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Biomedical subjects

T R Ward

Publications and source records attributed to T R Ward.

At least 19 recordsLinked to original sources
PubMed2001

Novel annulene dications from methylated [2.2]metacyclophane monoenes and [e]-ring benzannelated dimethyldihydropyrene.

Tetramethyl- and hexamethyl-substituted [2.2]metacyclophane monoenes (10 and 11) are transformed into their corresponding trans-dimethyldihydroethanophenanthrenium dications (14(2+) and 15(2+)) in FSO(3)H x SbF(5) (4:1) and FSO(3)H x SbF(5) (1:1) with SO(2)ClF or SO(2) as the solvent; these 10 pi-dications are equivalent to the C-4/C-5 diprotonated dications of the 2,7-dimethyl derivative of trans-DMDHP, 3a. The trans-12c,12d-dimethyl-12c,12d-dihydrobenzo[e]pyrene (6) reacts with FSO(3)H/SO(2)ClF under surprisingly mild conditions to give initially a persistent diprotonated dication (6H(2)(2+)) and, subsequently, the oxidation dication (6(2+)); the 6(2+):6H(2)(2+) ratio reaches 4:1 after 1 week at low temperature. Protonation of the anti-metacyclophane (13) was also examined. Charge delocalization mode and tropicity in the resulting dications are gauged via detailed NMR studies at 500 MHz.

Bridged-Ring Compounds↗
PubMed2001

Neurotransmitter concentrations and binding at dopamine receptors in rats after maternal exposure to 3,4,3',4'-tetrachlorobiphenyl: the role of reduced thyroid hormone concentrations.

Polychlorinated biphenyls (PCBs) are environmental contaminants, which accumulate in the food chain and are transferred to the offspring during prenatal development through the placenta and postnatally via breast milk. It is reported that PCBs exert effects on thyroid hormone levels and brain neurotransmitter levels. Both actions may alter neuronal development. The aim of the present study was to investigate, if PCB-induced effects on concentrations of catecholamines and serotonin can be attributed to PCB-induced reductions in thyroid hormone concentrations. In addition, binding to dopamine D(1) and D(2) receptors was examined. Time-mated Wistar rats were treated prenatally with 1 mg 3,4,3',4'-tetrachlorobiphenyl (PCB 77)/kg bodyweight or the vehicle. A third group serving as the positive control received perinatally 5 mg propylthiouracil (PTU)/l drinking water. There were no overt toxic signs in dams or offspring. Thyroid hormone measurements demonstrated effects in dams and offspring up to postnatal day 40. In particular, total T(4) in serum and in the thyroid were decreased in PCB- and PTU-treated dams and offspring. Only PTU exposed rats exhibited significantly increased concentrations of TSH in the serum and pituitary. Measurement of neurotransmitters revealed changes in the PCB-exposed offspring at PND 40, while PTU-treatment was without effect. Dopamine and DOPAC were increased in the medial prefrontal cortex. In adulthood, there were no PCB-related effects on thyroid hormones and neurotransmitters. Binding studies of dopamine D1 and D2 receptors demonstrated that PCB and PTU had no influence on receptor concentration and affinity. Comparison of PCB 77 exposed offspring to PTU exposed offspring demonstrated differential effects on TSH and neurotransmitter levels, the latter result indicating that not all PCB-induced effects on the nervous system can be ascribed to decreases in thyroid hormone concentrations.

Journal Article↗
PubMed2001

Neurotoxicological outcomes of perinatal heptachlor exposure in the rat.

