Significant Concentration-related Increase Research Articles

Microplastic-induced damage in early embryonal development of sea urchin Sphaerechinus granularis

Two microplastic sets, polystyrene (PS) and polymethyl methacrylate (PMMA), were tested for adverse effects on early life stages of Sphaerechinus granularis sea urchins. Microparticulate PS (10, 80 and 230 μm diameter) and PMMA (10 and 50 μm diameter) were tested on developing S. granularis embryos from 10 min post-fertilisation (p-f) to the pluteus larval stage (72 h p-f), at concentrations ranging from 0.1 to 5 mg L−1. Both PS and PMMA exposures resulted in significant concentration-related increase of developmental defects and of microplastic uptake in plutei. Moreover, embryo exposures to PS and PMMA (5 and 50 mg L−1) from 10 min to 5 h p-f resulted in a significant increase of cytogenetic abnormalities, expressed as significantly increased mitotic aberrations, while mitotoxicity (as % embryos lacking active mitoses) was observed in embryos exposed to PS, though not to PMMA. When S. granularis sperm suspensions were exposed for 10 min to PS or to PMMA (0.1–5 mg L−1), a significant decrease of fertilisation success was observed following sperm exposure to 0.1 mg L−1 PS, though not to higher PS concentrations nor to PMMA. Sperm pretreatment, however, resulted in significant offspring damage, as excess developmental defects in plutei, both following sperm exposure to PS and PMMA, thus suggesting transmissible damage from sperm pronuclei to the offspring. The overall results point to relevant developmental, cytogenetic and genotoxic effects of PS and PMMA microplastics to S. granularis early life stages, warranting further investigations of other microplastics and other target biota.

Assessment of bendamustine-induced genotoxicity in eukaryotic cells

Bendamustine, an anticancer drug with alkylating properties, is widely used to treat hematological malignancies. Since the nitrogen mustard family alkylators induce DNA damages and have been associated with an elevated risk of second malignancy, current study evaluates the cytotoxic, mutagenic, and recombinogenic effects of bendamustine by using, respectively the mitotic index assay, the in vitro mammalian cell micronucleus test (Mnvit) and the chromosome aberration (CA) test in human peripheral lymphocytes, and the in vivo homozygotization assay in Aspergillus nidulans, which detects the loss of heterozygosity (LOH) due to somatic recombination. Bendamustine (6.0 µg/ml, 9.0 µg/ml, and 12.0 µg/ml) induced a statistically significant concentration-related increase in the frequencies of micronuclei and a significant reduction in the cytokinesis block proliferation index (CBPI) rates when compared to negative control. In the CA test, bendamustine significantly increased the frequencies of structural aberrations at the three tested concentrations when compared to the negative control. Aspergillus nidulans diploids, obtained after bendamustine treatment (6.0 µg/ml, 12.0 µg/ml, and 24.0 µg/ml), produced, after haploidization, homozygotization index (HI) rates higher than 2.0 and significantly different from the negative control. Since bendamustine showed genotoxic effects in all tested concentrations, two of them corresponding to the peak plasma concentrations observed in cancer patients treated with bendamustine, data provided in the current research work may be useful to identify the most appropriate dosage regimen to achieve the efficacy and safety of this anticancer medication.

The effects of perfluorinated chemicals on adipocyte differentiation in vitro

The 3T3-L1 preadipocyte culture system has been used to examine numerous compounds that influence adipocyte differentiation or function. The perfluoroalkyl acids (PFAAs), used as surfactants in a variety of industrial applications, are of concern as environmental contaminants that are detected worldwide in human serum and animal tissues. This study was designed to evaluate the potential for PFAAs to affect adipocyte differentiation and lipid accumulation using mouse 3T3-L1 cells. Cells were treated with perfluorooctanoic acid (PFOA) (5–100 µM), perfluorononanoic acid (PFNA) (5–100 µM), perfluorooctane sulfonate (PFOS) (50–300 µM), perfluorohexane sulfonate (PFHxS) (40–250 µM), the peroxisome proliferator activated receptor (PPAR) PPARα agonist Wyeth-14,643 (WY-14,643), and the PPARγ agonist rosiglitazone. The PPARγ agonist was included as a positive control as this pathway is critical to adipocyte differentiation. The PPARα agonist was included as the PFAA compounds are known activators of this pathway. Cells were assessed morphometrically and biochemically for number, size, and lipid content. RNA was extracted for qPCR analysis of 13 genes selected for their importance in adipocyte differentiation and lipid metabolism. There was a significant concentration-related increase in cell number and decreased cell size after exposure to PFOA, PFHxS, PFOS, and PFNA. All four PFAA treatments produced a concentration-related decrease in the calculated average area occupied by lipid per cell. However, total triglyceride levels per well increased with a concentration-related trend for all compounds, likely due to the increased cell number. Expression of mRNA for the selected genes was affected by all exposures and the specific impacts depended on the particular compound and concentration. Acox1 and Gapdh were upregulated by all six compounds. The strongest overall effect was a nearly 10-fold induction of Scd1 by PFHxS. The sulfonated PFAAs produced numerous, strong changes in gene expression similar to the effects after treatment with the PPARγ agonist rosiglitazone. By comparison, the effects on gene expression were muted for the carboxylated PFAAs and for the PPARα agonist WY-14,643. In summary, all perfluorinated compounds increased cell number, decreased cell size, increased total triglyceride, and altered expression of genes associated with adipocyte differentiation and lipid metabolism.

