine may be related to the greater orotate excretion in rats fed arginine-devoid diets. The metabolic responses to dietary arginine deficiency were similar in young and aged rats. In general, concentrations of amino acids in plasma, liver and spleen were higher in aged rats.

Nitrogen Metabolism in Sickle Cell Anemia: Free Amino Acids in Plasma and Urine

Twenty-four hour urinary levels and fasting plasma concentrations of free amino acids (AA) were evaluated in adult sickle cell anemia (HbSS) and age-matched HbAA subjects with comparable daily energy (E) and protein (N) intake. HbSS elicited significant reduction in the sum of plasma indispensable (EAA) with no change in the dispensable (NAA) amino acids, resulting in a prominent (P less than 0.01) reduction in the EAA/NAA ratio from 71% to 55%. Arg (-38%), Leu (-32%), Val (-28%) and His (-32%) were among the AA most severely affected. Despite a twofold increase in 24-hour urine volume in HbSS compared with HbAA subjects, total urinary losses of EAA and NAA were markedly reduced in the former group, with Arg (-53%) and Gly (-56%) levels most prominently affected. Compared with HbAA controls, HbSS subjects showed a three-fourfold increase in 24-hour urinary orotate excretion that had no relationship to amount of N intake. The results indicated that adult subjects with HbSS, who consumed adequate N and E as per the RDA for healthy individuals, behaved like normal HbAA controls on a low protein diet. There was evidence that the HbSS subject might be in a precarious state with respect to sufficiency of several amino acids, particularly L-Arg, which is now classified as conditionally indispensable for the human.

Late-onset ornithine transcarbamylase deficiency in male patients

We report on 21 male patients who presented after 28 days of age with ornithine transcarbamylase (OTC) deficiency, which we define as late-onset OTC deficiency. These patients appeared normal at birth, but irritability, vomiting, and lethargy, which were often episodic, later developed. The age at presentation ranged from 2 months to 44 years. Biochemical testing revealed hyperammonemia, hyperglutaminemia, hypocitrullinemia, increased urinary orotate excretion, and decreased liver OTC activity measured in vitro, which ranged from 0% to 15% of normal. Male patients who were older at presentation had a somewhat different pattern of presenting symptoms and were more likely to die. These data illustrate the phenotypic variability of OTC deficiency. Unexplained episodes of repetitive or protracted vomiting in association with progressive alterations in behavior or neurologic findings should suggest the diagnosis of a urea cycle defect (or another symptomatic inborn error of metabolism), regardless of the age or medical history of the patient.

The urea cycle in the rett syndrome

The in vivo functioning of the urea cycle in the Rett syndrome (RS) was investigated using alanine load test. The test was carried out in girls with RS and their mothers. These results were compared with those on normal females, males and obligate carriers for ornithine carbamoyltransferase deficiency. Post load hyperammonaemia was not seen in any of the RS girls who were not on medication (valproate). Increased orotate excretion was found in some of the RS girls and mothers. The pattern of urinary excretion of orotate in RS girls and their mothers after a standardised alanine load was similar to that found in female carriers of OCT deficiency. This finding can be explained by a mutant OCT, subject to the Lyon effect, and linked to the gene abnormality in RS or by the presence of an abnormal X-linked mitochondrial protein synthesised in cytoplasma and positioned in close proximity to OCT. Such a protein abnormality could also affect mitochondrial functions in other cells, e.g. in the developing brain.

Effect on lipid in liver and serum, and some urinary components of dietary supplement of excess lysine given to previously starved or non-starved rats.

In order to obtain further information on the changes in liver lipids, either a basal or a lysineexcess diet was refed to previously starved rats or fed to previously non-starved rats. Liver lipid accumulation was observed in previously starved rats refed the lysine-excess diet for 7 days, but not in rats without previous starvation. The liver lipid did not accumulate with another 8 days' feeding (15 days' refeeding). The addition of methionine alone or in combination with threonine to the lysineexcess diet had no effect on the liver lipid level. The decrease in serum triacylglycerol in rats refed the lysine-excess diet was preceded by lipid accumulation in the liver. Urinary potassium during the initial two days increased with refeeding and feeding. Marked excretion of orotate was observed for 2 days from the initiation of refeeding of the lysine-excess diet and it then decreased. Thus, such a marked increase in the urinary excretion of orotate might be associated with the stimulation of orotate biosynthesis and with lipid accumulation in the liver.

Determination of urinary orotate excretion by high-performance liquid chromatography

A new reversed-phase high-performance liquid chromatography procedure for the determination of urinary orotate excretion is described. It is a selective, sensitive and rapid method, suitable for the differentiation of inherited metabolic diseases with abnormal orotate metabolism.

Effect of partial ornithine carbamoyltransferase deficiency on urea synthesis and related biochemical events.

