Pharmacokinetics and hepatoprotective effects of 2-methylaminoethyl-4,4'-dimethoxy-5 ,6,5',6'dimethylenedioxybiphenyl-2-carboxylic acid-2'-carboxylate monohydrochloride in rats with CCl4-induced acute hepatic failure

The pharmacokinetics and hepatoprotective effects of 2-methylaminoethyl-4,4'-dimethoxy-5,6,5',6'-dimethylenedioxybip henyl-2-carboxylic acid-2'-carboxylate monohydrochloride (DDB-S) have been investigated in rats with CCl4-induced acute hepatic failure. To study the pharmacokinetics of DDB-S, rats were divided into a control group and a CCl4-intoxicated group. DDB-S 50 mg kg(-1) was administered by intravenous bolus injection to both groups of rats. In the CCl4-intoxicated rats the plasma concentrations of DDB-S were significantly higher, the area under the plasma concentration-time curve from time zero to time infinity was significantly greater (6-46 vs 3.34 mg min mL(-1)), and the total body (7.74 vs 15.0 mL min(-1) kg(-1)), renal (2.55 vs 5.10 mL min(-1) kg(-1)), nonrenal (5.07 vs 9.65 mL min(-1) kg(-1)), and biliary (1.48 vs 2.69 mL min(-1) kg(-1)) clearances were significantly slower compared with the control rats. This could be due to decreased hepatic cytochrome P450 activity and impaired kidney function induced by CCl4. To study the hepatoprotective effects of DDB-S, rats were divided into three groups, control rats and CCl4-intoxicated rats with or without DDB-S pretreatment (50 mg kg(-1) i.p.). The effects of DDB-S pretreatment on CCl4-induced liver injury were considerable; the serum levels of alanine transaminase, aspartate transaminase, and alkaline phosphatase were significantly lower by 54.3, 44.6 and 67.2%, respectively, compared with the CCl4-intoxicated-only group. In an in-vitro study, rat hepatocytes were exposed to fresh medium containing 10 mM CCl4 and various concentrations of DDB-S (10 or 100 microg mL(-1)). The levels of alanine transaminase and aspartate transaminase in the medium were measured as an indicator of hepatocyte injury. DDB-S dose-dependently decreased the levels of alanine transaminase and aspartate transaminase compared with CCl4-intoxication only. These results indicate that DDB-S has hepatoprotective activity.     

Characterization of the selectivity and mechanism of cytochrome P450 inhibition by dimethyl-4,4'-dimethoxy-5,6,5',6'-dimethylenedioxybiphenyl-2,2'-dicarboxylate.

In vitro studies with human liver microsomes and cytochrome P450 (P450) prototype substrates were performed to characterize the selectivity and mechanism of inhibition of P450 by dimethyl-4,4'-dimethoxy-5,6,5',6'-dimethylenedioxybiphenyl-2,2'-dicarboxylate (DDB). DDB was found to be a strong inhibitor of testosterone 6beta-hydroxylation activity (CYP3A4) with a K(i) value of 0.27 +/- 0.21 microM. At higher concentrations, DDB marginally inhibited caffeine N(3)-demethylation (CYP1A2), diclofenac 4'-hydroxylation (CYP2C9), and dextromethorphan O-demethylation (CYP2D6) activities, but this compound had no effect on CYP2A6-, CYP2C19-, and CYP2E1-mediated reactions. Spectral analysis indicated that the formation of metabolite-P450 complex having absorbance at 456 nm was concentration-dependent; 5 to 33% of the total P450 was complexed in rat and human liver microsomes after a 5-min incubation with DDB. In addition, microsomal incubations with DDB in the presence of NADPH resulted in a loss of spectral P450 content, which was restored after adding K(3)Fe(CN)(6). This complex formation resulted in a time-dependent loss of CYP3A-catalyzed marker activity (testosterone 6beta-hydroxylation) in human liver microsomes. The inhibition was only partially restored upon dialysis. These results collectively suggest that formation of a metabolite-CYP3A complex with DDB was responsible for the CYP3A-selective time-dependent loss of catalytic function of CYP3A.     

