Repeated amiodarone exposure in the rat: toxicity and effects on hepatic and extrahepatic monooxygenase activities

Amiodarone is a potent and efficacious antiarrhythmic agent, yet associated with its use are life-threatening pulmonary fibrosis and hepatotoxicity. We have investigated the susceptibility of the male Sprague-Dawley rat to pulmonary and hepatic toxicity after repeated exposure to amiodarone and the effects of such exposure on hepatic and extrahepatic drug metabolizing enzymes. Animals received amiodarone (200 mg.kg-1.day-1 i.p., 5 days/week) for 1 week followed by 150 mg.kg-1.day-1 (5 days/week) for 3 additional weeks. No signs of pulmonary fibrosis or hepatotoxicity were observed, based on histological examination, lung hydroxyproline content, and plasma alanine aminotransferase activity. Analysis of tissues revealed extensive accumulation of amiodarone and desethylamiodarone in lung and liver, but concentrations were significantly lower in animals treated for 4 weeks than for 1 week. In a separate experiment, rats received amiodarone 150 mg.kg-1.day-1 i.p. (5 days/week) for 1 or 4 weeks. No differences in tissue concentrations of amiodarone and desethylamiodarone were detected between animals treated for 1 or 4 weeks. This regimen did not affect hepatic or extrahepatic monooxygenase activities. These results indicate that, in the male Sprague-Dawley rat, there is no observable pulmonary or hepatic toxicity following short-term amiodarone exposure, and there is enhanced elimination of amiodarone and desethylamiodarone when the daily dose of amiodarone is decreased after 1 week from 200 to 150 mg/kg.     

Placental transfer of halogenated benzenes (pentachloro-, pentachloronitro-, and hexabromo-) in rats.

The present study was carried out to provide information on the placental transfer of three organohalogens of environmental concern. Pentachloro-, pentachloronitro-, and hexabromobenzene were administered per os to rats daily on days 6 through 15 of gestation at level of 40, 100, and 200 mg/kg body weight. On day 22, the dams were killed and fetuses removed by caesarean section. Maternal brain, heart, kidney, liver, spleen and adipose tissue as well as whole fetus, fetal liver and fetal brain were analyzed for organohalogen residue by GLC. Pentachlorobenzene accumulated in the fetus to a greater extent than hexabromobenzene. In maternal tissues pentachlorobenzene accumulated to the greatest extent in adipose tissue, followed by liver, spleen, brain, heart and kidney. With hexabromobenzene, the greatest accumulation was observed in adipose tissue, followed by spleen, liver, heart, kidney and brain. Pentachloronitrobenzene was not detected (0.05 p.p.m.) in any maternal or fetal tissue.     

Effects of desethylamiodarone on thyroid hormone metabolism in rats: comparison with the effects of amiodarone

Desethylamiodarone is the principal metabolite of amiodarone. Amiodarone is a class III antiarrhythmic agent, which acts by lengthening repolarization in the myocardium, an effect that is identical to that produced by hypothyroidism. Amiodarone is known to alter thyroid hormone metabolism, and it has been suggested that the mechanism underlying its antiarrhythmic action is the induction of a myocardial but not generalized hypothyroidism. Since the serum levels of desethylamiodarone reach those of the parent compound during chronic amiodarone therapy, it has been suggested that at least part of amiodarone's pharmacological effects may be attributable to the additive effects of the metabolite. Therefore, we investigated the effects of desethylamiodarone on thyroid hormone metabolism and compared them with those of amiodarone in rats. We have shown that chronic treatment with desethylamiodarone decreased serum T3, markedly increased serum reverse T3 with no significant change in serum T4. These effects are similar to those of amiodarone. The data suggest that the chronic effects of amiodarone on thyroid hormone metabolism may be due at least in part to the actions of desethylamiodarone.     

