Diethylstilbestrol potentiates and testosterone antagonizes the action of 3-methylcholanthrene on benzo(a) pyrene metabolism in hep G2 cells

We have used the human hepatoma cell line, Hep G2, to examine the ability of hormones and xenobiotics to modulate the hepatic induction of benzo(a)pyrene hydroxylase and epoxide hydrolase. Hep G2 cells were cultured in Eagle's Minimum Essential Medium supplemented with 10% fetal calf serum. 3-Methylcholanthrene, diethylstilbestrol, testosterone propionate, and combinations of 3-meth-ylcholanthrene, and each of the hormones were added directly to the culture media. We subsequently studied the metabolism of benzo(a)pyrene using cell lysates of the Hep G2 cells. Metabolites were quantitated by high-performance liquid chromatography (HPLC) using fluorodetection. Exposure to 3-methyl-cholanthrene alone resulted in an eightfold increase in total benzo(a)pyrene metabolites with a change of the predominant metabolite from the 3-hydroxy-benzo(a)pyrene to the carcinogenic pathway of the benzo(a)pyrene-7,8-diol. Diethylstilbestrol and testosterone propionate resulted in small, but significant, decreases in metabolism of benzo(a)pyrene. When exposed in combination with 3-methyl-cholanthrene, testosterone propionate antagonized and diethylstilbestrol potentiated the metabolism of benzo(a)pyrene. 3-Methylcholanthrene, diethylstilbestrol, and combinations of 3-methylcholanthrene and diethylstilbestrol or testosterone propionate resulted in increased epoxide hydrolase activity as compared to controls. These results, carried out in a human hepatoma cell line, lend support to a concern for potentiated toxicity and carcinogenicity following exposure to complex chemical mixtures.     

Assessment of Amyloid β Protein in Cerebrospinal Fluid as an Aid in the Diagnosis of Alzheimer's Disease

Abstract: The principal constituent of amyloid plaques found in the brains of individuals with Alzheimer's disease (AD) is a 39–42-amino-acid protein, amyloid β protein (Aβ). This study examined whether the measurement of Aβ levels in CSF has diagnostic value. There were 108 subjects enrolled in this prospective study: AD (n = 39), non-AD controls (dementing diseases/syndromes; n = 20), and other (n = 49). CSF was obtained by lumbar puncture, and Aβ concentrations were determined using a dual monoclonal antibody immunoradiometric sandwich assay. The mean Aβ value for the AD group (15.9 ± 6.8 ng/ml) was not significantly different from that for the non-AD control group (13.0 ± 7.1 ng/ml; p = 0.07), and substantial overlap in results were observed. Aβ values did not correlate with age ( r = −0.05, p = 0.59), severity of cognitive impairment ( r = 0.22, p = 0.21), or duration of AD symptoms ( r = 0.14, p = 0.45). These findings are in conflict with other reports in the literature; discrepant results could be due to the instability of Aβ in CSF. Aβ immunoreactivity decays rapidly under certain conditions, particularly multiple freeze/thaw cycles. Use of a stabilizing sample treatment buffer at the time of lumbar puncture allows storage of CSF without loss of Aβ reactivity. In conclusion, the total CSF Aβ level is not a useful marker for current diagnosis of AD.     

Microtubule Depolymerization Inhibits Ethanol-Induced Enhancement of GABAA Responses in Stably Transfected Cells

Abstract: We studied whether microtubule organization is important for actions of ethanol on GABA_A ergic responses by testing the effects of microtubule depolymerization on ethanol enhancement of GABA action in mouse L(tk⁻) cells stably transfected with GABA_A receptor α₁β₁γ_2L subunits. The microtubule-disrupting agents colchicine, taxol, and vinblastine completely blocked ethanol-induced enhancement of muscimol-stimulated chloride uptake. β-Lumicolchicine, a colchicine analogue that does not disrupt microtubules, had no effect on ethanol action. Colchicine did not alter the potentiating actions of flunitrazepam or pentobarbital on muscimol-stimulated chloride uptake. Thus, colchicine specifically inhibited the potentiating action of ethanol. From these findings, we conclude that intact microtubules are required for ethanol-induced enhancement of GABA_A responses and suggest that a mechanism involving microtubules produces posttranslational modifications that are necessary for ethanol sensitivity in this cell system.     

Differential Inhibition of Secretagogue-Stimulated Sodium Uptake in Adrenal Chromaffin Cells by Activation of D4 and D5 Dopamine Receptors

