Effects of liver ischemia on hepatic protein synthesis in vitro and in vivo

When an in vitro system is used to study the influence of ischemia on hepatic protein synthesis, an important question is whether alterations observed in vitro reflect changes in vivo. In the present study the effects of liver ischemia on protein synthesis were investigated in rats both in vitro and in vivo. Liver ischemia was induced by hepatic artery ligation. Protein synthesis in vitro was determined from leucine incorporation into proteins in liver slices incubated in a medium containing ¹⁴C-leucine (0.5 mmol/l) and in vivo from leucine incorporation into hepatic proteins after intraportal injection of a tracer dose of ¹⁴C-leucine. Leucine incorporation rate in non-ischemic liver was 0.16 pmol * g pror¹ h-¹ in vitro and 19.6 μmol g prot-¹. h^-1 in vivo. After hepatic artery ligation protein synthesis in vitro was reduced by about 60% and in vivo by about 80%. Thus, the relative changes were of the same magnitude in vitro and in vivo. This indicates that an in vitro system can be used to evaluate the effects of liver ischemia on hepatic protein synthesis.     

Acute effects of selected hepatotoxic agents on polyribosomes and protein synthesis in the livers of rats fed purified diets containing hepatocarcinogens

This investigation was concerned with the effects of certain hepatocarcinogens ingested for long- or short-term intervals upon selected responses of rat liver to specific hepatotoxic stimuli. Rats fed ad libitum for long (3 to 45 weeks) periods or force-fed for short (3 days) periods purified diets containing a single hepatocarcinogen, ethionine, N-2-fluorenylacetamide, 3′-methyl-4-dimethylaminoazobenzene, or thioacetamide, were subjected to the acute administration of a single hepatotoxic agent, such as puromycin, actinomycin D, sparsomycin, hypertonic NaCl or CCl₄, and hepatic protein synthesis (in vitro) and polyribosomal aggregation were evaluated. Relating to in vitro protein synthesis, in long-term experiments, puromycin induced less inhibition in the experimental groups than in the control (basal diet) group; and actinomycin D inhibited all groups similarly; while in the short-term experiments, puromycin induced less inhibition in the experimental groups than in the control group; actinomycin D inhibited he basal and ethionine groups more than the other groups; sparsomycin induced variable degrees of inhibition, with the ethionine group showing the most and the N-2-fluorenylacetamide and thioacetamide groups showing the least; hypertonic NaCl markedly inhibited all groups similarly; and CCl₄ induced somewhat variable degrees of inhibition, with the thioacetamide group showing less than the other groups. In a few long-term experiments where hepatomas were evaluated in relation to surrounding liver tissue, puromycin administration induced less inhibition in the hepatomas than in the surrounding liver while actinomycin D administration induced similar inhibition in both groups. In general, in all experiments the degree of polyribosomal disagregation was parallel to that of inhibition of protein synthesis (in vitro). The results reveal that the livers of rats fed purified diets containing selected hepatocarcinogens responded in a variable manner in regard to protein synthesis (in vitro) and status of polyribosomal aggregation to the acute administration of selected hepatotoxic agents.     

Protein Synthesis with Special Reference to S-100 Protein in Brain Slices from Rats Receiving a Restricted Protein Supply

The synthesis of S-100 protein and that of soluble and total proteins was investigated using cerebral slices from rats fed a 20% or 3% protein containing diet for 6 days. Incorporation of radioactive amino acids into S-100 protein was significantly higher when rats were fed a diet containing 20% protein. No significant differences were obtained in the radioactivity incorporated into total or soluble proteins between the 2 dietary groups. ¹⁴C-leucine of a specific radioactivity of 55 mCi/mmol or 3.2 mCi/mmol incorporated with time into total protein was similar for the 2 dietary groups. The time-dependent uptake of ¹⁴C-leucine by the slices and the inulin space remained unaffected by the dietary conditions used; and amino acid analyser estimates of the free amino acid pool showed no significant differences. Brain wet weight was 1.54 ± 0.02 g and 1.39 ± 0.02 g for protein-fed and protein-restricted rats respectively. The corresponding body weight increased by 7.8 g/day or fell by 0.5 g/day. Although the differences observed in total protein synthesis were small the synthesis of a nervous tissue specific protein S-100 was markedly affected by short-term protein restriction.     

Nucleolar-cytoplasmic transport of RNA in livers of rats fed a deficient diet.