Distribution of glutamine synthetase and an inverse relationship between glutamine synthetase expression and intramuscular glutamine concentration in the horse

Glutamine plays important roles in the interorgan transport of nitrogen, carbon and energy but little is known about glutamine metabolism in the horse. In this study we determined the tissue distribution of glutamine synthetase expression in three Standardbred mares. Expression of glutamine synthetase was highest in kidney and mammary gland, and relatively high in liver and adipose tissue. Expression was lower in gluteus muscle, thymus, colon and lung, and much lower in small intestine, pancreas and uterus. The pattern of glutamine synthetase expression in the horse is similar to that of other herbivores and it is likely that skeletal muscle, liver, adipose tissue and lungs are the major sites of net glutamine synthesis in this species. Expression did not differ between adipose tissue depots but did vary between different muscles. Expression was highest in gluteus and semimembranous muscles and much lower in diaphragm and heart muscles. The concentration of intramuscular free glutamine was inversely correlated with expression of glutamine synthetase (r=-0.81, p=0.0017). The concentration of free glutamine was much higher in heart muscle (21.6+/-0.9 micromol/g wet wt) than in gluteus muscle (4.19+0.33 micromol/g wet wt), which may indicate novel functions and/or regulatory mechanisms for glutamine in the equine heart.     

Antibody biodistribution coefficients

Tissue vs. plasma concentration profiles have been generated from a physiologically-based pharmacokinetic model of monoclonal antibody (mAb). Based on the profiles, we hypothesized that a linear relationship between the plasma and tissue concentrations of non-binding mAbs could exist; and that the relationship may be generally constant irrespective of the absolute mAb concentration, time, and animal species being analyzed. The hypothesis was verified for various tissues in mice, rat, monkey, and human using mAb or antibody-drug conjugate tissue distribution data collected from diverse literature. The relationship between the plasma and various tissue concentrations was mathematically characterized using the antibody biodistribution coefficient (ABC). Estimated ABC values suggest that typically the concentration of mAb in lung is 14.9%, heart 10.2%, kidney 13.7%, muscle 3.97%, skin 15.7%, small intestine 5.22%, large intestine 5.03%, spleen 12.8%, liver 12.1%, bone 7.27%, stomach 4.98%, lymph node 8.46%, adipose 4.78%, brain 0.351%, pancreas 6.4%, testes 5.88%, thyroid 67.5% and thymus is 6.62% of the plasma concentration. The validity of using the ABC to predict mAb concentrations in different tissues of mouse, rat, monkey, and human species was evaluated by generating validation data sets, which demonstrated that predicted concentrations were within 2-fold of the observed concentrations. The use of ABC to infer tissue concentrations of mAbs and related molecules provides a valuable tool for investigating preclinical or clinical disposition of these molecules. It can also help eliminate or optimize biodistribution studies, and interpret efficacy or toxicity of the drug in a particular tissue.     

Distribution of apolipoprotein A-I, C-II, C-III, and E mRNA in fetal human tissues. Time-dependent induction of apolipoprotein E mRNA by cultures of human monocyte-macrophages

Recently developed molecular probes for human apolipoprotein (apo) genes have been used to study the specificity of human tissue expression of the apo A-I, apo C-II, apo C-III, and apo E genes. We have found that apo E mRNA was present in all tissues examined. On the basis of total RNA concentration the relative abundance of apo E mRNA expressed as a percentage of the liver value is as follows: adrenal gland and macrophages, 74-100%; gonads and kidney, 12-15%; spleen, brain, thymus, ovaries, intestine, and pancreas, 3-9%; heart, 1.5%; stomach, striated muscle, and lung, less than 1%. The relative concentration of apo E mRNA in cultures of human peripheral blood monocyte-macrophages increases dramatically as a function of time in culture, and after 5 days, it compares to that of liver. The human tissues shown to synthesize apo E mRNA were also examined for their ability to synthesize apo A-I, apo C-II, and apo C-III mRNA. The relative abundance of apo A-I, apo C-III, and apo C-II mRNA expressed as a percentage of the liver value is as follows: apo A-I, intestine, 50%; apo A-I, pancreas and gonads, 12%; apo A-I, kidney, 4%; apo A-I, adrenal, 2.5%; apo A-I, ovaries and heart, 1%; apo A-I, stomach and thymus, less than 1%; apo C-III, intestine, 62%; apo C-III, pancreas, 7%; apo C-II, intestine, 3%; apo C-II, pancreas, less than 1%. The knowledge of tissue specificities in the synthesis of apolipoproteins is important for our understanding of the regulation of apolipoproteins and lipoprotein metabolism.     