Antitumor agents. 293. Nontoxic dimethyl-4,4'-dimethoxy-5,6,5',6'-dimethylenedioxybiphenyl-2,2'-dicarboxylate (DDB) analogues chemosensitize multidrug-resistant cancer cells to clinical anticancer drugs

Novel dimethyl-4,4'-dimethoxy-5,6,5',6'-dimethylenedioxybiphenyl-2,2'-dicarboxylate (DDB) analogues were designed and synthesized to improve their chemosensitizing action on KBvin (vincristine-resistant nasopharyngeal carcinoma) cells, a multidrug resistant cell line overexpressing P-glycoprotein (P-gp). Structure-activity relationship analysis showed that aromatic and bulky aliphatic side chains at the 2,2'-positions effectively and significantly sensitized P-gp overexpressing multidrug resistant (MDR) cells to anticancer drugs, such as paclitaxel (TAX), vincristine (VCR), and doxorubicin (DOX). DDB derivatives 16 and 23 showed 5-10 times more effective reversal ability than verapamil (VRP) for TAX and VCR. Analogue 6 also exhibited five times greater chemosensitizing effect against DOX than VRP. Importantly, no cytotoxicity was observed by the active DDB analogues against both non-MDR and MDR cells, suggesting that DDB analogues serve as novel lead compounds for the development of chemosensitizers to overcome the MDR phenotype. The mechanism of action studies demonstrated that effective inhibition of P-glycoprotein by DDB analogues dramatically elevated the cellular concentration of anticancer drugs.     

Assessment of changes in liver blood flow after food intake—comparison of ICG clearance and echo-Doppler

Echo-Doppler measurements of portal venous blood flow in intrahepatic branches and indocyanine green (ICG) clearance after continuous i.v. infusion as measure for liver blood flow were compared to evaluate the increase in splanchnic blood flow after food intake. It was shown that both methods assessed the changes in flow in a similar manner. Changes in blood flow in intrahepatic portal vein branches measured with echo-Doppler adequately predicted the change in ICG concentrations. Hence, echo-Doppler measurements of hepatic portal blood flow in intrahepatic branches can be used to estimate changes in total liver blood flow.     

Pharmacokinetic interaction between oltipraz and dimethyl-4,4'-dimethoxy-5,6,5',6'-dimethylene dioxybiphenyl-2,2'-dicarboxylate (DDB) after single intravenous and oral administration to rats

The aim of this study was to report the pharmacokinetic interaction between oltipraz (50 mg kg(-1)) and dimethyl-4,4'-dimethoxy-5,6,5',6'-dimethylene dioxybiphenyl-2,2'-dicarboxylate (DDB, 10 mg kg(-1)) after single intravenous and oral administration to rats. After intravenous administration of oltipraz plus DDB, the area under the plasma concentration-time curve from time zero to time infinity (AUC) of oltipraz was significantly greater (1440 vs 1740 microg min mL(-1)) than that after oltipraz alone. This was not due to slower clearances of oltipraz after oltipraz plus DDB since the total body, renal and nonrenal clearances were comparable between the two groups of rats. It could be due to a decrease in tissue binding of oltipraz by DDB. The apparent volume of distribution at steady state (Vd(ss)) of DDB was significantly smaller (7060 vs 4650 mL kg(-1)) than after oltipraz alone. After oral administration of oltipraz plus DDB, the AUC of olitpraz was also significantly greater (479 vs 583 microg min mL(-1)) than that after oltipraz alone. This was not due to increased absorption of oltipraz from the rat gastrointestinal tract after oltipraz plus DDB but again could be due to a decrease in Vd(ss) of oltipraz by DDB. However, after both intravenous and oral administration, the pharmacokinetic parameters of DDB were comparable between DDB alone and DDB plus oltipraz, indicating that oltipraz did not greatly affect the pharmacokinetics of DDB in rats.     

The plasma clearance of indocyanine green in rats with acute renal failure: Effect of dose and route of administration

The pharmacokinetics of various doses (1–7.5 mg/kg i.v.) of indocyanine green (ICG) have been studied in control rats and rats with glycerol-induced acute renal failure (ARF). The pharmacokinetic changes seen at a dose of 1 mg/kg, after jugular vein administration, were significant decreases in uraemic rats in the rate of entry of ICG into the liver (k₁₂) and in the rate of movement of dye from liver to plasma (k₂₁). Greater and more numerous changes in pharmacokinetic parameters were recorded in experiments conducted using 4.0 and 7.5 mg/kg ICG. The results from these experiments showed that in addition to significant decreases in k₁₂ and k₂₁ there was a significant reduction in the rate constant for transfer of dye from liver to bile (k₂₃). These changes were accompanied by a significant decrease in plasma clearance.In a separate series of experiments steps were taken to reduce the degree of uraemia produced by glycerol injection. The findings from these experiments showed no significant pharmacokinetic differences between control and mildly uraemic animals after administration of a dose of 7.5 mg/kg ICG. This suggests that the kinetic changes described above were a consequence of renal failure and not a direct hepato-toxic effect of glycerol.     