Electron microscopic autoradiographic studies were conducted on livers of young female rats that were force-fed a complete or threonine-devoid diet for 3 days. Groups of control and experimental rats were killed on the fourth morning 15 or 60 min after intraperitoneally administering [³H]orotic acid. The electron microscopic autoradiographs of liver sections revealed grains attributed to RNA overlying or adjacent to several small nucleoli of hepatocytes of control rats. In contrast, the livers of the experimental rats revealed grains overlying single enlarged nucleoli and often they were present at the nucleolar-nuclear membrane junctions. Grains appearing in the cytoplasm of the livers of experimental rats were observed in the vicinity of the nucleolar base where it was attached to the nuclear membrane. Thus our results suggest that newly synthesized nucleolar RNA of the livers of rats force-fed a threonine-devoid diet migrates in the nucleolus toward its base at the nucleolar-nuclear membrane junction and then exits from the nucleus through the nuclear pores to enter the cytoplasm.     

Smooth endoplasmic reticulum proliferation and increased cell multiplication in cultured hepatocytes of the newborn rat in the presence of phenobarbital

Twenty-eight-day old male Sprague-Dawley rats were fed diets containing 2, 5, 10, 15, 25, or 50% protein and varying levels of fat and energy for 8 weeks and were then killed. The livers of rats fed the 2% lactalbumin protein diet ad libitum but not the others showed visible fat. This difference was confirmed histologically. Chemical analyses of the livers indicated that the rats fed the 2% protein diets ad libitum had experienced a substantial reduction in liver cell size but only a very slight reduction in the lipid content of each cell. As a consequence, their livers showed a net increase in the amount of lipid per g of tissue and this gave them a fatty appearance. The lipids formed large fat globules suspended in a greatly reduced amount of protoplasm; there were no fat globules outside the cells. Therefore, the fatty liver of protein deficiency was a physical phenomenon rather than a metabolic defect. Rats fed the 2% protein diets in restricted amounts also had shrunken cells but they did not manifest fatty liver because there was a relatively greater reduction in liver cell lipid than in liver cell size. There was a reduced amount of protein in the livers of rats fed 2 and 5% protein diets which can be explained by a reduction in liver RNA and in Protein/RNA ratio. Protein/RNA ratio was enhanced by starvation. Liver protein content showed a biphasic response to variation in dietary fat level with its lowest value in rats fed diets containing 11.9% fat.     

Induced biochemical and physiological changes in young and adult growing rats fed on a vegetable or animal protein diet

The influence of diets containing faba bean or casein as sources of protein were studied in rats at two stages of development. A significant impairment of growth rate, carcass, liver and skeletal muscle were found in young and adult rats fed for a period of 10 days on raw legume. Urinary urea output and the activities of three amino acid degrading enzymes: arginase, alanine aminotransferase and arginine succinate synthetase were all affected by the dietary protein and the stage of development. Urinary creatinine excretion was higher in the adult rats, while serum cholesterol was slightly increased in the young ones. Changes in plasma zinc may be attributed to a reduced zinc bioavailability to rats from the faba bean diet. Other biochemical parameters measured (glucose, triglycerides and plasma proteins) remained unchanged in all the experimental groups. Liver DNA and RNA content (mg/g tissue) decreased with age in both dietary groups, which were accompanied by an increase in tissue size. Furthermore, liver RNA concentration (primarily a measure of protein synthesis capacity) was enhanced in the adult legume fed rats. In this context, it is suggested that other organs (particularly muscle, with lower amino acid requirements for protein synthesis as a consequence of the stunting of growth) could contribute to increase the amino acid supply to liver in the animals fed on the faba bean diet.     

Age decreased steady-state concentrations of genistein in plasma, liver, and skeletal muscle in Sprague-Dawley rats

Soy isoflavones are associated with low incidence of cardiovascular diseases (CVD) and hormone-dependent cancers, but no solid information is available on the relative deposition of isoflavones in the body as a function of age. One-year-old (adult) male Sprague-Dawley rats were fed control diet or one of three high-genistein isoflavone (HGI) diets at a dose of 62, 154, or 308 genistein mg/kg (ppm) diet for 5 weeks; 2-year-old (old) were fed a dose of 154 or 308 ppm. Steady-state genistein concentrations in plasma, liver, and gastrocnemius muscle of the adult rats after 12 h fast revealed a linear dose-dependent manner (P < or = 0.0001). However, there was no such relationship in the old rats. Nevertheless, old rats fed the 308 ppm genistein diet had significantly lower steady-state genistein concentrations in plasma and liver than the adult rats did (P < or = 0.05); but similar genistein concentration in muscle. The results of this study indicate that steady-state genistein concentrations in tissues of adult rats after 12 h fast exhibited a dose-dependent fashion and were diminished in specific tissues by age.     