Immunochemical characteristics of the peroxidases of several mouse tissues]

Enzymes catalyzing peroxidase reaction of a lysosomal fraction in bone marrow, leucocytes, spleen, thyroid gland, stomach, kidney, heart, lungs, brain and skeletal muscle of mice were investigated by immunochemical methods. A high level of peroxidase activity was discovered in leucocytes, bone marrow, spleen, heart and lung, a lower activity appeared to be characteristic of liver, thyroid gland and kidney. The peroxidase activities in brain, skeletal muscle and stomach were low. The reaction of immunoprecipitation with myeloperoxidase-specific antiserum revealed considerable antigenic distinctions between the enzymes catalysing peroxidase reaction in various tissues of mice.     

Pathological effects of the radiation protector WR-151327 in mice

The systemic effects of the radiation protective agent, S-3-(3-methylaminopropylamino) propylphosphorothioic acid (WR-151327), were studied in unirradiated B6CF1 male mice. Fifty mice were injected intraperitoneally with 540 mg/kg WR-151327, and groups of five mice were sacrificed at 14-day intervals up to and including 140 days post-treatment. Ten mice served as sham-injected controls. A necropsy was performed and gross morphological abnormalities were noted. Tissues (brain, eyes, harderian gland, salivary glands, sternal bone marrow, thyroid, lung, thymus, esophagus, trachea, skeletal muscle, heart, liver, kidney, adrenal gland, spleen, small intestine, pancreas, and testes) were fixed in 10% formalin, embedded in paraffin, and sectioned. Slides were routinely stained with hematoxylin and eosin while Alizarin red stain was used to test specifically for the presence of calcium salts. Histopathological effects of WR-151327 were restricted to the testes, salivary gland, and pancreas. The caudal pole of the testes was observed to undergo progressive changes from coagulation necrosis to dystrophic calcification. The cells of the submandibular salivary gland showed mainly hyperchromatic nuclei while the pancreas showed enlarged islets of Langerhans.     

Lipoprotein lipase and hepatic lipase mRNA tissue specific expression, developmental regulation, and evolution

Lipoprotein lipase (LPL) and hepatic lipase (HL) enzyme activities were previously reported to be regulated during development, but the underlying molecular events are unknown. In addition, little is known about LPL evolution. We cloned and sequenced a complete mouse LPL cDNA. Comparison of sequences from mouse, human, bovine, and guinea pig cDNAs indicated that the rates of evolution of mouse, human, and bovine LPL are quite low, but guinea pig LPL has evolved several times faster than the others. 32P-Labeled mouse LPL and rat HL cDNAs were used to study lipase mRNA tissue distribution and developmental regulation in the rat. Northern gel analysis revealed the presence of a single 1.87 kb HL mRNA species in liver, but not in other tissues including adrenal and ovary. A single 4.0 kb LPL mRNA species was detected in epididymal fat, heart, psoas muscle, lactating mammary gland, adrenal, lung, and ovary, but not in adult kidney, liver, intestine, or brain. Quantitative slot-blot hybridization analysis demonstrated the following relative amounts of LPL mRNA in rat tissues: adipose, 100%; heart, 94%; adrenal, 6.6%; muscle, 3.8%; lung, 3.0%; kidney, 0%; adult liver, 0%. The same quantitative analysis was used to study lipase mRNA levels during development. There was little postnatal variation in LPL mRNA in adipose tissue; maximal levels were detected at the earliest time points studied for both inguinal and epididymal fat. In heart, however, LPL mRNA was detected at low levels 6 days before birth and increased 278-fold as the animals grew to adulthood.(ABSTRACT TRUNCATED AT 250 WORDS)     

Levels and distribution of zinc,copper, magnesium,and calcium in rats fed different levels of dietary zinc