Validated stability-indicating HPLC method for the determination of dimethyl-4,4'-dimethoxy-5,6,5',6'-dimethylene dioxybiphenyl-2,2'-dicarboxylate (DDB) and its degradation products

High-performance liquid chromatographic method was developed for the quantitative determination of dimethyl-4,4'-dimethoxy-5,6,5',6'-dimethylene dioxybiphenyl-2,2'-dicarboxylate (DDB) and its degradation products. Forced degradation studies were performed on bulk sample of DDB using acid (1N hydrochloric acid), alkaline (0.1N sodium hydroxide), oxidation (0.33% hydrogen peroxide), heat (70 degrees C) and photolytic degradation. The chromatographic method was fine tuned using the samples generated from forced degradation studies. Good resolution between the peaks corresponds to degradation products and the analyte was achieved on 5 microm ODS column (Luna, Phenomenex, USA). The mobile phase consists of a mixture of acetonitrile and water (60:40, v/v). Quantitation was achieved with UV detection at 235 nm based on peak area. The proposed HPLC method was utilized to investigate the kinetics of acidic, alkaline and oxidative degradation processes of DDB at different temperatures and the apparent pseudo first-order rate constant, half-life and activation energy were calculated. The pH-rate profiles of degradation of DDB in Britton-Robinson buffer solutions within the pH range 2-11 were studied. The developed method was validated with respect to linearity, accuracy, precision, robustness and forced degradation studies prove the stability-indicating power of the method.     

No effect of diltiazem on the hepatic clearance of indocyanine green in the rats

In order to investigate the effect of the pretreatment with various doses of diltiazem (DTZ) on the pharmacokinetics of indocyanine green (ICG) at steady state, especially the hepatic blood clearance due to the change of hepatic blood flow, the following experiments were carried out with ICG, a hepatic function test marker, not metabolized in liver and only excreted in bile. The intravenous bolus injection (3,780 μg/kg) and the constant-rate infusion (10, 100 μg/kg/hr) or ICG into the left femoral vein were made in order to check the steady-state plasma concentration (C _ss of 10 μg/ml) of ICG at 20, 25 and 30 min. Following a 90-min washout period, the intravenous bolus injection (108, 430, 860 and 1,720 μg/kg) and the constant-rate infusion (108, 433, 866 and 1,730 μg/kg/hr) of DTZ into the right femoral vein were made and the achievement of the steady-state plasma levels (C _ss of 50, 200, 400 and 800 ng/ml) of DTZ were conformed at 60, 70 and 80 min. During the steady state of DTZ, the intravenous bolus injection (3,780 μg/kg) and the constant-rate infusion (10,200 μg/kg/hr) of ICG into the left femoral vein were made and also the steady-state plasma concentration of ICG was checked at 20, 25 and 30 min. The plasma concentrations of DTZ and ICG were determined using a high performance liquid chromatographic technique. At the steady state, the hepatic blood clearance of ICG was obtained from the plasma concentration and blood-to-plasma concentration ratio (R _b ) of ICG. The pretreatment with various doses of DTZ did not influence the plasma concentrations,R _B and plasma free fraction (f _p ) of ICG. So the hepatic blood clearance of ICG was independent of concentration of DTZ. The hepatic blood clearance of ICG could be affected by both hepatic blood flow and hepatic intrinsic clearance. But there was no change of the hepatic blood clearance of ICG between the control and the DTZ-pretreated rats in this study. So it may be suggested that DTZ does not influence hepatic blood flow.     

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin on indocyanine green blood clearance in rhesus monkeys.