Effects of Cyclic Starvation-Feeding and of Splenectomy on the Development of Hemosiderosis in Rat Livers

The development of siderosis of liver and spleen was investigated in rats subjected alternately to periods of starvation and periods of feeding of diets rich in iron (0.71% or 1.23% Fe) or of control diets, during periods ranging up to 245 days. With 0.71% iron in the diet, cyclic starvation-feeding markedly enhanced the accumulation of iron in rat livers by comparison to feeding ad libitum even though rats fed ad libitum ingested far greater total amounts of iron than cyclically fed rats. With 1.23% iron in the diet, the concentration of iron in livers reached more or less the same plateau in cyclically starved-fed rats and in rats fed ad libitum (betwen 4 and 5 mg Fe/g wet weight); but the mean rate of accumulation of iron in the livers of cyclically starved and fed rats was more than twice that in rats fed ad libitum, whereas mean ingestion of iron per feeding day was only 16% higher in the former group. Surgical removal of the spleen enhanced the accumulation of iron in the liver in cyclically starved-fed rats and in rats fed ad libitum. Histologically, siderosis of the liver was moderate in rats fed the diet with 0.71% iron but was severe in rats fed the diet with 1.23% iron and most severe in those without spleens. Stainable iron was deposited in hepatocytes and in Kupffer cells. None of the rats developed cirrhosis of the liver. The data suggest that in rats a barrier to the absorption of iron from the gut, or to its later utilization, is surmounted if the concentration of iron in the food exceeds a certain limit value, somewhere between 0.71 and 1.23%. With iron in the food below this value, cyclic starvation-feeding markedly potentiates accumulation of iron in the liver in the course of several months, but siderosis is moderate. With iron in the food above the limit value, cyclic starvation-feeding and feeding ad libitum can equally lead to massive siderosis of the liver.     

Effects of short-term dietary change from high fat to high carbohydrate diets on the storage and utilization of glycogen and triacylglycerol in untrained rats

The effects of short-term diet change from high fat (F) to high carbohydrate (C) (or vice versa) on the storage and utilization of glycogen and triacylglycerol (TG) in muscle and liver were studied in untrained rats. Rats were fed on an F or C diet for 28 days. For an additional 3 days, half of the rats in both F and C groups were fed the same diets as before (F-F and C-C) and the other half of the rats were switched to the counterpart diets (F-C and C-F). On the final day of the experiment, half of the rats in each diet group were exercised by swimming for 1.5 h and the other half were rested. Short-term diet change from F to C diets increased, but the change from C to F diets decreased, glycogen stores of soleus and plantaris muscles and liver, resulting in no difference in glycogen stores between F-C and C-C, and between F-F and C-F. The dietary change also had an affect on TG stores of red gastrocnemius muscle and liver - however, muscle TG stores were still higher in F-C than in C-C and C-F, and there were no differences in liver TG stores between F-C and C-F. Exercise decreased muscle glycogen contents markedly in F-C and C-C, whereas, it decreased muscle TG concentrations in F-F and C-F. Liver glycogen depletion was lower in F-C than in other groups. Lipolytic activities of epididymal adipose tissue at rest and postexercise were no differences between F-F and F-C, and were higher in F-C than in C-C and C-F. -adrenergic receptor binding was determined with [¹²⁵I] iodocyanopindolol, and maximal numbers of -adrenergic receptor of plasma membrane from perirenal adipose tissue were approximately 170%–200% higher in F-C than in other groups at rest and postexercise. These results suggested that short-term C diet fed rats adapted to F diet enhanced not only glycogen stores of muscle and liver but also did not decrease lipolytic activity of adipose tissue with increased -adrenergic receptor density, resulting in the preservation of energy reserves (glycogen and TG) of muscle at rest, and liver glycogen sparing during exercise.     