Effects of altered dietary zinc on levels of zinc, copper, magnesium, and calcium in organ and peripheral tissues were studied. When rats fed a zinc-deficient diet (1.3 μg Zn/g) for 28 d were compared with rats fed a control diet (37.5 μg Zn/g), levels of zinc were slightly lower in plasma, hair, and skin and 50% lower in femur and pancreas, whereas the levels of copper were higher in all tissue except plasma. Magnesium levels were higher than controls in the heart and lower in the spleen, whereas the calcium levels were lower in plasma, lung, spleen, kidney, and skin and strikingly higher in brain, hair, and femur. When rats fed a zinc-supplemented diet (1.0 mg Zn/g) were compared to the same conrols, levels of zinc in these were higher in all organs and peripheral tissues studied, except heart, lung, and liver; copper levels were higher in liver, kidney, and spleen; magnesium levels were significantly higher in the spleen, but were little affected in other tissues, although calcium levels were higher in pancreas, spleen, kidney, and skin and lower in plasma and hair. These data indicate that overall copper organ and peripheral tissue levels are affected inversely, and zinc and calcium levels directly, by zinc nutriture.     

Mechanisms of digoxin-amiodarone interaction in the rat

Amiodarone and digoxin are often used in combination and clinical experience suggests that amiodarone may increase serum digoxin levels and toxicity. We have investigated the influence of amiodarone on digoxin pharmacokinetics and tissue distribution in the rat. Forty-nine rats were injected with 10 mg/kg amiodarone sc three times a day for 7 days, while 49 others were injected with saline only. On the eighth day, all the rats received 0.5 mg/kg digoxin ip; 4, 5, 6, 7, 8, 10, and 12 hr later, groups of 7 amiodarone-pretreated and control animals were sacrificed, and plasma, heart, liver, muscle, brain, and kidney digoxin concentrations measured by radioimmunoassay. Data were analyzed by two-way ANOVA, with group comparisons using the Waller-Duncan multiple comparison procedure. Digoxin levels were significantly higher in the plasma, heart, muscle, and kidney of the amiodarone-pretreated rats at most points of measurement (P less than 0.05) whereas liver digoxin levels were elevated at 8, 10, and 12 hr. Kidney/plasma, heart/plasma, muscle/plasma, and especially liver/plasma ratios in the control groups significantly exceeded the values found in the amiodarone-pretreated group at most time points. Concentrations of digoxin in brain were not changed. This suggests that the volume of distribution is significantly altered in the amiodarone-pretreated group. Amiodarone increases plasma digoxin levels in rats as it does in humans, but the mechanism is unclear.     

Model to evaluate the toxic effect of drugs: vincristine effect in the mass of organs and in the distribution of radiopharmaceuticals in mice

There are evidences that the biodistribution of radiopharmaceuticals can be modified by some drugs. As chemotherapeutic drugs present important toxic effects, we studied the vincristine effect in the mass of organs and are trying to develop a model to evaluate the action of chemotherapeutic drug using the biodistribution of radiopharmaceuticals. Vincristine was administered (n=15) into female Balb/c mice, the organs isolated and their mass determined. To study the vincristine effect in the biodistribution of technetium-99m-dimercaptosuccinic acid (99mTc-DMSA) or technetium-99m-diethylenetriaminepentaacetic acid (99mTc-DTPA), vincristine (0.03 mg) was administered in the animals (n=15) in three doses. 99mTc-DMSA or 99mTc-DTPA was injected 1h after the last dose. After 0.5h, the animals were sacrificed and the percentage of radioactivity (%ATI) and the percentage of radioactivity per gram of tissue (%ATI/g) in each organ were calculated. The results have shown that the mass decreased significantly (Wilcoxon test, P<0.05) in thymus, spleen, ovary, uterus, kidneys, pancreas. The %ATI to 99mTc-DMSA increased in lung, pancreas, heart, thyroid, brain, and bone, and the %ATI/g increased in uterus, ovary, spleen, thymus, kidney, lung, liver, pancreas, heart, thyroid, brain and bone. To 99mTc-DTPA, the %ATI increased in uterus, ovary, spleen, thymus, kidney, lung, liver, stomach, heart and bone, and the %ATI/g increased in uterus, ovary, spleen, thymus, kidney, lung, liver, stomach, heart and bone. The results were statistically significant (Wilcoxon test). The results can be explained by the metabolization, therapeutic, toxicological or immunosupressive action of the vincristine. This model, probably, should be used to evaluate the toxic effect of various drugs.     

