ABSTRACT
Aroclor 1254 is a widely studied commercial polychlorinated biphenyl (PCB) mixture which, by definition, contains 54% chlorine by weight. Recent reports indicate substantial differences in the congener composition among Aroclor lots and hence their biologic effects. We designed the current study to compare the effects of two lots of Aroclor 1254 (lots 6024 and 124-191). We analyzed these two lots for PCB congeners, polychlorinated dibenzofurans (PCDFs), polychlorinated naphthalenes (PCNs), and polychlorinated dibenzodioxins (PCDDs). We used previously established techniques for analyzing intracellular Ca(2+) buffering and protein kinase C (PKC) translocation to test their biologic activity in neuronal preparations. PCB congener-specific analysis indicated that ortho and non-ortho congeners in these two lots varied in their percent contribution. Among all congeners, the percentages of non-ortho congeners (PCBs 77, 81, 126, and 169) were higher in lot 6024 (2.9% of total) than in lot 124-191 (0.02% of total). We detected no dioxins in these two lots (< 2 ppb). Although there are some differences in the congener composition, total PCNs were similar in both lots: 171 ppm in lot 6024 and 155 ppm in lot 124-191. However, total PCDFs were higher in lot 6024 (38.7 ppm) than in lot 124-191 (11.3 ppm). When we tested these two Aroclors on Ca(2+) buffering and PKC translocation in brain preparations, the effects were significantly different. Although lot 124-191 was more potent on PKC translocation than lot 6024, lot 6024 was slightly more active on Ca(2+) buffering than lot 124-191. These effects could not be attributed to the differences in the percentage of non-ortho congeners or PCDFs because they were inactive on these two parameters. The effects could not be attributed to PCNs because the levels were almost similar. The effects seen with two lots of Aroclor 1254 in neuronal cells were also not predicted based on the TCDD toxic equivalents (TEQs), although TEQs predicted the effects on ethoxyresorufin-O-deethylase (EROD) or methoxyresorufin-O-deethylase (MROD) activities. It is possible that the differential effects seen in neuronal cells could be caused by differences in the composition of ortho-congeners in these two mixtures, because PCBs with ortho-lateral substitutions can exhibit different activities on the selected neurochemical end points. Because of these differential effects with different lot numbers, the composition of Aroclor mixtures used in investigations should be disclosed.
Subject(s)
Antithyroid Agents/adverse effects , Antithyroid Agents/chemistry , Cytochrome P-450 CYP1A1/metabolism , Polychlorinated Dibenzodioxins/analogs & derivatives , Protein Kinases/metabolism , Animals , Benzofurans/analysis , Brain/drug effects , Brain/physiology , Calcium/pharmacokinetics , Cytochrome P-450 CYP1A1/drug effects , Female , Isomerism , Male , Neurons/drug effects , Neurons/enzymology , Polychlorinated Biphenyls/analysis , Polychlorinated Dibenzodioxins/analysis , Protein Kinases/drug effects , Protein Transport , Rats , Rats, Long-Evans , Reproducibility of Results , Risk Assessment , Soil Pollutants/analysisABSTRACT
Aroclor 1254 is a commercial mixture of polychlorinated biphenyls (PCBs), which is defined as being 54% chlorine by weight. However, the congener composition varies from lot to lot. Two lots which have been used in toxicity studies, 124-191 and 6024 (AccuStandard), were analyzed for their congener composition. Lot 6024 has approximately 10 times the dioxin toxic equivalents (TEQ) of lot 124-191. The purpose of this study was to determine if the difference in the TEQ of the two lots explains the different in vivo responses seen on a weight basis. Male Long-Evans rats (70 days old) were treated orally with a single dose of 0-1,000 mg/kg of each lot. Hepatic ethoxy-, methoxy-, and pentoxyresorufin O-deethylase (EROD, MROD, and PROD, respectively) activities as well as serum thyroxine (T(4)) concentrations and measures of oxidative stress were determined 4 days after treatment. Results, on a weight basis, indicate that lot 6024 led to a greater induction of EROD, MROD, and PROD but not total T(4) reduction. The differences in TEQ between the lots explained the differential induction of EROD and MROD but did not account for the induction of PROD nor decreases in T(4). PROD induction is not due to dioxin-like congeners, whereas the decrease in serum T(4) levels may involve multiple mechanisms. Effects on the antioxidants ascorbic acid and uric acid were seen only at the highest mass dose for both lots and were not explained by the difference in TEQ. These results illustrate that the differences in the TEQ explain the differences in the strict dioxin-like effects (EROD, MROD induction), but the non-dioxin-like congeners cause other effects that are not associated with the aryl hydrocarbon receptor (e.g., PROD). In addition, supra-additive effects also occur in the mixture (T(4), oxidative stress). Thus, current results demonstrate that overall toxicity cannot be predicted on the basis of the TEQ values. It is also critical that the lot number is reported in studies conducted with Aroclor 1254 because the congener composition and therefore the effects observed can be very different.