The developing nervous system has been identified as a potential target of pesticide exposure. Heptachlor is a cyclodiene pesticide that was widely used for many years, and for which inadvertent exposure to children and fetuses took place in the early 1980s; yet little is known regarding the developmental neurotoxicity of it and other cyclodienes. The aim of this study was to determine whether perinatal heptachlor exposure results in persistent alterations in nervous system function. Pregnant Sprague-Dawley dams were dosed from gestational day (GD) 12 to postnatal day (PND) 7, whereupon the rat pups were dosed directly until PND 21 (group A) or PND 42 (group B). Dose levels were 0, 0.03, 0.3, or 3 mg/kg/day, po. There were no dose-related effects on maternal weight, litter size, or pup growth. GABA(A) receptor binding (using [(35)S] tert-butylbicyclophosphorothionate; TBPS) and GABA-stimulated Cl- flux were evaluated in control and high-dose brain tissues taken on PND 7, 21, and 43. The B(max) values for [(35)S]-TBPS binding in brainstem, but not cortex, were decreased in female rats across all ages tested. There were no such changes in male rats, nor were K(D) values altered in either tissue or gender. GABA-stimulated Cl- flux was decreased in female cortex synaptoneurosomes only on PND 21. The ontogeny of the righting response (PND 2-5) was delayed in the high-dose females. All subsequent testing took place a week to months after dosing ceased. The functional observational battery (FOB) showed treatment-related, but not necessarily dose-related, changes in different aspects of the rat's reactivity and activity levels. Group-A rats also showed altered within-session habituation of motor activity. There were no heptachlor-related differences in motor activity following challenge with a range of chlordiazepoxide doses. Cognitive assessments were conducted in both groups of rats. There were no statistically significant differences among treatment groups in a one-trial passive avoidance test, although there was a trend toward less learning. In group B, rats (both sexes), heptachlor altered spatial learning in the Morris water maze during two weeks of daily training (2 trials/day). On probe trials, heptachlor-treated rats did not show significant preference for the correct quadrant (all dose groups in males, high dose in females). These rats did not show alterations on subsequent working-memory training (where the platform position was relearned each day). Thus, perinatal exposure to heptachlor produced neurochemical and persistent neurobehavioral changes, including alterations in spatial learning and memory.

Animals↗
PubMed2001

Developmental exposure of rats to a reconstituted PCB mixture or aroclor 1254: effects on long-term potentiation and [3H]MK-801 binding in occipital cortex and hippocampus.

The central nervous system is one of the target organs for polychlorinated biphenyls (PCBs). We measured the effects of maternal exposure of Long-Evans rats to a mixture of PCB congeners reconstituted according to the pattern found in human breast milk (reconstituted mixture, RM) on long-term potentiation (LTP) in two brain regions. Exposure of the dams via food started 50 days prior to mating and was terminated at birth. In the first experiment, adult male and female offspring were exposed maternally to 40 mg/kg of the RM or the commercial mixture Aroclor 1254 (A1254). LTP and paired-pulse inhibition were measured in slices of the visual cortex. In addition, the binding of [3H]MK-801 to the N-methyl-D-aspartate (NMDA) receptor-ion channel as well as the [3H]muscimol binding to the GABA-A receptor in membrane preparations from the occipital cortex and hippocampus were determined. LTP as well as [3H]MK-801 binding were significantly reduced in the cortex following PCB exposure, while [3H]MK-801 binding in the hippocampus was not affected. In a succeeding experiment, LTP was determined in cortical and hippocampal slices from rats at postnatal days 10 to 20, following exposure to 0, 5, or 40 mg/kg of the RM. Cortical LTP was significantly affected by the RM while no effects were seen in hippocampal LTP. Taking the two experiments together, PCB exposure significantly reduced LTP, as well as [3H]MK-801 binding, in the cortex and had no effect in the hippocampus. The LTP deficits can only partly be related to the reduction of binding sites to the NMDA receptor; other PCB-induced neurochemical changes have to be assumed.

Administration, Oral↗
PubMed2001

Differential effects of two lots of aroclor 1254: congener-specific analysis and neurochemical end points.

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.

Animals↗
PubMed2000

Widespread aneuploidy revealed by DNA microarray expression profiling.

Expression profiling using DNA microarrays holds great promise for a variety of research applications, including the systematic characterization of genes discovered by sequencing projects. To demonstrate the general usefulness of this approach, we recently obtained expression profiles for nearly 300 Saccharomyces cerevisiae deletion mutants. Approximately 8% of the mutants profiled exhibited chromosome-wide expression biases, leading to spurious correlations among profiles. Competitive hybridization of genomic DNA from the mutant strains and their isogenic parental wild-type strains showed they were aneuploid for whole chromosomes or chromosomal segments. Expression profile data published by several other laboratories also suggest the use of aneuploid strains. In five separate cases, the extra chromosome harboured a close homologue of the deleted gene; in two cases, a clear growth advantage for cells acquiring the extra chromosome was demonstrated. Our results have implications for interpreting whole-genome expression data, particularly from cells known to suffer genomic instability, such as malignant or immortalized cells.

Aneuploidy↗
PubMed2000

Iha: a novel Escherichia coli O157:H7 adherence-conferring molecule encoded on a recently acquired chromosomal island of conserved structure.