Glutathione reductase activity and flavin concentration in guinea-pig tissues.

Glutathione reductase (GR) activity and flavin concentration were studied in systemic tissues (brain, heart, lung, liver, spleen, stomach, pancreas, muscle, kidney, testis) and blood components (erythrocytes and plasma) from male guinea-pigs. GR activity and the flavin concentration were high in kidney and liver, and low in muscle. GR activity in erythrocytes was found in a range of tissues, but flavin concentration in erythrocytes was lower than in any tissues. GR was saturated with flavin adenine dinucleotide (FAD) in almost all tissues, but not in muscle or erythrocytes.     

SCN efferents to peripheral tissues: implications for biological rhythms.

The suprachiasmatic nucleus (SCN) is the principal generator of circadian rhythms and is part of an entrainment system that synchronizes the animal with its environment. Here, we review the possible communication of timing information from the SCN to peripheral tissues involved in regulating fundamental physiological functions as revealed using a viral, transneuronal tract tracer, the pseudorabies virus (PRV). The sympathetic nervous system innervation of the pineal gland and the sympathetic outflow from brain to white adipose tissue were the first demonstrations of SCN-peripheral tissue connections. The inclusion of the SCN as part of these and other circuits was the result of lengthened postviral injection times compared with those used previously. Subsequently, the SCN has been found to be part of the sympathetic outflow from the brain to brown adipose tissue, thyroid gland, kidney, bladder, spleen, adrenal medulla, and perhaps the adrenal cortex. The SCN also is involved in the parasympathetic nervous system innervation of the thyroid, liver, pancreas, and submandibular gland. Individual SCN neurons appear connected to more than one autonomic circuit involving both sympathetic and parasympathetic innervation of a single tissue, or sympathetic innervation of two different peripheral tissues. Collectively, the results of these PRV studies require an expansion of the traditional roles of the SCN to include the autonomic innervation of peripheral tissues and perhaps the modulation of neuroendocrine systems traditionally thought to be controlled solely by hypothalamic stimulating/inhibiting factors.     

Neuropeptide control of thyroid blood flow and hormone secretion in the rat

The effects of peptide HI (PHI), neuropeptide Y (NPY), and substance P (SP) on thyroid blood flow and hormone levels were studied in anesthetized rats. Regional blood flows were determined using radioactive microspheres. No change in heart rate or mean left ventricular pressure occurred during these neuropeptide infusions (0.625 micrograms iv over 2 min). PHI treatment resulted in a four-fold increase in thyroid blood flow. Blood flows to the pancreas and salivary gland also increased during PHI treatment. Infusions of NPY or SP did not significantly alter thyroid blood flow. However, SP decreased blood flow to the spleen and small intestine. These neuropeptides had no effect on blood flows to the adrenal, kidney, brain, heart, and adipose tissues. Following PHI, NPY, and SP infusions, plasma triiodothyronine and thyroxine levels were not different from values in saline-treated rats. This study demonstrates that PHI, like vasoactive intestinal peptide, is a potent thyroidal vasodilator at a dose that does not affect circulating thyroid hormone secretion.     

