Intracellular redox state and control of gluconeogenesis in perfused chicken liver

The role of the cellular redox state in the control of gluconeogenesis was studied in hemoglobin-free perfused chicken liver, by fluorimetric measurement of the redox states of intracellular pyridine nucleotides. The aminotransferase inhibitor, aminooxyacetate, completely inhibited gluconeogenesis from lactate in the perfused rat liver and to a small extent in the perfused chicken liver. In chicken liver, the highest rate of glucose production was seen with lactate, followed by fructose, pyruvate, and glycerol. When compared at 5 mM, the rate of glucose production from pyruvate was only 10% of that from lactate. Glucose production from a pyruvate/lactate mixture decreased with increasing proportions of pyruvate, together with redox changes of pyridine nucleotides to a more oxidized state. Increased reduction of pyridine nucleotides upon infusion of ethanol was associated with an increased glucose production from pyruvate, and the increase was abolished during octanoate infusion. This abolishment was accompanied by an increase in the acetoacetate to beta-hydroxybutyrate ratio with an oxidation of pyridine nucleotides. The octanoate-inhibited gluconeogenesis occurred at the higher lactate concentration (10 mM) with a transient oxidation of pyridine nucleotides. No significant inhibition was observed at 1 mM lactate, although an instant reduction of pyridine nucleotides was taking place. The rate of beta-hydroxybutyrate generation during octanoate infusion was 2.2 times higher at 1 mM than at 10 mM lactate. The inhibitory effect of octanoate on glyconeogenesis was completely relieved by the addition of NH4Cl. The results demonstrate that the regeneration of NADH in the cytosol is limited in chicken liver, and that gluconeogenesis is regulated, in part, by alteration in the redox states of mitochondria and cytosol.     

Amebic liver abscess: changing trends over 20 years

The metabolism of [2,3-13C]succinic acid dimethyl ester ([2,3-13C]-SAD) 10 mmol/L was examined in hepatocytes from overnight-fasted normal rats, 3-day starved rats, and overnight-fasted hereditarily diabetic Goto-Kakizaki (GK) rats. The amount of 13C-labeled succinate, fumarate, malate, lactate, alanine, and aspartate released by the hepatocytes was much higher in fasted normal rats than in starved or diabetic animals. Although the integrated areas of the 13C2 and 13C3 signals assigned to double-labeled malate, lactate, or alanine were not significantly different, the amount of single-labeled malate, lactate, alanine, and aspartate was higher in C3- versus C2-labeled isotopomers. The release of 13C-labeled glucose by the hepatocytes was lower in fasted versus starved or diabetic rats. Virtually all hexose molecules double-labeled in the C1-C2-C3 and/or C6-C5-C4 moieties corresponded to the [1,2-13C] and/or [5,6-13C] isotopomers. However, in the case of the single-labeled species, 13C-labeling of C1 (or C6) exceeded that of C2 (or C5). Both the single- and double-labeled molecules enriched with 13C in the C1-C2-C3 moiety were less abundant than those labeled in the C6-C5-C4 moiety, with such asymmetry being most marked in overnight-fasted normal rats, less pronounced in diabetic animals, and virtually absent in starved rats. These findings document that SAD is efficiently metabolized in hepatocytes, with its use as a gluconeogenic precursor being influenced by the nutritional and hormonal status of the animals. The present experiments also reinforce the view that asymmetrical labeling of glucose by 13C-labeled precursors is modulated by the relative contribution of exogenous and endogenous nutrients to the production of triose phosphates incorporated into the hexose.     

Regulation of energy metabolism of the heart during acute increase in heart work

We determined the contribution of all major energy substrates (glucose, glycogen, lactate, oleate, and triglycerides) during an acute increase in heart work (1 microM epinephrine, afterload increased by 40%) and the involvement of key regulatory enzymes, using isolated working rat hearts exhibiting physiologic values for contractile performance and oxygen consumption. We accounted for oxygen consumption quantitatively from the rates of substrate oxidation, measured on a minute-to-minute basis. Total beta-oxidation (but not exogenous oleate oxidation) was increased by the work jump, consistent with a decrease in the level of malonyl-CoA. Glycogen and lactate were important buffers for carbon substrate when heart work was acutely increased. Three mechanisms contributed to high respiration from glycogen: 1) carbohydrate oxidation was increased selectively; 2) stimulation of glucose oxidation was delayed at glucose uptake; and 3) glycogen-derived pyruvate behaved differently from pyruvate derived from extracellular glucose. Despite delayed activation of pyruvate dehydrogenase relative to phosphorylase, glycogen-derived pyruvate was more tightly coupled to oxidation. Also, glycogen-derived lactate plus pyruvate contributed to an increase in the relative efflux of lactate versus pyruvate, thereby regulating the redox. Glycogen synthesis resulted from activation of glycogen synthase late in the protocol but was timed to minimize futile cycling, since phosphorylase a became inhibited by high intracellular glucose.     