Abstract: Recent studies have demonstrated that D1-selective and D2-selective dopamine receptor agonists inhibit catecholamine secretion and Ca²⁺ uptake into bovine adrenal chromaffin cells by receptor subtypes that we have identified by PCR as D5, a member of the D1-like dopamine receptor subfamily, and D4, a member of the D2-like dopamine receptor subfamily. The purpose of this study was to determine whether activation of D5 or D4 receptors inhibits influx of Na⁺, which could explain inhibition of secretion and Ca²⁺ uptake by dopamine agonists. D1-selective agonists preferentially inhibited both dimethylphenylpiperazinium- (DMPP) and veratridine-stimulated ²²Na⁺ influx into chromaffin cells. The D1-selective agonists chloro-APB hydrobromide (CI-APB; 100 µ M ) and SKF-38393 (100 µ M ) inhibited DMPP-stimulated Na⁺ uptake by 87.5 ± 2.3 and 59.7 ± 4.5%, respectively, whereas the D2-selective agonist bromocriptine (100 µ M ) inhibited Na⁺ uptake by only 22.9 ± 5.0%. Veratridine-stimulated Na⁺ uptake was inhibited 95.1 ± 3.2 and 25.7 ± 4.7% by 100 µ M CI-APB or bromocriptine, respectively. The effect of CI-APB was concentration dependent. A similar IC₅₀ (∼18 µ M ) for inhibition of both DMPP- and veratridine-stimulated Na⁺ uptake was obtained. The addition of 8-bromo-cyclic AMP (1 m M ) had no effect on either DMPP- or veratridine-stimulated Na⁺ uptake. These observations suggest that D1-selective agonists are inhibiting secretagogue-stimulated Na⁺ uptake in a cyclic AMP-independent manner.     

On the origin of lateral asymmetry

Lateral asymmetry refers to unequal fluorescent intensity between adjacent regions of sister chromatids. It has been observed in the centromeric regions of mitotic chromosomes of mouse or human origin when cells are grown in 5-bromo-2-deoxyuridine (BrdU) for a single round of DNA synthesis. The chromosome-orientation fluorescence in situ hybridization (CO-FISH) technique was used with pseudodiploid mouse cells to show that the regions of asymmetrical brightness coincide with major satellite repetitive DNA, and that the more heavily BrdU-substituted chromatid is the one that fluoresces less brightly. These observations support a 20 year old hypothesis on the origin of lateral asymmetry. Other observations suggest that differential loss of DNA from the heavily substituted chromatid also contributes to lateral asymmetry.     

Foraging ecology of Gentoo Penguins Pygoscelis papua at Macquarie Island during the period of chick care

The diet, diving behaviour, swimming velocity and foraging range of Gentoo Penguins Pygoscelis papua were studied at Macquarie Island during the breeding season in the 1993–1994 austral summer. Gentoo Penguins are considered to be inshore feeders, and at Macquarie Island the diet and estimated foraging ranges supported this. The diet consisted of 91.6% fish and 8.3% squid, by mass. The dominant prey taxa were the fish Gymnoscopelus sp. and Paranotothenia magellanica. A mixture of pelagic and benthic prey was consumed, with a greater proportion of benthic species occurring later in the season. The penguins exhibited a strong diurnal pattern in their diving behaviour. Deep diving (≥30 m) began near sunrise (03.00 h) and finished close to sunset (21.00 h). Diving at night was less common and very shallow (<10 m). Early in the breeding season, dive profiles indicated that birds were probably following vertically migrating pelagic prey through the water column and were foraging in waters over 100 m deep. Later in the season, more uniform, shallower depths were used, suggesting an increase in benthic foraging activity. These changes in dive pattern and depth were consistent with the habitat preferences of prey species found in the diet. Gentoo Penguins swam at 1.04 m per s and had a maximum potential foraging range of about 26 km for single-day trips. They tended to forage within 14 km of the colony, with a mean range of 5.4 km. This range encompassed the deep ocean habitat to the west and east of the island and a shallow area to the north.     

Dust and ammonia in horse stalls with different ventilation rates and bedding

Airborne particles and ammonia were monitored in horse stalls managed under four conditions. Two ventilation rates, high (27 air changes per h) and low (5 air changes per h) and two bedding types, paper and straw, were employed. At both ventilation rates, the number of airborne particles generated while the stalls were mucked out was higher with straw than with paper. Particles were more efficiently cleared at the higher ventilation rate in both the straw and paper stalls. Ammonia measurements reflected an accumulation over time. In the stalls with low ventilation, ammonia levels were significantly higher than in those stalls with high ventilation regardless of bedding type. Management decisions and their relationships to respiratory disorders are discussed.     

Shoot regeneration from tissue-cultured leaves of the American cranberry (Vaccinium macrocarpon)

A method for shoot regeneration from leaf explants in two cultivars of cranberry (Vaccinium macrocarpon Ait.) is described. Modified Anderson's medium supplemented with combinations of thidiazuron (TDZ) with or without 1 M NAA (-naphthaleneacetic acid) was used to optimize shoot regeneration. The effect of light or dark incubation was also determined. Maximum regeneration was obtained in the light in the presence of 10 M TDZ and 1 M NAA. While this medium was suitable for leaf explants obtained from shoot cultures, regeneration did not occur from leaves collected from greenhouse-grown plants. Elongation of the regenerated shoot tips did not occur until explants were transferred to growth regulator-free medium at which time only a minority of shoots elongated. Elongated shoots could be dissected away from leaf tissue, rooted easily, and acclimitized to ambient conditions.Abbreviations NAA -naphthaleneacetic acid - TDZ 1-phenyl-3-(1,2,3-thiadiazol-5-yl) urea