Effect of exercise on Hepatic cholesterof of rats fed diets high in saturated or unsaturated fats

Summary Rats have been fed diets containing 24.2% coconut or corn oil or an equal mixture of each for 14–18 weeks. Half of the animals in each dietary group were exercised by running in motor-driven work wheels throughout the entire experimental period. During the final 10–14 weeks, these exercised animals ran continuously for 60 min at 1.0 mph or faster each day. Comparisons between sedentary groups revealed that hepatic cholesterol and excretion of digitonin precipitated sterols in the feces increased (P < 0.01) as the per cent of unsaturated fat (corn oil) in the ingested food increased. In contrast, total liver lipid decreased (P < 0.01) as the consumption of corn oil increased. No change in plasma cholesterol occurred in the sedentary rats in response to the three diets. Hepatic cholesterol of the exercised groups was significantly less (P < 0.05–P < 0.01) than that of their respective control groups (same diet). However, the group fed the corn oil diet had a significantly higher (P < 0.01) liver cholesterol after exercise than did the exercised group fed the coconut oil diet. Liver lipid was reduced (P < 0.01) by exercise in the corn oil and mixed corn-coconut oil fed groups. Plasma cholesterol and sterol excretion were unchanged by the exercise program.This investigation supported by Research Grant HE 08262 from the National Heart Institute, National Institutes of Health, U.S. Public Health Service.     

Scanning microdensitometry of glycogen zonation in the livers of rats adapted to a controlled feeding schedule and to 30, 60, or 90% casein diets

The effect of diet composition on diurnal changes in glycogen zonation patterns in rat liver was investigated in individually-caged male Sprague-Dawley rats adapted to the 2 + 22 controlled feeding and lighting schedule and to diets containing 30% casein/55% carbohydrates, 60% casein/25% carbohydrates, or 90% casein (30 rats/dietary group). Three rats from each dietary group were killed at the following times relative to the onset of feeding (0 min):?60, ?30, 0, 15, 30, 45, 60, 90, 120, and 180 min. Glycogen in cryostat sections from the median and right lateral lobes of the liver was fixed and stained by standard techniques. The optical density of glycogen at points along the path between the central and portal veins of a given lobule was determined, and lobular glycogen gradients of replicate animals were integrated to form a composite lobular glycogen distribution profile. In the period from ?60 to 0 min, liver glycogen levels were similar for rats on any of the diets, and the glycogen concentration was similar in periportal (P), midlobular (M), and centrilobular (C) hepatocytes. During the 0- to 45-min period, diet-related glycogen depletion occurred (90 > 60 ? 30% casein) by asymmetrical glycogen loss (P > M ? C hepatocytes) from the liver lobules. Similar food intake curves occurred for all diets. During the 45- to 180-min period, asymmetrical glycogen accumulation began in lobular parenchymal cells (P > M ? C hepatocytes), and the rate of accumulation was related to dietary composition (30 > 60 ? 90% casein). The differential responses of parenchymal cells within liver lobules to physiological stimuli resulted in glycogen distributional changes that were rapid and of large magnitude. Our results are consistent with the hypothesis that periportal and midlobular hepatocytes are more metabolically responsive and active than centrilobular hepatocytes.     

The preference-aversion behavior of rats for nutritionally-controlled diets containing oil or fat

The preference-aversion behavior of Sprague-Dawley rats for semi-purified diets containing various levels of either corn oil, animal fat (lard) or hydrogenated vegetable oil (Crisco) was determined in brief and long-term, two-choice preference tests. After the first day of exposure, rats selected more calories from the control-unadulterated diet than from experimental diets containing either oil or fat. Only in the first hour of exposure (with diets containing either 5% Crisco, 15 or 25% lard) or in the first day (with diets containing 5% Crisco, 5 or 15% lard) was a preference for experimental diets observed. There were significant effects of oil and fat concentration and of exposure time upon preference for experimental diets, such that preference for the experimental diets was reduced as levels of oil or fat and as the time of experiment increased. Changes in diet selection had no major effect on total caloric intake measured as the sum of both experimental and control dietary choices, although total intake on a daily basis tended to decrease during the middle portion (2-4 days) of the study, then recover by the end of the study. The results suggest that postingestional factors related to the dietary oil and fat (rather than the sensory properties of the diets) directed the long-term preference behavior of rats for semi-purified diets adulterated with various levels of either corn oil, animal fat (lard) or hydrogenated vegetable oil (Crisco).     

Biological Evaluation of Protein Quality and Safety of a Lipoprotein Concentrate from Niger Seed* (Guizotia Abyssinica Cass.)