Radiotherapy results in early stage low grade nodal non-Hodgkin''s lymphoma

Severe lactic acidosis usually accompanies intense endurance exercise. It has been postulated that glycogen depletion working in concert with elevated muscle and plasma lactate levels lead to a concomitant reduction in pH. Their cumulative effect during prolonged physical exertion now leads to muscular fatigue and eventually limit endurance capacity. Therefore in the present study, dichloroacetate (DCA), a compound which enhances the rate of pyruvate oxidation thus reducing lactate formation, has been evaluated in a validated rat model of sub-maximal exercise performance. Male rats (350 g) were divided into two groups (control-saline, i.v. and DCA 5 mg/kg, i.v.) and were exercised to exhaustion in a chamber (26 degrees C) on a treadmill (11 m/min, 6 degrees incline). When compared to controls, the DCA-treated rats had longer run times (169 vs 101 min) and a decreased heating rate (0.020 vs 0.029 degrees C/min). In addition, DCA attenuated the increase in plasma lactate (28 vs 40 mg/dl) and significantly reduced both the rate and absolute amount of depletion of muscle glycogen stores. These results suggest that the activation of pyruvate dehydrogenase activity by DCA resulted in a reduction in the rate of glycogenolysis in addition to decreasing lactate accumulation by presumably limiting the availability of pyruvate for conversion to lactate, therefore increasing muscle carbohydrate oxidation via the TCA cycle. Thus DCA effected a significant delay in muscle fatigue.     

Endotoxin modulates arachidonic acid-induced glycogenolysis in the perfused rat liver

Effects of endotoxin on arachidonic acid (AA)-induced hepatic glycogenolysis were examined in perfused rat liver. In normal rat liver, infusion of AA increased oxygen consumption and glucose production concurrently. In rats injected with lipopolysaccharide (LPS) 6 h before, AA increased glucose production but suppressed oxygen consumption. The changes in LPS-injected rat were abolished by a thromboxane (Tx) A2 receptor antagonist. The release of Tx B2 by AA increased after LPS-injection. These results suggest that priming of hepatic macrophage by endotoxin in vivo enhances Tx synthesis, resulting in modulating hepatic glycogenolysis.     

Promotion of hepatic lipid oxidation and gluconeogenesis as a strategy for appetite control

There is considerable evidence that hepatic vagal afferents monitor the availability of liver glycogen and glucose metabolites, and that this mechanism participates in appetite regulation. Thus, promotion of gluconeogenesis and liver glycogen storage may enhance satiety. Hepatic lipid oxidation drives gluconeogenesis by positive allosteric modulation of pyruvate carboxylase and fructodiphosphatase. The rate-limiting enzyme for hepatic lipid oxidation, carnitine acyltransferase I, is activated by exogenous carnitine, and inhibited by malonyl coA. The lipogenesis inhibitor (-)-hydroxycitrate--a natural fruit acid found in the Brindall berry--can decrease production of malonyl coA in hepatocytes by potent inhibition of citrate lyase; many studies demonstrate that (-)-hydroxycitrate can reduce body fat accumulation in growing rats, owing in large part to a reduction in appetite. Joint administration of (-)-hydroxycitrate and carnitine should therefore promote hepatic lipid oxidation, gluconeogenesis, and satiety. Thermogenic effects as well as a reduction of the respiratory quotient can also be predicted. If this technique proves clinically useful in weight management, it could be used in conjunction with chromium picolinate and soluble fiber supplements, which appear to aid hunger control at the level of the hypothalamus and terminal ileum, respectively.     