Subject(s)
Antithyroid Agents/adverse effects , Cytochrome P-450 CYP1A1/metabolism , Cytochrome P-450 CYP2B1/metabolism , Cytochrome P-450 Enzyme System/metabolism , Oxidative Stress , Oxidoreductases/metabolism , Thyroxine/blood , Administration, Oral , Animals , Cytochrome P-450 CYP1A1/drug effects , Cytochrome P-450 CYP2B1/drug effects , Cytochrome P-450 Enzyme System/drug effects , Enzyme Induction , Isomerism , Liver/drug effects , Liver/enzymology , Male , Oxidoreductases/drug effects , Rats , Rats, Long-Evans , Reproducibility of Results , Thyroxine/drug effectsABSTRACT
Pregnant Long-Evans rats were exposed to 0, 1 or 6 mg/kg/day of Aroclor 1254 (A1254; Lot no. 124-191), a commercial mixture of polychlorinated biphenyls (PCBs), from gestation day (GD) 6 through postnatal day (PND) 21. At 128-140 days of age, male and female offspring were tested for visual-, somatosensory- and peripheral nerve-evoked potentials. The evoked responses increased in amplitude with larger stimulus intensities, and gender differences were detected for some endpoints. In contrast, developmental exposure to A1254 failed to significantly affect the electrophysiological measures. A subset of the animals were tested for low-frequency hearing dysfunction using reflex modification audiometry (RMA). An elevated threshold for a 1-kHz tone was observed, replicating previous findings of A1254-induced auditory deficits [Hear. Res. 144 (2000) 196; Toxicol. Sci. 45(1) (1998) 94; Toxicol. Appl. Pharmacol. 135(1) (1995) 77.]. These findings indicate no statistically significant changes in visual-, somatosensory- or peripheral nerve-evoked potentials following developmental exposure to doses of A1254 that produce behavioral hearing deficits. However, subtle changes in the function of the visual or somatosensory systems cannot be disproved.
Subject(s)
Auditory Threshold/drug effects , Evoked Potentials, Somatosensory/drug effects , Evoked Potentials, Visual/drug effects , Administration, Oral , Animals , Female , Male , Photic Stimulation , Pregnancy , Prenatal Exposure Delayed Effects , Rats , Rats, Long-EvansABSTRACT
Previous research has demonstrated the sensitivity of the developing rat to the hypothyroxinemic and ototoxic effects of perinatal exposure to Aroclor 1254 (A1254). We tested the hypothesis that postnatal exposure via lactation is the major cause of the ototoxicity by cross fostering animals at birth. Primiparous rats (22-24/dose) received 0 or 6 mg/kg A1254 (po in corn oil) from gestation day (GD) 6 to postnatal day (PND) 21. On the day of birth, half of the treated litters and half of the control litters were cross-fostered, resulting in the following groups: Ctrl/Ctrl (controls); A1254/A1254 (perinatal exposure); A1254/Ctrl (prenatal exposure only); and Ctrl/A1254 (postnatal exposure only). We assessed offspring at a number of ages for: serum thyroid hormone concentrations, liver and brain concentrations of PCBs, body weight, mortality, age of eye opening, auditory startle amplitudes, and auditory thresholds for 1 kHz and 40 kHz tones. Circulating thyroxine (T(4)) concentrations were sharply reduced at GD 21 in the A1254-exposed group, and on PND 3, 7, 14, and 21 in the A1254/A1254 and the Ctrl/A1254 groups. Smaller decreases in T(4) were observed in the A1254/Ctrl group on PND 3, 7, and 14. PCB concentrations in the liver on PND 21 were sharply elevated in the A1254/A1254 and Ctrl/A1254 groups. Much smaller increases were seen in the A1254/Ctrl group. Age of eye-opening and startle amplitudes were unaffected by treatment. A1254 exposure caused permanent hearing deficits (20 dB increase) at the low frequency (1 kHz) in the A1254/A1254 and Ctrl/A1254 groups. The present findings demonstrated that the critical period for the ototoxicity of developmental A1254 exposure is within the first few postnatal weeks in the rat. This effect is consistent with the greater degree of postnatal hypothyroxinemia resulting from the greater magnitude of exposure that occurs postnatally via lactation.