The mechanisms used by Shiga toxin (Stx)-producing Escherichia coli to adhere to epithelial cells are incompletely understood. Two cosmids from an E. coli O157:H7 DNA library contain an adherence-conferring chromosomal gene encoding a protein similar to iron-regulated gene A (IrgA) of Vibrio cholerae (M. B. Goldberg, S. A. Boyko, J. R. Butterton, J. A. Stoebner, S. M. Payne, and S. B. Calderwood, Mol. Microbiol. 6:2407-2418, 1992). We have termed the product of this gene the IrgA homologue adhesin (Iha), which is encoded by iha. Iha is 67 kDa in E. coli O157:H7 and 78 kDa in laboratory E. coli and is structurally unlike other known adhesins. DNA adjacent to iha contains tellurite resistance loci and is conserved in structure in distantly related pathogenic E. coli, but it is absent from nontoxigenic E. coli O55:H7, sorbitol-fermenting Stx-producing E. coli O157:H-, and laboratory E. coli. We have termed this region the tellurite resistance- and adherence-conferring island. We conclude that Iha is a novel bacterial adherence-conferring protein and is contained within an E. coli chromosomal island of conserved structure. Pathogenic E. coli O157:H7 has only recently acquired this island.

Adhesins, Bacterial↗
PubMed2000

Acquisition of the rfb-gnd cluster in evolution of Escherichia coli O55 and O157.

The rfb region specifies the structure of lipopolysaccharide side chains that comprise the diverse gram-negative bacterial somatic (O) antigens. The rfb locus is adjacent to gnd, which is a polymorphic gene encoding 6-phosphogluconate dehydrogenase. To determine if rfb and gnd cotransfer, we sequenced gnd in five O55 and 13 O157 strains of Escherichia coli. E. coli O157:H7 has a gnd allele (allele A) that is only 82% identical to the gnd allele (allele D) of closely related E. coli O55:H7. In contrast, gnd alleles of E. coli O55 in distant lineages are >99.9% identical to gnd allele D. Though gnd alleles B and C in E. coli O157 that are distantly related to E. coli O157:H7 are more similar to allele A than to allele D, there are nucleotide differences at 4 to 6% of their sites. Alleles B and C can be found in E. coli O157 in different lineages, but we have found allele A only in E. coli O157 belonging to the DEC5 lineage. DNA 3' to the O55 gnd allele in diverse E. coli lineages has sequences homologous to tnpA of the Salmonella enterica serovar Typhimurium IS200 element, E. coli Rhs elements (including an H-rpt gene), and portions of the O111 and O157 rfb regions. We conclude that rfb and gnd cotransferred into E. coli O55 and O157 in widely separated lineages and that recombination was responsible for recent antigenic shifts in the emergence of pathogenic E. coli O55 and O157.

Alleles↗
PubMed2000

Ribosomal DNA replication fork barrier and HOT1 recombination hot spot: shared sequences but independent activities.

In the ribosomal DNA of Saccharomyces cerevisiae, sequences in the nontranscribed spacer 3' of the 35S ribosomal RNA gene are important to the polar arrest of replication forks at a site called the replication fork barrier (RFB) and also to the cis-acting, mitotic hyperrecombination site called HOT1. We have found that the RFB and HOT1 activity share some but not all of their essential sequences. Many of the mutations that reduce HOT1 recombination also decrease or eliminate fork arrest at one of two closely spaced RFB sites, RFB1 and RFB2. A simple model for the juxtaposition of RFB and HOT1 sequences is that the breakage of strands in replication forks arrested at RFB stimulates recombination. Contrary to this model, we show here that HOT1-stimulated recombination does not require the arrest of forks at the RFB. Therefore, while HOT1 activity is independent of replication fork arrest, HOT1 and RFB require some common sequences, suggesting the existence of a common trans-acting factor(s).

DNA Replication↗
PubMed1999

Effects of the chlorotriazine herbicide, cyanazine, on GABA(A) receptors in cortical tissue from rat brain.