Differential effects of nonselective nitric oxide synthase (NOS) and selective inducible NOS inhibition on hepatic necrosis, apoptosis, ICAM-1 expression, and neutrophil accumulation during endotoxemia

The roles of nitric oxide derived from either the constitutive endothelial NO synthase (eNOS or NOS3) or the inducible NOS (iNOS or NOS2) in hepatic injury during endotoxemia remain controversial. To investigate this further, rats received a bolus of lipopolysaccharide (LPS) following implantation of osmotic pumps containing one of two nonselective NOS inhibitors (NMA or NAME), one of two inducible NOS inhibitors (NIL or AG), or saline. The inhibitors were infused continuously into the liver via the portal vein. Treatment of LPS-injected rats with NMA and NAME resulted in 106 and 227% increases, respectively, in circulating hepatic enzyme levels compared to LPS-treated control rats. In contrast, infusion of the iNOS-selective inhibitors had no effect on the LPS-induced hepatic necrosis. In rats receiving NAME, LPS induced greater neutrophil infiltration and ICAM-1 expression than in the LPS + saline group, whereas NIL infusion did not. The increased hepatic necrosis and PMN infiltration in the LPS + NAME group was partially prevented by a simultaneous infusion of a liver-selective NO donor. Inhibition of PMN accumulation using an anti-ICAM-1 antibody or by PMN depletion using vinblastine pretreatment, however, did not reverse the increased necrosis with NAME infusion during endotoxemia. In contrast to the assessment for necrosis, increased apoptosis was observed in the livers of LPS-treated rats receiving infusions of either NAME or NIL, but not with LPS alone. These data indicate that NO produced by eNOS may be adequate to prevent necrosis by a mechanism independent of PMN, while induced NO appears to prevent apoptosis.     

Mechanisms of insulin resistance during acute endotoxemia

We characterized the mechanisms underlying acute endotoxin-induced alterations in glucose metabolism and determined the extent to which catecholamines mediate these changes. Acute endotoxemia was induced in chronically catheterized awake rats by a bolus injection of lipopolysaccharide (LPS; 1 mg/kg; LD10). Basal glucose turnover (Rt; infusion of [5-3H]glucose), in vivo insulin action on overall glucose utilization (euglycemic clamp), glycolysis, and glycogen synthesis were determined in four groups of rats. These groups received 1) LPS (LPS rats; n = 6), 2) saline (control rats; n = 6), 3) LPS and alpha beta-blockade (alpha beta-blockade and LPS rats; n = 9), or 4) saline and alpha beta-blockade (alpha beta-blockade control rats; n = 9). In the basal state, LPS induced hypotension and transient hyperglycemia. These changes were associated with glycogen depletion in both skeletal muscle and liver, and increased Rt. During hyperinsulinemia, whole body glucose disposal was 37% decreased (105 vs. 166 mumol/kg.min; P < 0.01). This whole body insulin resistance was characterized by decreased glycogen synthesis and glycogen synthase activity, but not by altered whole body glycolysis. alpha beta-Blockade abolished transient hyperglycemia, increased Rt, and accelerated basal liver glycogen depletion (45 vs. 105 mmol/kg dry, LPS and alpha beta-blockade rats vs. LPS rats; P < 0.05), but inhibited muscle glycogenolysis. alpha beta-Blockade did not reverse the insulin resistance induced by endotoxin. These data suggest that catecholamines counteract the LPS-induced increase in basal glucose turnover and stimulate muscle glycogenolysis during acute endotoxemia. These effects might explain the better preservation of hepatic glycogen in the absence than in the presence of alpha beta-blockade and serve as a defense mechanism against hypoglycemia. Catecholamines do not seem to be the immediate causes of insulin resistance during acute endotoxemia.     

Suppression of glycolysis is associated with an increase in glucose cycling in hepatocytes from diabetic rats

Rates of cycling between glucose and glucose 6-phosphate and between glucose and pyruvate, and the effects of these cycles on glucose metabolism, were compared in hepatocytes isolated from fasted normal or streptozotocin-induced diabetic rats. In diabetic hepatocytes the rate of glucose phosphorylation was 30% lower than that in normal hepatocytes, and there was a doubling of the rate of glucose/glucose 6-phosphate cycling. In addition, the rate of glycolysis was 60% lower in diabetic hepatocytes. This inhibition of glycolysis and stimulation of glucose/glucose 6-phosphate cycling appeared to be a consequence of the elevated rates of endogenous fatty acid oxidation observed in diabetic hepatocytes. The proportion of glycolytically derived pyruvate that was recycled to glucose was more than doubled in hepatocytes from diabetic rats compared with normal animals. This increase also appeared to be linked to the high rates of endogenous fatty acid oxidation in diabetic cells. As a consequence of the increased rates of both these cycles, 85% of all glucose molecules taken up by diabetic hepatocytes were recycled to glucose, compared with only 50% in normal hepatocytes. Glucose cycling is therefore likely to make a substantial contribution to the hyperglycemia of diabetes.     