Growing rats were fed diets containing a lipoprotein concentrate prepared from niger seeds, as the sole source of protein. Protein efficiency ratio (PER) and productive protein value (PPV) were calculated for two levels of protein in the diet, 11.1 % and 22.4 %, respectively. When the protein content of the diet was 11.1 % the PER was 2.37 (for a feeding period of three weeks). The corresponding PPV was 42.0. A short term toxicological feeding study was carried out on rats fed with these diets for a 90-day period. No significant diet-related changes were observed in the growth, appearance, behaviour and survivals of the animals. Hematological findings were normal. The histopathological examinations of the main organs gave normal results. Electron microscopy showed an increased amount of multi-vesicular bodies (possibly lysosomes) as well as an increased content of fat droplets in the cytoplasm of liver cells. The significance of these changes remains unclear.     

Protein synthesis with special reference to S-100 protein in brain slices from rats receiving a restricted protein supply.

The synthesis of S-100 protein and that of soluble and total proteins was investigated using cerebral slices from rats fed a 20% or 3% protein containing diet for 6 days. Incorporation of radioactive amino acids into S-100 protein was significantly higher when rats were fed a diet containing 20% protein. No significant differences were obtained in the radioactivity incorporated into total or soluble proteins between the 2 dietary groups. 14C-leucine of aspecific radioactivity of 55 mCi/mmol or 3.2 mCi/mmol incorporated with time into total protein was similar for the 2 dietary groups. The time-dependent uptake of 14C-leucine by the slices and theinulin space remained unaffected by the dietary conditions used; and amino acid analyser estimates of the free amino acid pool showed no significant differences. Brain wet weight was 1.54+/-0.02 g and1.39+/-0.02 g for protein-fed and protein-restricted rats respectively. The corresponding body weight increased by 7.8 g/day or fell by 0.5 g/day. Although the differences observed in total protein synthesis were small the synthesis of a nervous tissue specific protein S-100 was markedly affected by short-term protein restriction.     

Haemorheological Characterisation of a Model of Diet-induced Hypercholesterolaemia in Rats

This investigation examined the haemorheological changes that occurred in rats during and after administration of high-fat diets containing 10% lard and 1% cholic acid, both with and without 2% cholesterol for a period of 2 weeks. Rats fed the high-fat diet enriched with cholesterol showed markedly impaired fluidity of whole blood, increases of blood viscosity, plasma low-density lipoprotein (LDL)-cholesterol, VLDL-cholesterol, and impaired skin microcirculation shortly after the start of this diet regimen. Additionally, increased liver mass, and lipid accumulation in hepatocytes were observed after two weeks. In contrast, rats fed the high-fat, cholesterol-free diet had less pronounced haemorheological disorders, increased liver mass and lipid accumulation in the liver than rats given the cholesterol-enriched diet. Histopathologically, there were no marked changes in blood vessels or liver in rats fed these diets for 2 weeks, except for fatty liver changes. Dietary induced hypercholesterolaemia therefore contributes to the haemorheological disorder, impaired microcirculation and fatty liver change.     

Maternal diet affects feeding behaviour of self-selecting weanling rats

Weanling rats have been shown to regulate protein intake when provided the opportunity. The objective of these studies was to determine the influence of protein concentration in the maternal diet on protein selection of the offspring. Dams were fed single diets containing 10, 20, 30, or 40% casein throughout gestation and lactation. Pups from all groups were then allowed to select from diets containing 10 and 60% casein for two weeks. Protein selection in the weanling rat was found to correlate with protein concentration in the maternal diet expressed either as g/100g BW (r=0.48, p<0.05) or protein concentration (r=0.65, p<0.01). In order to differentiate between a gestational and lactational effect a cross-over study was performed. Pups from dams fed 10% casein during gestation and/or lactation selected 17% protein concentration, lower than the 25.1% selected by pups from dams receiving 40% casein through gestation and lactation. Therefore, feeding behaviour, as indicated by protein selection, is influenced by maternal diet during both gestation and lactation. The use of the self-selection paradigm has been proposed as a behavioural model for the investigation of the functional significance of maternal nutrition.     