Chlorotriazine herbicides disrupt luteinizing hormone (LH) release in female rats following in vivo exposure. Although the mechanism of action is unknown, significant evidence suggests that inhibition of LH release by chlorotriazines may be mediated by effects in the central nervous system. GABA(A) receptors are important for neuronal regulation of gonadotropin releasing hormone and LH release. The ability of chlorotriazine herbicides to interact with GABA(A) receptors was examined by measuring their effects on [3H]muscimol, [3H]Ro15-4513 and [35S]tert-butylbicyclophosphorothionate (TBPS) binding to rat cortical membranes. Cyanazine (1-400 microM) inhibited [3H]Ro15-4513 binding with an IC50 of approximately 105 microM (n=4). Atrazine (1-400 microM) also inhibited [3H]Ro15-4513 binding, but was less potent than cyanazine (IC50 = 305 microM). However, the chlorotriazine metabolites diaminochlorotriazine, 2-amino-4-chloro-6-ethylamino-s-triazine and 2-amino-4-chloro-6-isopropylamino-s-triazine were without significant effect on [3H]Ro15-4513 binding. Cyanazine and the other chlorotriazines were without effect on [3H]muscimol or [35S]TBPS binding. To examine whether cyanazine altered GABA(A) receptor function, GABA-stimulated 36Cl- flux into synaptoneurosomes was examined. Cyanazine (50-100 microM) alone did not significantly decrease GABA-stimulated 36Cl- flux. Diazepam (10 microM) and pentobarbital (100 microM) potentiated GABA-stimulated 36Cl- flux to 126 and 166% of control, respectively. At concentrations of 50 and 100 microM, cyanazine decreased potentiation by diazepam to 112 and 97% of control, respectively, and decreased potentiation by pentobarbital to 158 and 137% of control (n = 6). Interestingly, at lower concentrations (5 microM), cyanazine shifted the EC50 for GABA-stimulated 36Cl- flux into synaptoneurosomes from 28.9 to 19.4 microM, respectively (n = 5). These results suggest that cyanazine modulates benzodiazepine, but not the muscimol (GABA receptor site) or TBPS (Cl- channel), binding sites on GABA(A) receptors. Furthermore, at low concentrations, cyanazine may slightly enhance function of GABA(A) receptors, but at higher concentrations, cyanazine antagonizes GABA(A) receptor function and in particular antagonizes the positive modulatory effects of diazepam and pentobarbital.

Animals↗
PubMed1999

Functional characterization of the S. cerevisiae genome by gene deletion and parallel analysis.

The functions of many open reading frames (ORFs) identified in genome-sequencing projects are unknown. New, whole-genome approaches are required to systematically determine their function. A total of 6925 Saccharomyces cerevisiae strains were constructed, by a high-throughput strategy, each with a precise deletion of one of 2026 ORFs (more than one-third of the ORFs in the genome). Of the deleted ORFs, 17 percent were essential for viability in rich medium. The phenotypes of more than 500 deletion strains were assayed in parallel. Of the deletion strains, 40 percent showed quantitative growth defects in either rich or minimal medium.

Culture Media↗
PubMed1998

Congener-specific distribution of polychlorinated biphenyls in brain regions, blood, liver, and fat of adult rats following repeated exposure to Aroclor 1254.

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.

Adipose Tissue↗
PubMed1998

Interactive effects of environmentally relevant polychlorinated biphenyls and dioxins on [3H]phorbol ester binding in rat cerebellar granule cells.

Polychlorinated biphenyls (PCBs) are persistent contaminants that exist as complex mixtures in the environment. One problem faced by risk assessors is that the possible interactive effects of specific PCB congeners and related chemicals found in environmental and biological samples have not been systematically investigated. Some PCBs perturb Ca2+ homeostasis and cause protein kinase C (PKC) translocation in neuronal cell cultures and in brain homogenate preparations at concentrations where no cytotoxicity is observed, and these systems are necessary for the growth and normal functioning of neurons. The changes in second messenger systems appear to be associated with the extent of noncoplanarity of the PCB molecule. We studied the interactive effects of selected PCB congeners, a PCB metabolite, and a dioxin on PKC translocation, as determined by [3H]phorbol ester binding in cerebellar granule cells. The binary combinations included coplanar and noncoplanar PCB congeners or PCB congeners with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)/PCB metabolite. In addition, we tested the interactive effects of several PCB congeners (three or more) found in environmental samples such as human milk and blood, contaminated fish, and brain samples from PCB-treated animals. The results indicated that 1) the coplanar congener [3,3',4, 4'-tetrachlorobiphenyl (TeCB)] did not alter the in vitro activity of the noncoplanar (2,2',5,5'-TeCB) or coplanar [4, 4'-dichlorobiphenyl (DCB)] congeners; 2) binary mixtures of active PCB congeners (2,2',4,4'-TeCB and 2,2'-DCB; 2,2'-DCB and 3,5-DCB; 2,2',3,5',6-PeCB and 2,2',4,4',5-PeCB) interact in a dose-additive manner; 3) TCDD did not alter the activity of either coplanar (3,3', 4,4'-TeCB) or noncoplanar (2,2',5,5'-TeCB) congeners; 4) the interaction between the parent PCB congener and hydroxy metabolite of PCB is additive; 5) PCB congener mixtures at the ratios found in environmental samples are biologically active; and 6) there was no indication of synergism in any of the combinations studied. These results suggest that the biological effects of binary mixtures of PCB congeners fit a dose-additive model, indicating that there is a specific site of action for these PCB congeners which is independent of the aryl hydrocarbon receptor. Environmental mixtures contain mostly noncoplanar PCB congeners, and because they appear to be biologically active, the potential human health risk by this group of chemicals should be considered in the risk assessment of PCBs.