Performance and cost-effectiveness of selective screening criteria for Chlamydia trachomatis infection in women. Implications for a national Chlamydia control strategy

At 9 mM glucose, experimental results show that mitochondrial phosphate depletion (induced by glucose phosphorylation, catalyzed by mitochondrial hexokinase) reduces the activities of the respiratory chain, oxidative phosphorylation, and glutaminase. Consequently, the 14C-lactate oxidation to 14CO2 is lowered in the presence of glucose. The fall of ATP level triggers a high aerobic glycolysis by deinhibiting fructose-6-P kinase. NADH, generated by enhanced glyceraldehyde-3-P dehydrogenase activity, increases the reducing power. Moreover, the lactate dehydrogenase (LDH) system is shifted toward lactate formation, while NAD+ is regenerated and the oligomycin-inhibited ATP production is replaced by the iodoacetate-inhibited ATP production. From 14CO2 production and lactate accumulation it is calculated that about 60% of 14C-glucose which disappears is channelled into extraglycolytic reactions. On the contrary, 82% of glucose below l mM is metabolized through non-glycolytic reactions. The pyruvate kinase-M2 (PK-M2) inhibition does not limit the glycolytic flow from 9 mM glucose, but it may cause sustained gluconeogenesis.     

Hyperpolarization of the liver cell membrane by palmitate as affected by glucose and lactate: implications for control of feeding

Since the membrane potential of liver cells being in contact with vagal afferents has been proposed to represent a major signal in metabolic control of food intake, we investigated the effect of palmitate, glucose and lactate on the membrane potential of hepatocytes with microelectrodes using superfused mouse liver slices. The mice used for the experiments were fed a fat-enriched diet (18% fat). Palmitate (0.5 mM) hyperpolarized the membrane of hepatocytes by 3-4 mV, and this hyperpolarization was not affected by 5-10 mM glucose and 0.5-1 mM lactate. Glucose alone did not influence the potential, even when mice fed a high carbohydrate diet were employed. At lactate concentrations > or = 2 mM the palmitate induced hyperpolarization was eliminated and 5 mM lactate or pyruvate alone hyperpolarized the liver cell membrane. Similar to the palmitate induced hyperpolarization, the lactate induced hyperpolarization was prevented by the K-channel blocker TEA, suggesting that activation of K channels is involved in the hyperpolarization. The results show that physiological concentrations of glucose and lactate do not affect the hyperpolarization of the liver cell membrane due to fatty acid oxidation. The implications of these findings with regard to control of food intake by fatty acid oxidation and lactate metabolism are discussed. The observations are consistent with a signal function of the hepatic membrane potential in physiological control of food intake by fatty acid oxidation. Hepatic lactate metabolism at supraphysiological lactate concentrations may also produce a satiety signal coded by the hepatic membrane potential.     

Instrumental and metabolic evaluation of patients affected by peripheral arterial occlusive disease (PAOD) following surgical revascularization surgery

Experiments were performed on eight subjects affected by peripheral arterial occlusive disease (PAOD) of the lower limbs. Each patient was submitted to Ecodoppler, angiography and the "Treadmill test". Two bioptic muscle of these patients. A sample was used for the spectrophotometric and spectrophotofluorimetric determinations of: glycogen, pyruvate, lactate, citrate, alpha-ketoglutarate, malate, aspartate, glutamate, AMP, ADP, ATP and creatine phosphate (CP). The other bioptic sample was used to determine the following enzyme activities: hexokinase, phosphofructokinase, pyruvate kinase, lactate dehydrogenase, citrate synthase, succinate dehydrogenase, malate dehydrogenase, total NADH cytochrome c reductase, cytochrome oxidase, aspartate aminotransferase and alanine aminotransferase. Patients showed an increase in lactate dehydrogenase, total NADH cytochrome c reductase and succinate dehydrogenase activities, a decrease in glycogen, ATP and CP concentrations. Telethermographic data showed patient muscle thermic emission quantitatively different from control group. The telethermographic test can be used as an additional diagnostic tool to determine and monitor the efficiency of a muscle undergoing metabolic failure.     