To assess effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on liver function adult male rhesus monkeys were treated with a single oral dose of acetone/corn oil (control) or 5, 25, or 75 micrograms/kg TCDD. Each monkey was used as its own control and indocyanine green (ICG) blood clearance and the following serum enzymes: glutamic pyruvate transaminase (SGPT), sorbitol dehydrogenase (SDH) and gamma glutamyl transpeptidase (gamma GTP), were measured at regular intervals for 4 weeks before and 17 weeks after treatment. In control monkeys ICG blood clearance and serum enzymes were similar before and after treatment. However, in the monkey that received 5 micrograms/kg TCDD there was a mild increase in ICG blood clearance followed by a slight decrease. The magnitude of this biphasic change was greater in monkeys that received 25 and 75 micrograms/kg TCDD and the decrease in clearance was invariably associated with a 1--2-week period before the monkeys died. SDH and SGPT activities were elevated at some time during the course of intoxication in all TCDD-treated monkeys but gamma GTP activity was not altered. The monkey treated with 5 micrograms/kg TCDD survived but monkeys treated with 25 and 75 micrograms/kg died 4--6 weeks after treatment. Light microscopy of the livers of TCDD-treated monkeys that died revealed fatty infiltration with minimal hepatocellular necrosis.     

The anti-fibrogenic effect of a pharmaceutical composition of [5-(2-pyrazinyl)-4-methyl-1,2-dithiol-3-thione] (oltipraz) and dimethyl-4,4'-dimethoxy-5,6,5',6'-dimethylene dioxybiphenyl-2,2'-dicarboxylate (DDB)

Liver fibrosis is a prepathological state wherein damaged liver tissues in chronic liver diseases, such as hepatitis, are not repaired to normal tissues, but converted to fibrous tissue. 5-(2-Pyrazinyl)-4-methyl-1,2-dithiol-3-thione (oltipraz), a cancer chemopreventive agent, is effective against a wide variety of chemical carcinogens. Recently, we reported that oltipraz inhibits liver fibrogenesis (Kang et al., 2002). In the present study, the effects of oltipraz in combination with dimethyl-4,4'-dimethoxy-5,6,5',6'-dimethylene dioxybiphenyl-2,2'-dicarboxylate (DDB) on dimethylnitrosamine (DMN)-induced liver fibrogenesis were assessed in rats. Oltipraz (30 mg/kg body weight, p.o., 3 times per week for 4 weeks) was found to inhibit the increases in plasma ALT, AST and bilirubin by DMN, whereas DDB (30 mg/kg body weight, p.o., 3 times per week for 4 weeks) attenuated the increases in the plasma ALT and bilirubin. The lowered plasma protein and albumin contents in DMN-treated rats were completely restored by oltipraz, but not by DDB. DDB decreases liver cell injury and inflammation through inhibition of nuclear factor-kB. DMN increased the accumulation of liver collagen, as indicated by the increase in the 4-hydroxyproline content in liver homogenates, which was reduced by treatment with oltipraz, but not by DDB. Given the differential effect between oltipraz and DDB, the potential enhancement of antifibrotic efficacy by the drugs was assessed in the animal model. Despite the minimal effect of DDB on DMN-induced fibrogenesis, DDB (5-25 mg/kg), administered together with oltipraz (25-5 mg/kg), showed an additive protective effect against hepatotoxicity and fibrosis induced by DMN, which was shown by the blood chemistry parameters and histopathological analysis. The adequate composition ratio of oltipraz to DDB was 5:1. These results provide information on the pharmaceutical composition, comprising of oltipraz and DDB as the active components, for the treatment and/or prevention of liver fibrosis and cirrhosis.     

The influence of taurocholate and dehydrocholate choleresis on plasma disappearance and biliary excretion of indocyanine green in the rat

Summary The pharmacokinetics of indocyanine green (ICG; 3.9 moles/kg and 12.9 moles/kg) were investigated in rats given infusion of either saline, taurocholate (106 moles/h) or dehydrocholate (106 or 268 moles/h). During the infusion of saline and taurocholate the plasma concentration of ICG decreased in a mono-exponential manner. However, with dehydrocholate the clearance of ICG from plasma showed two phases with different half lives. The half life of the rapid component (2.2 min) was about the same as the one found in the control experiments.After injection of 12.9 moles/kg ICG the biliary excretion of the dye increased by 138% during taurocholate administration, while an equimolar dehydrocholate infusion resulted in a mean increament of 55%. Under these circumstances the bile flow was stimulated by 195% and 297% resp.With the lower dose of ICG (3.9 moles/kg) however, there was no stimulation of the biliary ICG excretion with taurocholate. At this dose level an infusion of dehydrocholate (106 mol/h) enchanced the biliary output of ICG by approximately 54%, while administration of 268 mol/h resulted in a slight but significant decrease of 31%.These observations can be explained by assuming interaction of the bile acids with the hepatic transport of ICG at different sites. The appearance of the second component of the plasma curve during dehydrocholate infusion is possibly related to a diminished hepatic storage capacity for ICG and is not due to an effect on the primary hepatic uptake or biliary output of the dye.     