Animals↗
PubMed1997

Empirical modeling of an in vitro activity of polychlorinated biphenyl congeners and mixtures.

The goal of this research is to predict an in vitro activity of polychlorinated biphenyl (PCB) congeners and their mixtures and to describe the relationship between this activity and chemical structure. The test system used multiple PCB concentrations on each cell culture plate in a repeated measures design, which improved precision for comparing between concentration levels. A weighted regression that accounted for this experimental design feature was used in fitting a nonlinear dose-response exponential model to the PCB concentration-activity data from an in vitro test system in which 3H-phorbol ester binding was measured in cerebellar granule cells exposed to different PCB congeners to test for their effects on protein kinase C translocation. The model allowed for the minimum level to be less than control, a common slope, and the estimation of the log of the concentration that produces an activity 50% above the control activity (E50) for 36 congeners and 3 commercial mixtures. Next, a weighted logistic regression using a second order response model in the variables Clortho, Clpara, and Clmeta was used to relate the estimated log E50s to indicators of chemical structure. This model was preferred over models that might seem more mechanistically based because in internal validation, it attained a smaller PRESS statistic (the sum of squares between all observed and predicted observations) than other models. Evidently, this second order model makes more efficient use of parameters than other models considered. Plots of the predictions of the logistic second order response model versus log Kow confirm the usual pattern that congeners with intermediate levels of log Kow are the more active. The data of three commercial mixtures were included in this regression by assuming a common combination index (ratio of observed E50 to predicted E50, assuming dose addition). The logistic model suggests that congeners with one, two, or three chlorine substitutions at the ortho position are more active than other congeners. Also, congeners with log Kow between 5.2 and 6.6 are generally more active. The estimated combination index indicated that the joint action of PCB congeners in the three commercial mixtures was less than dose additive. The error sum of squares was significantly large, which may indicate a lack of fit of the logistic model. Empirical Bayes estimates (EBE) are weighted averages of model predictions and observations of E50s and can be better estimates than the fitted model when there is a lack of fit. The PRESS statistic for the EBE indicated larger prediction error than for the logistic model, but the EBE provided better estimates of commercial mixture E50s based on dose addition. This may indicate that the logistic model is not incorporating all the information in the single congener data needed to predict mixtures.

Animals↗
PubMed1996

Increased [3H]phorbol ester binding in rat cerebellar granule cells and inhibition of 45Ca2+ sequestration in rat cerebellum by polychlorinated diphenyl ether congeners and analogs: structure-activity relationships.