The role of malonyl-coa in the coordination of fatty acid synthesis and oxidation in isolated rat hepatocytes

Fatty acid synthesis and fatty acid oxidation were examined in rat hepatocytes under a variety of experimental conditions. In cells from fed animals, glucagon acutely switched the direction of fatty acid metabolism from synthesis to oxidation. Addition of lactate plus pyruvate had the opposite effect. The inhibitory action of glucagon on fatty acid synthesis and its stimulatory effect on fatty acid oxidation were largely, but not completely, offset by the simultaneous addition of lactate plus pyruvate. Changes in cellular citrate and malonyl-CoA levels indicated that glucagon exerted its inhibitory effect on fatty acid synthesis at two levels: (i) blockade of glycolysis; and (ii) partial inhibition of a more distal step, probably acetyl-CoA carboxylase. Under all conditions, fatty acid oxidation was related in a linear and reciprocal fashion to the rate of fatty acid synthesis and the tissue malonyl-CoA content. The latter fluctuated through a range of 1 to 6 nmol per g wet weight of cells. Since malonyl-CoA inhibits carnitine acyltransferase I of liver mitochondria with a Ki in the region of 1 to 2 micron, the present studies support the concept that this compound plays a pivotal role in the coordination of hepatic fatty acid synthesis and oxidation. The ketogenic effect of glucagon on liver appears to be manifested in large part through the ability of the hormone to reduce the tissue malonyl-CoA concentration.     

Effects of glucagon-like peptide 1 on the kinetics of glycogen synthase a in hepatocytes from normal and diabetic rats

Glucagon-like peptide 1(7-36)amide (GLP-1) is currently under investigation as a possible tool in the treatment of non-insulin-dependent diabetes mellitus. In addition to enhancing nutrient-stimulated insulin release, the peptide also favors glycogen synthesis and glucose use in liver, muscle, and adipose tissue. GLP-1 also activates glycogen synthase a in hepatocytes from both normal and diabetic rats. In the present study, the kinetic aspects of such an activation were investigated in hepatocytes from normal rats and from animals rendered diabetic induced by injection of streptozotocin, either in the adult age (insulin-dependent diabetes mellitus model) or in days 1 or 5 after birth (non-insulin-dependent diabetes mellitus models). GLP-1 increased, in a dose-dependent manner, glycogen synthase a activity in the hepatocytes from all groups studied. The activation of the enzyme reached a steady state within 1 min exposure to GLP-1, which, at 10(-12) M, caused a half-maximal activation. When comparing fed vs. overnight-starved normal rats, a somewhat lower basal activity of glycogen synthase a in fasted animals (P < 0.05) coincided with a greater relative increment in reaction velocity in response to GLP-1. The basal activity of glycogen synthase a and the relative extent of its inhibition by glucagon or activation by insulin and GLP-1 were modulated by the extracellular concentration of D-glucose. The activation of glycogen synthase a by either insulin or GLP-1 resulted not solely in an increase in maximal velocity but also in a decrease in affinity of the enzyme for uridine diphosphate-glucose; in diabetic animals, the capacity of insulin or GLP-1 to increase the maximal velocity and Michaelis-Menten constant were less marked than in normal rats. In conclusion, this study indicates that the GLP-1-induced activation of glycogen synthase a displays attributes of rapidity, sensitivity, and nutritional dependency that are well suited for both participation in the physiological regulation of enzyme activity and therapeutic purpose.     

Bacterial lipopolysaccharide induces uncoupling protein-2 expression in hepatocytes by a tumor necrosis factor-alpha-dependent mechanism

The liver is a target for bacterial lipopolysaccharide (LPS) and participates in the metabolic response to endotoxemia. Recently published evidence indicates that LPS increases the expression of mitochondrial uncoupling protein-2 (UCP-2) mRNAs in several tissues, including the liver. Because hepatocytes in the healthy liver do not express UCP-2, LPS was thought to induce UCP-2 in liver macrophages, which express UCP-2 constitutively. However, the present studies of cultured peritoneal macrophages indicate that LPS reduces steady state levels of UCP-2 mRNAs in these cells. In contrast, UCP-2 mRNAs are induced in hepatocytes isolated from LPS treated rats and transfection of these hepatocytes with UCP-2 promoter-reporter constructs demonstrates substantial increases in UCP-2 promoter activity. LPS induction of hepatocyte UCP-2 expression is virtually abolished by prior treatment of rats with neutralizing antibodies to tumor necrosis factor alpha (TNF). Futhermore, TNFalpha treatment induces UCP-2 mRNA accumulation in primary cultures of hepatocytes from healthy rats. Thus, hepatocytes are likely to be important contributors to endotoxin-related increases in liver UCP-2 via a mechanism that involves the LPS-inducible cytokine, TNFalpha.     