Long-term pharmacokinetics of 2,2',4,4',5,5'-hexachlorobiphenyl (6-CB) in rats with constant adipose tissue mass

Long-term (280 days) pharmacokinetics of 2,2',4,4',5,5'-hexachlorobiphenyl (6-CB) was studied in rats with constant adipose tissue mass. This was achieved by feeding the animals 50% of their mean ad libitum food intake. 6-CB was administered as a single iv injection of 0.6 mg/kg. Tissues and excreta were analyzed at various time points from 4 to 280 days. After the redistribution phase, all tissue concentrations declined with terminal half-lives of 431-478 days, and concentration in adipose tissue was 1000 times higher than in blood. The corresponding ratios were: for skin 40, lung 30, liver 25, brain 10, and muscle 10. From day 4 on only adipose tissue, skin, and muscle contained significant amounts of 6-CB. Between 2 and 4 weeks adipose tissue and skin reached a maximum corresponding to 68 and 15% of the dose, respectively. After 280 days these values declined to 38 and 7% of the dose. Fecal excretion during this period was 43% of the dose with a terminal half-life of 478 days. Polar metabolites (1.5% of dose) were detectable in urine only. Extrapolation of fecal excretion kinetics yields a total excretion value of 99% of the dose at infinite time. Thus, in the rat with constant adipose tissue mass, 6-CB shows first order kinetics with reversible storage and total excretion. This is in sharp contrast with the situation of increasing adipose tissue, i.e. ad libitum feeding, which is characterized by irreversible storage in adipose tissue and limited excretion.     

Effect of 2,2',4,4'-tetrahydroxybenzophenone (BP2) on steroidogenesis in testicular Leydig cells

Endocrine disruptors (EDs) affect the function of animal reproductive systems. Recently, 2,2',4,4'-tetrahydroxybenzophenone (BP2), which is a component of UV protection products, was found to be an ED that interferes with the thyroid hormone (TH) axis. However, BP2 activity in the testis has not been well addressed. In this study, we have examined the effects of BP2 on steroidogenesis in testicular Leydig cells in connection with thyroid hormone signaling, which is known to play an important role in testicular development and function. Our study showed that BP2 affected the expression of steroidogenic enzyme genes in testicular Leydig cells, which is differentially regulated by thyroid hormone/thyroid hormone receptor (TR) signaling. In MA-10 Leydig cell line, TR/T3 signaling increased the expression of P450c17 and P450scc, while it decreased the expression of StAR and 3β-HSD. Interestingly, BP2 affected the expression of steroidogenic enzyme genes in a manner opposite to that of T3 signaling. BP2 downregulated the TRα/T3-activation of P450c17 and P450scc expression while enhancing the TRα/T3-repression of StAR and 3β-HSD expression. Transient transfection analyses with promoter-reporter constructs revealed that BP2 altered the expression of steroidogenic enzyme genes by affecting the cAMP and Nur77-activated promoter activity of P450c17, StAR, and 3β-HSD. Animal experiments with mice revealed that BP2 decreased the production of testosterone in the testis by affecting the expression of some steroidogenic enzyme genes in vivo. Together, these findings elucidate a molecular mechanism of BP2 action underlying testicular steroidogenesis and also suggest that BP2 acts, in part, as a thyroid antagonist that affects steroidogenesis in the testis.     

Tissue disposition, excretion and metabolism of 2,2',4,4',6-pentabromodiphenyl ether (BDE-100) in male Sprague-Dawley rats

The absorption, disposition, metabolism and excretion study of orally administered 2,2',4,4',6-pentabromodiphenyl ether (BDE-100) was studied in conventional and bile-duct cannulated male rats. In conventional rats, >70% of the radiolabelled oral dose was retained at 72 h, and lipophilic tissues were the preferred sites for disposition, i.e. adipose tissue, gastrointestinal tract, skin, liver and lungs. Urinary excretion of BDE-100 was very low (0.1% of the dose). Biliary excretion of BDE-100 was slightly greater than that observed in urine, i.e. 1.7% at 72 h, and glucuronidation of phenolic metabolites was suggested. Thiol metabolites were not observed in the bile as had been reported in other PBDE metabolism studies. Almost 20% of the dose in conventional male rats and over 26% in bile-duct cannulated rats was excreted in the faeces, mainly as the unmetabolized parent, although large amounts of non-extractable radiolabel were also observed. Extractable metabolites in faeces were characterized by mass spectrometry. Monohydroxylated pentabromodiphenyl ether metabolites were detected; mono- and di-hydroxylated metabolites with accompanying oxidative debromination were also observed as faecal metabolites. Tissue residues of [(14)C]BDE-100 in liver, gastrointestinal tract and adipose tissue contained only parent material. The majority of the 0-72-h biliary radioactivity was associated with an unidentified 79-kDa protein or to albumin.     