The biochemical response to an intravenous alanine load of 0.25 g/kg was studied in nine adult female relatives of children with ornithine carbamoyltransferase deficiency. Six were classified as affected by partial deficiency and three as unaffected. The plasma ammonium concentration showed no change after the alanine load in the unaffected group, but marked increases occurred in all but one of the affected groups. The maximum rate of urea synthesis after the alanine load was decreased by 37% (P = 0.02) and delayed by 43% (P = 0.02) in the affected group. In the affected group a low rate of urea synthesis was associated with high urinary orotate excretion, high maximum plasma ammonium concentration and delay in the time taken to reach the maximum rate of urea synthesis (Kendall concordance W = 0.55, P less than 0.05). The effects of a higher dose of alanine and of oral protein were compared. The alanine load of 0.25 g of alanine/kg body weight was shown to provide an adequate stimulus to urea synthesis with a more rapid return of ammonium concentration to the pre-load level than with the protein load. The implication of these results in determining the distribution of flux control of urea synthesis, the discrepancy between them and predicted results and the necessary modifications to quantitative simulations are discussed.

The role of hepatic ornithine transcarbamylase deficiency in the orotic aciduria of pregnant mice

Groups of normal and heterozygote sparse-fur (spf) mutant mice were studied at various stages of gestation, to assess the effects of normal pregnancy on orotate excretion, hepatic mitochondrial urea cycle enzymes and any predisposition to the development of fatty liver. Results show a higher total daily excretion of urinary orotate by normal pregnant mice on the 8th and 15th days of gestation, which came to within the usual basal range of excretion of non-pregnant mutant heterozygotes with hereditary ornithine transcarbamylase deficiency. Liver ornithine transcarbamylase and carbamyl phosphate synthetase-I activities were reduced in pregnant mice on the 16th day of gestation ( P < 0.05). No fatty change, bile stasis or glycogen depletion was discernible on optical microscopy in normal or mutant mice. Nonspecific changes were seen on ultrastructural examination. Orotic aciduria seen in pregnant mice may be directly related to a physiological deficiency of liver ornithine transcarbamylase. However, the depletion of both the mitochondrial urea cycle enzymes, seen on the 16th day of pregnancy, may be indicative of a metabolic stress at the mitochondrial level.

Hepatotoxicity of sodium valproate in ornithine trasncarbamylase-deficient mice

Susceptibility to sodium valproate (SV) hepatotoxicity was investigated in male sparse-fur mutant (spf/Y) mice with X-linked ornithine transcarbamylase (OTC) deficiency, as compared to normals (+/Y). SV was given in drinking water, in increasing concentrations of 0, 0.05, 0.15 and 0.25%. Actual SV intake was similar in both groups. There were no significant changes in orotate excretion, but α-amino nitrogen increased progressively with SV intake in both groups. Valproate-treated animals also had a significant increase in hepatic carbamyl phosphate synthetase- I (CPS-I) activity. OTC-deficient spf/Y mice showed 33% mortality and morbidity at 0.05–0.15% valproate, while normal mice remained non-symptomatic. spf/Y Mice also showed a higher incidence of hepatocellular necrosis, microvesicular steatosis and polymorphic infiltration. Centrilobular necrosis was seen only in symptomatic OTC-deficient mice, indicating an idiosyncratic hepatotoxic response which may be different from the dose-related effects seen in all SV-treated mice. Electron microscopy of liver sections from severely affected spf/Y mice showed marked abnormalities of mitochondria, which appeared swollen or rounded. The rough endoplasmic reticulum was dilated and filled with a flocculent material. It is postulated that the idiosyncratic response in OTC-deficient mice may be caused by an interaction between a metabolic aberration of mitochondria and toxic metabolites of valproate.

1245 IDIOSYNCRATIC HEPATOTOXIC RESPONSE TO SODIUM VALFROATE (SV) IN A SUB-POPULATION OF SPARSE-FUR (spf) MUTANT MICE

Groups of six normal +/Y and six spf/Y mice with X-linked ornithine transcarbamylase (OTC) deficiency were given SV in drinking water in increasing concentrations of 0, 0.05, 0.15 and 0.25 % for 3 days each. Two out of six spf/Y mice became symptomatic at low intakes (34-85 mg/kg/d). One of these died and the other was sacrificed. Its plasma NH3 was 440 μM as compared to 255 ± 42 in other spf/Y. Four spf/Y and all +/Y survived the treatment (intake: 612-715 mg/kg/d). SV intake was similar in surviving animals. There were no significant changes in orotate excretion; α-amino N increased progressively. Liver carbamyl phosphate synthetase (CPS-I) increased significantly in treated mice, while no changes were seen in OTC activity.Severely affected spf/Y mice showed centrilobular necrosis with cytoplasmic vacuolization and microvesicular steatosis, indicating an idiosyncratic/hepatotoxic response. All other treatted mice showed random single cell necrosis. On electron microscopy, the severely affected spf/Y had swollen mitochondria, dilated RER and glycogen depletion. The results indicate the existence of a susceptible sub-population in spf/Y mice which develops SV toxlcity at low doses, and can be differentiated from other spf/Y mice by a different histopathologic response.(* Present address: Sandoz Ltd., CH-4002, Basel, Switzerland) (Research sponsored by the Canadian Liver Foundation)