Effect of aniline on ethanol oxidation and carbohydrate metabolism in isolated hepatocytes

The addition of aniline to isolated hepatocytes derived from fasted rats and incubated with ethanol, caused a 30-60% decrease in the rate of ethanol oxidation. The degree of inhibition was dependent on aniline concentration, 5 mM causing near-maximal inhibition. Aniline reduced the activity of alcohol dehydrogenase in a noncompetitive manner, but had no effect on aldehyde dehydrogenase activity nor on reducing-equivalent transfer between the cytoplasm and mitochondria. The inhibition of alcohol dehydrogenase by aniline was associated with a decrease in the inhibitory effects of ethanol on glycolysis. Aniline, added to hepatocytes in the presence or absence of ethanol, inhibited gluconeogenesis from lactate and pyruvate, but not from sorbitol or fructose.     

Biochemical compartmentation of fish tissues. I. Brain energy reserves and its metabolic products

The quantitative distribution of glycogen, lactate and pyruvate in the brain was studied in 4 regions of 9 fishes. The highest glycogen, lactate and pyruvate content was present in major carps followed by cat fishes and snake headed fishes. Glycogen and lactate contents were highest in the medulla oblongata while the highest pyruvate level was observed in the cerebellum. The observed differential distribution of glycogen, lactate and pyruvate in the different regions of the brain is discussed in relation to their functional differention and may depend on the nature of the diet, on the environment and growth rate, etc., of the fishes.     

Long-term maintenance of functional rat hepatocytes in primary culture by additions of pyruvate and various hormones

Mature adult rat hepatocytes were cultured as monolayers in serum-free Williams medium E containing 10(-7) M each of insulin (Ins), dexamethasone (Dex) and triiodothyronine (T3) and 30 mM pyruvate. The hepatocytes remained morphologically intact for at least 14 days, during which period they maintained normal liver functions such as the expressions of cytochrome P-450 mRNA and glucokinase and secretion of albumin. They also retained the ability to resume proliferation. Cells cultured with pyruvate had a much higher ATP level than those without pyruvate, suggesting that pyruvate can sustain functional hepatocytes for a long period in culture in the presence of Ins, Dex and T3, probably by producing enough energy for their maintenance.     

Leptin increases glucose transport and utilization in skeletal muscle in vitro

1. The present study examines the effect of leptin on glucose transport and metabolism in incubated soleus muscle from male lean albino rats. 2. Insulin (100 microU/ml) increased glucose uptake by twofold while the leptin group (100 nmol/l) reached 75% of the insulin response after 1 hr of incubation. However, leptin did not potentiate the insulin effect on glucose uptake in soleus muscle. 3. Leptin elicited a significant increase (27.7%) in total lactate production, accompanied by a three-fold increment in glycogen synthesis from [U-14C]D-glucose. 4. Insulin raised glycogen synthesis by sixfold. The leptin plus insulin group increased glycogen synthesis by eightfold, which is equivalent to the sum of the separated leptin and insulin groups. 5. Leptin per se exerts an insulin-like effect stimulating glucose uptake, glycogen synthesis, and lactate formation and also seems to potentiate the effect of insulin on glucose incorporation into glycogen in incubated soleus muscle.     

Hepatotoxicity of halogenated inhalational anesthetics studied in rats hepatocytes

Effects of 2% halothane, 1.5% sevoflurane, 1.5% enflurane, and 1.2% isoflurane on hepatic dysfunction were studied using rat hepatocytes incubated in media containing 95% or 5% O2. The effects of anesthetics on hepatic perfusion were eliminated by incubation of hepatocytes for 45 minutes with each combination of anesthetic and oxygen concentration. After incubation, viability of hepatocytes was assayed by the LDH latency test. Enzyme (GPT, GOT, LDH) activities, lactate concentration and pyruvate concentration in the incubation medium were measured. The concentrations of adenine nucleotides and inorganic phosphorous in the liver were determined. Anesthetics administered in 95% O2 did not produce significant decreases in viability and enzyme release compared to 95% O2 alone. Halothane, sevoflurane, and isoflurane administered in 5% O2 produced significant decreases in viability and enzyme releases compared to 95% O2 alone. In groups administered 95% O2 there was a significant relationship between viability and energy charge in hepatocytes (P < 0.01). In the 5% O2 groups, there were significant relationships between viability and ATP in hepatocytes (P < 0.01) or L/P ratio in incubation medium (P < 0.01). These results suggest that the combination of anesthetics and hypoxia produce hepatotoxicity. Destruction of energy status might be the cause of hepatotoxicity.