2',4',6'-Tris(methoxymethoxy) chalcone induces apoptosis by enhancing Fas-ligand in activated hepatic stellate cells

Suppression of hepatic stellate cell (HSC) activation and proliferation, and induction of apoptosis in activated HSCs have been proposed as therapeutic strategies for the treatment and prevention of the hepatic fibrosis. We previously showed that 2′,4′,6′-tris(methoxymethoxy) chalcone (TMMC), a synthesized chalcone derivative, inhibits platelet-derived growth factor-induced HSC proliferation at 5–20 μM. Here, we showed that TMMC induces apoptosis in activated HSCs at higher concentrations (30–50 μM), but is not cytotoxic to primary hepatocytes. Moreover, TMMC induces hyperacetylation of histone by inhibiting histone deacetylase (HDAC) in activated HSCs. Interestingly, TMMC treatment remarkably increased Fas-ligand (FasL) mRNA expression in a dose-dependent manner. Cycloheximide treatment reversed the induction of TMMC on apoptosis, indicating that de novo protein synthesis was required for TMMC-induced apoptosis in activated HSCs. In addition, FasL synthesis by TMMC is closely associated with maximal procaspase-3 proteolytic processing. In vivo, TMMC reduced activated HSCs in CCl₄-intoxicated rats during liver injury recovery, as demonstrated by α-smooth muscle actin expression in rat liver. TMMC treatment also resulted in apoptosis, as demonstrated by cleavage of poly(ADP-ribose) polymerase in rat liver. In conclusion, TMMC may have therapeutic potential by inducing HSC apoptosis for the treatment of hepatic fibrosis.     

Effect of 3,3',4,4'-tetrachlorobiphenyl and 2,2',4,4',5,5'-hexachlorobiphenyl on the induction of hepatic lipid peroxidation and cytochrome P-450 associated enzyme activities in rats

Polychlorinated biphenyls (PCBs) are environmental contaminants that have been widely used for various industrial purposes. In spite of numerous studies on PCBs, however, their mechanism of toxicity remains unknown. The role of cytochrome P-450 in PCBs induced hepatic lipid peroxidation is controversial. Therefore, the present study was undertaken to study the mechanism of action of two PCBs and their role in cytochrome P-450 induction and lipid peroxidation, determined in vivo and during the incubation of subcellular fractions. We also examined whether agonist/antagonist activities between the two PCBs were occurring. Two PCBs were studied: 3,3',4,4'-tetrachlorobiphenyl (PCB-77), a non-ortho-substituted, coplanar PCB; and 2,2',4,4',5,5'-hexachlorobiphenyl (PCB-153), a di-ortho-substituted, non-planar PCB. Groups of male Sprague-Dawley rats were given a single i.p. injection of one of the two PCBs (at doses of 30, 150, or 300 micromol/kg), both PCBs (at doses of 30 or 150 micromol/kg), or vehicle alone. Rats were sacrificed after 2, 6, or 24 h; or 2, 6, or 10 days. Cytochrome P-450 induction occurred as early as 2 h with PCB-77 and 24 h with PCB-153. Significant increases in thiobarbituric acid reactive substances (TBARS) content in liver tissue occurred 2, 6 and 10 days after treatment with PCB-77 and PCB-153; it was unclear whether these PCBs were synergistic in their induction of TBARS formation. Liver microsomal fractions incubated with NADPH only showed increased TBARS formation at the highest doses of PCB-77 and PCB-153 after 6 days. The results indicate that both PCBs induced cytochrome P-450 enzymes and enhanced lipid peroxidation in liver and subcellular fractions but with different potencies and onsets of action. The results also indicate a larger time difference between cytochrome P-450 induction and lipid peroxidation for PCB-77. Thus, both PCB-77 and PCB-153 are toxic to cells, but may act via different mechanisms to induce their effects.     