Subject(s)
Hearing Disorders/chemically induced , Lactation/physiology , Polychlorinated Biphenyls/toxicity , Thyroid Hormones/blood , Aging/metabolism , Animals , Auditory Threshold/drug effects , Behavior, Animal/drug effects , Body Weight/drug effects , Eye/growth & development , Female , Liver/metabolism , Male , Milk/chemistry , Polychlorinated Biphenyls/pharmacokinetics , Pregnancy , Rats , Rats, Long-Evans , Reflex, Startle/drug effects , Thyroxine/blood , Triiodothyronine/bloodABSTRACT
Earlier reports from our laboratory have indicated that polychlorinated biphenyls (PCBs) affect signal transduction mechanisms in brain, including Ca2+ homeostasis, phosphoinositol hydrolysis, and protein kinase C (PKC) translocation in mature neurons and adult brain homogenate preparations. Present studies were designed to investigate whether there were any brain region-, gender-, or age-dependent effects of PCBs on 45Ca2+-uptake by two subcellular organelles, microsomes and mitochondria. We have studied in vitro effects of a widely studied commercial PCB mixture, Aroclor 1254R, on 45Ca2+-uptake by microsomes and mitochondria in cerebellum, frontal cortex and hippocampus of postnatal day (PND) 7, 21, and 90-120 (adult) male and female Long-Evans (LE)-rats. In general, microsomal and mitochondrial 45Ca2+-uptake in selected brain regions increased with age; PND 7Subject(s)
Brain/drug effects
, Calcium/metabolism
, /toxicity
, Age Factors
, Animals
, Brain/metabolism
, Brain/ultrastructure
, Calcium-Transporting ATPases/metabolism
, Female
, Male
, Microsomes/drug effects
, Microsomes/metabolism
, Mitochondria/drug effects
, Mitochondria/metabolism
, Rats
, Sex Factors
ABSTRACT
Polychlorinated biphenyls (PCBs) are ubiquitous environmental contaminants, some of which may be neurotoxic. In vitro studies from this laboratory indicated that noncoplanar PCBs perturbed intracellular signal transduction mechanisms including Ca2+ homeostasis, receptor-mediated inositol phosphate production, and translocation of protein kinase C (PKC). In the present study, we examined the effects of PCBs in vivo by dosing adult male Long-Evans rats orally with Aroclor 1254 (0, 10, or 30 mg/kg/day; 5 days/week for 4 weeks) in corn oil. At 24 h after the last dose, rats were tested for motor activity in a photocell device for 30 min. Immediately, the rats were euthanized, blood was collected for thyroid hormone analysis, and brains were removed, dissected into regions (cerebellum, frontal cortex, and striatum), and subcellular fractions were obtained for neurochemical analysis. Following Aroclor 1254 treatment, body weight gain in the high-dose group was significantly lower than the control and low-dose groups. Horizontal motor activity was significantly lower in rats dosed with 30 mg/kg Aroclor 1254. Ca2+ buffering by microsomes was significantly lower in all three brain regions from the 30 mg/kg group. In the same dose group, mitochondrial Ca2+ buffering was affected in cerebellum but not in cortex or striatum. Similarly, total cerebellar PKC activity was decreased significantly while membrane-bound PKC activity was significantly elevated at 10 and 30 mg/kg. PKC activity was not altered either in cortex or the striatum. Neurotransmitter levels in striatum or cortex were slightly altered in PCB-exposed rats compared to controls. Furthermore, repeated oral administration of Aroclor 1254 to rats did not significantly alter forebrain tyrosine hydroxylase immunoreactivity or enzymatic activity. Circulating T4 (total and free) concentrations were severely depressed at both doses in Aroclor 1254-exposed rats compared to control rats, suggesting a severe hypothyroid state. These results indicate that (1) in vivo exposure to a PCB mixture can produce changes in second messenger systems that are similar to those observed after in vitro exposure of neuronal cell cultures; (2) second messenger systems seem to be more sensitive than alterations in neurotransmitter levels or tyrosine hydroxylase involved in dopamine synthesis during repeated exposure to PCBs; and (3) the observed motor activity changes were independent of changes in striatal dopamine levels.