Tissue distribution and molecular heterogeneity of bovine thiol:protein-disulphide oxidoreductase (disulphide interchange enzyme)

1. The distribution of thiol:protein-disulphide oxidoreductase (disulphide interchange enzyme) in 17 bovine tissue extracts was determined by rocket immunoelectrophoresis and by measuring the reductive cleavage of insulin. 2. The relative concentration (per mg total protein) was found to be in the order: Pancreas greater than liver greater than lymph node greater than testes, fat tissue greater than parotid gland, brain, spleen, lung greater than small intestine, spinal cord, large intestine, kidney greater than paunch, aorta greater than skeletal muscle greater than heart. 3. The distribution of specific activity showed a similar pattern, irrespectively of whether glutathione or L-cysteine was used as cosubstrate. 4. The concentration varied 200-fold and the specific activity 400-fold between pancreas and heart muscle, respectively. 5. Crossed immunoelectrophoresis demonstrated that a fast-migrating form of the enzyme was the only one present in almost all tissues, but 15% of the enzyme in liver was a slow-migrating form and 50% in heart muscle a medium-migrating form. 6. The lung contains a species having partial immunological identity to the enzyme. 7. Purified enzyme from bovine liver has a somewhat lower mobility than the fast-migrating form in extract. 8. The results seem to support the general view that the enzyme is involved in synthesis of disulphide-bonded extracellular proteins, although the presence of the enzyme in tissues like fat, brain, spinal cord, skeletal muscle and heart indicates other cellular functions as well.     

野生成年树鼩主要脏器重量及脏器系数的测定分析

目的对野生成年树鼩的体重和主要脏器重量进行测定,计算其脏器系数。方法测定60只野生成年树鼩体重及11个主要脏器重量,并计算其脏器系数。进行脏器重量、脏器系数的性别间比较分析及Kendall和谐系数分析。结果性别间比较心、肺重量差异极显著（P〈0.01）,脑、肾上腺、胰腺重量之间差异显著（P〈0.05）;肾上腺、胰腺系数差异有极显著性（P〈0.01）,心、肺、肾系数之间差异均达到了显著水平（P〈0.05）。Kendall和谐系数（W）分析表明,动物与其个体各主要器官整体发育协调性较好。结论野生成年树鼩心重量、肺重量、脑重量、肾上腺重量、胰腺重量、肾上腺系数、胰腺系数、心系数、肺系数、肾系数性别间存在差异。     

Effects of aging on cardiac output, regional blood flow, and body composition in Fischer-344 rats

The purpose ofthis study was to determine the effects of maturation and aging oncardiac output, the distribution of cardiac output, tissue blood flow(determined by using the radioactive-microsphere technique), and bodycomposition in conscious juvenile (2-mo-old), adult (6-mo-old), andaged (24-mo-old) male Fischer-344 rats. Cardiac output was lower injuvenile rats (51 ± 4 ml/min) than in adult (106 ± 5 ml/min) oraged (119 ± 10 ml/min) rats, but cardiac index was not differentamong groups. The proportion of cardiac output going to most tissuesdid not change with increasing age. However, the fraction of cardiacoutput to brain and spinal cord tissue and to skeletal muscle wasgreater in juvenile rats than that in the two adultgroups. In addition, aged rats had a greater percentcardiac output to adipose tissue and a lower percent cardiac output tocutaneous and reproductive tissues than that in juvenile and adultrats. Differences in age also had little effect on mass-specificperfusion rates in most tissues. However, juvenile rats had lower flowsto the pancreas, gastrointestinal tract, thyroid and parathyroidglands, and kidneys than did adult rats, and aged rats had lower flowsto the white portion of rectus femoris muscle, spleen, thyroid andparathyroid glands, and prostate gland than did adult rats. Body massof juvenile rats was composed of a lower percent adipose mass and agreater fraction of brain and spinal cord, heart, kidney, liver, andskeletal muscle than that of the adult and aged animals. Relative tothe young adult rats, the body mass of aged animals had a greaterpercent adipose tissue mass and a lower percent skeletal muscle andskin mass. These data demonstrate that maturation and aging have asignificant effect on the distribution of cardiac output but relativelylittle influence on mass-specific tissue perfusion rates in conscious rats. The old-age-related alterations in cardiac output distribution toadipose and cutaneous tissues appear to be associated with theincreases in percent body fat and the decreases in the fraction of skinmass, respectively, whereas the decrease in the portion of cardiacoutput directed to reproductive tissue of aged rats appears to berelated to a decrease in mass-specific blood flow to the prostate gland.