Our previous reports indicate that ortho-substituted non-coplanar polychlorinated biphenyl (PCB) congeners perturbed neuronal Ca2+-homeostasis in vitro, altered agonist-stimulated inositol phosphate accumulation, and caused protein kinase C (PKC) translocation. The structure-activity relationship (SAR) with 24 PCB congeners was consistent with a chlorination pattern that favored non-coplanarity while those with chlorination that favored coplanarity were less active. To test the hypothesis that coplanarity (or lack thereof) is a significant factor in the activity of PCBs, studies with related classes of chemicals such as the polychlorinated diphenyl ethers (PCDEs), in which coplanarity is more difficult to achieve regardless of degree and pattern of chlorination, were initiated. The selected PCDEs and their analogs are predicted to be active, since they are non-coplanar in nature. The effects of these chemicals were studied using the same measures for which PCBs had differential effects based on structural configuration. These measures include PKC translocation as determined by [3H]-phorbol ester ([3H]PDBu) binding in cerebellar granule cells and 45Ca2+ sequestration as determined by 45Ca2+ uptake by microsomes and mitochondria isolated from adult rat cerebellum. All the PCDE congeners studied, increased [3H]PDBu binding in a concentration-dependent manner. The order of potency was 2,4,4'-trichlorodiphenyl ether > 4,4'-dichlorodiphenyl ether > diphenyl ether, 3,3',4,4'-tetrachlorodiphenyl ether and, 2,2',4,4',5- and 2,3',4,4',5-pentachlorodiphenyl ethers. The structurally related diphenyl ether nitrofen and diphenyl ethanes o,p'-1,1,1-trichloro-2,2-bis[p-chlorophenyl]ethane (DDT) and p,p'-DDT increased [3H]PDBu binding to a similar extent (28-35% stimulation at 100 microM). All PCDE congeners and their analogs inhibited 45Ca2+ sequestration by microsomes and mitochondria. Of all the chemicals, unchlorinated diphenyl ether was the least active. These results are in agreement with previous SAR findings in which non-coplanar PCBs are active and support our hypothesis that the extent of coplanarity determined by a pattern of chlorination on certain aromatic hydrocarbons can weaken their potency in vitro, although the extent of chlorination is also important.

Animals↗
PubMed1996

Inhibition of microsomal and mitochondrial Ca2+-sequestration in rat cerebellum by polychlorinated biphenyl mixtures and congeners. Structure-activity relationships.

Recent studies from our laboratory indicate that polychlorinated biphenyl (PCB) congeners in vitro perturbed signal transduction mechanisms including cellular Ca2+-homeostasis and protein kinase C translocation. We have now investigated the structure-activity relationship (SAR) of three PCB mixtures, 24 PCB congeners and one dibenzofuran for their effects on microsomal and mitochondrial Ca2+-sequestration in rat cerebellum. Ca2+-sequestration by these intracellular organelles was determined using radioactive 45CaCl2. All three mixtures studied, Aroclor 1016, Aroclor 1254 and Aroclor 1260, were equally potent in inhibiting microsomal and mitochondrial Ca2+-sequestration with IC50 values of 6-8 microM. 1,2,3,7,8-Pentachlorodibenzofuran had no effect on Ca2+-sequestration by these organelles. The SAR among the congeners revealed: (1) congeners with ortho-/meta- or ortho-, para-chlorine substitutions were the most potent in inhibiting microsomal and mitochondrial Ca2+-sequestration (IC50 = 2.4-22.3 microM); (2) congeners with only para- but without ortho-substitutions were not effective in inhibiting Ca2+-sequestration by microsomes and mitochondria; (3) increased chlorination was not related to the effectiveness of these congeners. The present SAR studies indicate that the effects of most PCB congeners in vitro may be related to an interaction at specific sites having preference for low lateral substitution or lateral content (meta- or para) in the presence of ortho-substitution.

Animals↗
PubMed1996

Organophosphorus compounds preferentially affect second messenger systems coupled to M2/M4 receptors in rat frontal cortex.

Recent reports indicate that organophosphate insecticides, in addition to inhibiting acetylcholinesterase activity, can bind directly at a subset of muscarinic receptors, which also bind cis-methyldioxolane with high affinity. Muscarinic receptors are known to act through at least two second messenger systems, either the stimulation of phosphoinositide turnover (mediated through the M1 and M3 receptor subtypes) or the inhibition of cAMP formation (mediated through the M2 and M4 receptor subtypes). We have investigated the action of the active forms of parathion, malathion, and chlorpyrifos (paraoxon, malaoxon, and chlorpyrifos oxon, respectively) on these second messenger systems in cortical slices from adult male Long-Evans rats. Paraoxon, malaoxon, and chlorpyrifos oxon (10(-8) to 10(-4) M) inhibited forskolin-stimulated cAMP formation in a concentration-dependent manner. The effect on cAMP formation was blocked by the muscarinic antagonist atropine (10 microM). These results suggest that paraoxon, malaoxon, and chlorpyrifos oxon can act as agonists at the M2 and/or M4 subset of muscarinic receptors. In addition, chlorpyrifos may have another site of action. In contrast, none of the organophosphates had any effect on basal or carbachol-stimulated phosphoinositide hydrolysis. The differential activity on these two second messenger systems make it unlikely that the observed effects on cAMP formation are due to increases in endogenous acetylcholine resulting from inhibition of acetylcholinesterase.

Adenylyl Cyclases↗