Subject(s)
Antithyroid Agents/toxicity , Brain/metabolism , Calcium/metabolism , Protein Kinase C/metabolism , Tyrosine 3-Monooxygenase/metabolism , Animals , Body Weight/drug effects , Buffers , Cerebellum/metabolism , Cerebral Cortex/metabolism , Dopamine/metabolism , Immunohistochemistry , Male , Microsomes/metabolism , Mitochondria/metabolism , Motor Activity/drug effects , Neurotransmitter Agents/analysis , Polychlorinated Biphenyls/toxicity , Rats , Rats, Long-Evans , Thyroid Hormones/metabolism , Time Factors , Visual Cortex/metabolismABSTRACT
Our previous in vitro studies with both isolated organelles and primary neuronal cell cultures found that intracellular signal transduction can be perturbed by some noncoplanar PCBs at exposure levels of =10 microM. However, it is not clear whether such concentrations are achievable in brain in vivo. Also, the pattern of congener disposition and quantities of the PCB accumulation in tissues of animals exposed to commercial PCB mixtures is not well studied. In the present study, we have conducted PCB congener-specific analysis in different brain regions, liver, blood, and fat of adult male Long-Evans rats dosed orally with Aroclor 1254 (0 or 30 mg/kg/day; once per day, 5 days/week for 4 weeks) in corn oil. Twenty-four hours after the last dose, rats were euthanized, and the brains were removed and dissected to obtain cerebellum, frontal cortex, and striatum. Liver, blood, and fat samples were also collected at the same time. Congener-specific analysis of PCBs was performed by high-resolution gas chromatography with electron capture detection. While PCB concentrations in control rat brain regions were less than 0.02 ppm, total PCB congeners in treated animals accumulated to much higher levels. Total levels in the frontal cortex, cerebellum, and striatum were 15.1 +/- 0.3, 13.1 +/- 1.7, and 8.2 +/- 2.6 ppm, respectively. The levels of PCBs in the fat, liver, and blood were 0.041, 0.002, and 0.001 ppm in control rats and 552, 38.3, and 1.6 ppm in treated rats, respectively. In addition to the differential total uptake between tissues, there was differential accumulation of PCBs with respect to the number of chlorines. In all the tissues, the more lightly chlorinated (tetra- and penta-) congeners accumulated less than their respective proportions in the parent Aroclor 1254 mixture. On the other hand, heavily chlorinated (hexa- to nona-) congeners accumulated more than the proportion of these congeners found in Aroclor 1254 mixture. This shift toward accumulation of heavily chlorinated congeners seems to be more pronounced in the brain than liver and fat. Predominant congeners (5-32% of total PCBs) detected in different brain regions, blood, liver, and fat are: 2,3,3',4',5,6- (no. 163) + 2,2',3,4,4',5- (no. 138) (coeluted); 2,2',4,4',5,5'- (no. 153) + 2, 2',3,3',4,6'- (no. 132) (coeluted); 2,3,3',4,4',5- (no. 156) + 2,2', 3,3',4,4',6- (no. 171) (coeluted); 2,3',4,4',5- (no. 118); 2,2',4,4', 5-(no. 99); and 2,3,3',4,4'- (no. 105). These congeners together accounted for about two thirds of the total PCB load in brain. All these predominant congeners are ortho-substituted and therefore are noncoplanar in nature. The total PCB concentrations accumulated in brain were as high as 50 microM (based on average molecular weight of 326.4 for Aroclor 1254) and, at these concentrations, intracellular second messengers were significantly affected in neuronal cultures and brain homogenate preparations in vitro. These results indicate that concentrations that altered Ca2+ disposition and second messenger systems in vitro are achievable in brain in vivo following repeated exposure.