Effect of exposure to 3,3',4,4',5-pentachlorobiphenyl (PCB 126) throughout gestation and lactation on development and spatial delayed alternation performance in rats

There is evidence that polychlorinated biphenyl (PCB) congeners have differential effects on endpoints of neurotoxicity depending on their chemical structure: specifically, that ortho-substituted congeners are neurotoxic whereas coplanar (dioxin-like) congeners are relatively inactive in producing neurotoxic effects. The effects of the coplanar congener 3,3',4,4',5-pentachlorobiphenyl (PCB 126) on developmental endpoints, hematology, serum biochemistry, and performance on a spatial delayed alternation task were assessed in Long-Evans rats. Dams were dosed with 0, 0.25, or 1.0 microg/kg/day Monday to Friday beginning 5 weeks before and continuing through gestation and lactation. The first 2-week breeding period produced 10, 8, and 13 litters in the three dose groups, respectively. Breeding females from the control and low-dose group that did not conceive were rebred after 76 days of dosing, producing 7 and 6 litters, respectively. Reduction in weight gain from birth to weaning at 21 days of age (DOA) was observed in both dose groups of Cohort 1 but not in Cohort 2. Males in Cohort 1 exhibited a slight decrease in anogenital distance normalized for weight. Changes in hematological and some serum biochemical parameters were observed in the pups at DOA 21 and/or 60. PCB 126 was detected in fat sampled at both DOA 21 and 60. PCB 126 was not detected in brain samples at 60 DOA in any group; analysis of Cohort 2 at DOA 21 revealed levels in the treated group about 1/100 of those in fat. On the spatial delayed alternation task, there was no convincing evidence for impairment as a result of PCB exposure, as assessed by overall accuracy of performance and measures of perseverative and other types of inappropriate responding. These data provide further evidence for the lack of neurotoxicity of dioxin-like PCB congeners. However, assessment of performance on additional behavioral indices is required before definitive conclusions may be drawn.     

Behavioral assessments of learning and attention in rats exposed perinatally to 3,3',4,4',5-pentachlorobiphenyl (PCB 126)

Evidence from humans suggests that cognitive dysfunction may result from perinatal exposure to polychlorinated biphenyls (PCBs), and the results of some animal research with PCBs have been interpreted in terms of possible impairment of attention. Long-Evans rats were fed 3,3',4,4',5-pentachlorobiphenyl (PCB 126), a coplanar congener, at doses of 0.25 or 1 microgram/kg/day [corrected] throughout gestation and nursing. Male offspring of these rats were trained as adults to perform 2 tests of attention for food reward. First, a cued target-detection task, modeled after Posner's covert orienting method for humans, was used to assess visuospatial attention. In this task, a visual target stimulus was presented in 1 visual hemifield on each trial, preceded either by a valid cue, an invalid cue, or no cue. A valid cue appeared in the same hemifield as the target, and an invalid cue appeared in the opposite hemifield. As expected, valid cues increased accuracy and speed of target detection and invalid cues decreased accuracy and speed; moreover, these effects were systematically related to changes in cue intensity and target duration. However, perinatal exposure to PCB 126 did not affect acquisition or performance of this task. The second task assessed sustained attention by means of a signal detection method in which a brief, spatially-constant but temporally unpredictable, visual signal indicated which of 2 responses would yield food. Varying the intensity of the signal greatly affected the probability of correctly reporting the signal. Perinatal exposure to PCB 126 did not affect acquisition of the response rule or performance of the task. Finally, all rats were challenged with chlordiazepoxide (CDP) at doses of 0, 3, 5, 8, or 12 mg/kg SC, 20 min before testing in the sustained attention task. In control rats, low doses (3, 5, and 8 mg/kg) of CDP reduced accuracy at low signal intensities only, suggesting an increase in visual threshold. The high dose of CDP reduced accuracy at all signal intensities and increased the false-alarm rate as well, suggesting an impairment of attention. The rats exposed perinatally to PCB 126 at 0.25 micrograms/kg [corrected] were unaffected by CDP, and those exposed to PCB 126 at 1 microgram/kg [corrected] showed a smaller decrement in performance after CDP than did the controls. Taken together, these data provide little support for the possibility that perinatal exposure to PCB 126 causes deficits in attention, but suggest that PCB 126 may alter GABA-mediated pathways in the CNS during development.