Effect of Glucagon on Cyclic AMP,Albumin Metabolism and Incorporation of 14C-Leucine into Proteins in Isolated Parenchymal Rat Liver Cells

Parenchymal rat liver cells were isolated by the collagenase method and incubated in Krebs-Henseleit buffer containing 0.5% gelatin. The basal level of cyclic AMP in isolated cells was 0.52 nmol per g liver wet wt. Glucagon (10^-10- 10^-6 M) caused a significant increase in the level of cyclic AMP. Maximum levels were obtained 2–15 min after addition of glucagon. Repeated administration of glucagon caused a new increase in cyclic AMP, but the response was lesser than after the first addition of glucagon, indicating refractoriness to glucagon. The rate of albumin secretion was 4.6 μg/min per g liver wet wt. This is about the rate found in the perfused liver. Glucagon (10^-8- 10^-6 M) inhibited albumin secretion and the incorporation of ¹⁴C-leucine into albumin, into total proteins in the medium and into total proteins in the cell suspension. The effect of glucagon on albumin secretion is compatible with an effect on the rate of synthesis. A positive correlation existed between the maximal level of cyclic AMP after glucagon administration and the inhibition of both albumin secretion and the incorporation of ¹⁴C-leucine.     

High fructose feeding of magnesium deficient rats is associated with increased plasma triglyceride concentration and increased oxidative stress.

The purpose of this study was to assess whether dietary carbohydrate could differentially influence the consequences of magnesium deficiency with particular emphasis on lipid metabolism and oxidative stress. Rats were fed a sucrose based or starch based diet either adequate or deficient in magnesium for two weeks. Magnesium deficient rats, as compared with rats fed magnesium adequate diets, displayed the usual decrease in plasma magnesium concentration. The classic symptoms of inflammation including hyperaemia, increased number of blood leukocytes and enlarged spleen weight were observed in these rats. Plasma TG and plasma apo B concentrations were also significantly increased. In addition, magnesium-deficient animals presented an increased susceptibility to lipid peroxidation of heart and liver tissues as shown by TBARS concentration. Regardless of magnesium status, sucrose feeding did not affect the magnesium plasma level and inflammatory parameters. Feeding rats the sucrose diets induced hypertriglyceridaemia and increased plasma apo B concentration. Heart and liver susceptibility to lipid peroxidation were significantly increased in rats fed the sucrose diets as compared with those fed the starch diets. Sucrose feeding in magnesium deficient rats was associated with higher plasma triglycerides concentration and higher tissue susceptibility to peroxidation as compared with magnesium deficient rats fed the starch diet. The results emphasised the potential detrimental and additional effect of sucrose feeding and magnesium deficiency on cardiovascular risk. Since the intake of magnesium has been reduced appreciably in industrialised countries while fructose consumption has been rapidly increased, the impact of this eating pattern should be clarified in humans.     

Cholesterol metabolism in squirrel monkeys: Analysis of long-term kinetic studies in plasma and body tissues

Thirty adult male Brazilian-type squirrel monkeys (Saimiri sciureus) were fed a semipurified diet containing 45% of the calories as fat. One milligram of cholesterol per calorie was added to the diet of one-half of the animals. After 4 months on their respective diets, a single intravenous dose of [³H] cholesterol was administered to each animal. At 2, 4, and 6 months following intravenous labeling, five animals from the cholesterol-fed group and five animals from the non-cholesterol-fed group were killed for analysis of tissue cholesterol specific activity. The analysis of ratios between cholesterol specific activity of various tissues to the specific activity of terminal blood samples of the cholesterol-fed group indicated to us that cholesterol in liver, spleen, adrenal gland, ileum, stomach, kidney, testes, lung, heart, skin, tendon, fascia, and skeletal muscle had reached equilibrium with plasma cholesterol radio-activity by 2 months. The tissue/plasma ratios of adipose tissue, aorta, and gallstones demonstrated a slower rate of exchange. In the non-cholesterol-fed group the tissue/plasma ratios of all the tissues mentioned above remained close to unity in all three subgroups, indicating equilibration with plasma by 2 months. As before, the tissue/plasma ratios in adipose tissue and aorta had a slower rate of exchange. In both groups, cerebrum and spinal cord had a very slow rate of exchange with plasma cholesterol. Computer analysis of the plasma cholesterol specific activity time curve suggested to us a significantly better three-exponential fit rather than a two-exponential fit in $\text{5}\text{14}$ of the cholesterol-fed group and in none of the non-cholesterol-fed group. The minimal mass of the miscible body pool of cholesterol derived from kinetic analysis agreed reasonably well with, but consistently overestimated, the whole-body miscible pool determined by carcass analysis in both groups.