An adaptation of the human HepaRG cells to the in vitro micronucleus assay

The in vitro micronucleus test is considered as an attractive tool for genotoxicity testing of chemicals because of its simplicity of scoring and wide applicability in different cell types. However, most of the cells currently in use are devoid of the enzyme equipment required for activation of promutagens in the genotoxic metabolites. We postulated that the human HepaRG cell line, which can express xenobiotic metabolising enzymes at levels close to those found in primary human hepatocytes and has retained the indefinite growth capacity of transformed cells, could represent a more suitable model for genotoxicity testing of chemicals requiring metabolic activation. Based on the recommendations of the Organisation for Economic Co-operation and Development test guideline TG 487 for testing of chemicals, HepaRG cell cultures containing >80% mature hepatocytes were treated in situ with various chemicals for 24 h followed by a 3-day mitogenic stimulation with epidermal growth factor without cytokinesis block. In such culture conditions, HepaRG cells underwent >1.5 cell cycle per cell during the mitogenic stimulation. While non-genotoxic compounds (mannitol and staurosporine) did not increase the rate of micronucleated mononucleated cells, all aneugens (colchicine, nocodazole and dichlorodiphenyldichloroethylene) as well as the direct acting clastogen methyl methanesulfonate and clastogens requiring metabolic activation (aflatoxin B1, benzo(a)pyrene and 2-nitrofluorene) induced a statistically significant concentration-related increase in the number of mono-micronucleated cells. The micronucleus test was also performed after 7-day repeat exposure of HepaRG cells to the chemicals. Noticeably, a time-dependent effect was obtained with the three clastogens requiring metabolic activation. In conclusion, our results obtained with HepaRG hepatocytes exposed to various genotoxic compounds requiring or not bioactivation, compared favorably with those reported in various other cell types. They support the view that metabolically competent HepaRG cells have unique potential benefits for testing genotoxic compounds using the in vitro micronucleus assay.

Perinatal programming of obesity later in life by the endocrine disruptor bisphenol A in a mouse model

The 3T3-L1 preadipocyte culture system has been used to examine numerous compounds that influence adipocyte differentiation or function. The perfluoroalkyl acids (PFAAs), used as surfactants in a variety of industrial applications, are of concern as environmental contaminants that are detected worldwide in human serum and animal tissues. This study was designed to evaluate the potential for PFAAs to affect adipocyte differentiation and lipid accumulation using mouse 3T3-L1 cells. Cells were treated with perfluorooctanoic acid (PFOA) (5–100 µM), perfluorononanoic acid (PFNA) (5–100 µM), perfluorooctane sulfonate (PFOS) (50–300 µM), perfluorohexane sulfonate (PFHxS) (40–250 µM), the peroxisome proliferator activated receptor (PPAR) PPARα agonist Wyeth-14,643 (WY-14,643), and the PPARγ agonist rosiglitazone. The PPARγ agonist was included as a positive control as this pathway is critical to adipocyte differentiation. The PPARα agonist was included as the PFAA compounds are known activators of this pathway. Cells were assessed morphometrically and biochemically for number, size, and lipid content. RNA was extracted for qPCR analysis of 13 genes selected for their importance in adipocyte differentiation and lipid metabolism. There was a significant concentration-related increase in cell number and decreased cell size after exposure to PFOA, PFHxS, PFOS, and PFNA. All four PFAA treatments produced a concentration-related decrease in the calculated average area occupied by lipid per cell. However, total triglyceride levels per well increased with a concentration-related trend for all compounds, likely due to the increased cell number. Expression of mRNA for the selected genes was affected by all exposures and the specific impacts depended on the particular compound and concentration. Acox1 and Gapdh were upregulated by all six compounds. The strongest overall effect was a nearly 10-fold induction of Scd1 by PFHxS. The sulfonated PFAAs produced numerous, strong changes in gene expression similar to the effects after treatment with the PPARγ agonist rosiglitazone. By comparison, the effects on gene expression were muted for the carboxylated PFAAs and for the PPARα agonist WY-14,643. In summary, all perfluorinated compounds increased cell number, decreased cell size, increased total triglyceride, and altered expression of genes associated with adipocyte differentiation and lipid metabolism.

Cytogenetic and developmental toxicity of cerium and lanthanum to sea urchin embryos

The aim of this study was to evaluate the toxicity of two rare earth elements (REE), cerium and lanthanum on sea urchin embryos and sperm. Sea urchin ( Paracentrotus lividus) embryos were reared for 72 h in Ce(IV)- or La(III)-contaminated seawater at concentrations ranging from 10 −8 to 10 −5 M. Cleaving embryos (5 h post-fertilization) were submitted to cytogenetic analysis, scoring mitotic activity and a set of mitotic aberrations. Embryological analysis was carried out to determine percent developmental anomalies and/or embryonic mortality. P. lividus sperm were suspended in Ce(IV) or La(III) (10 −8–10 −5 M) for 1 h, and percent fertilized eggs were scored in cleaving embryos that were cultured up to pluteus stage to score any developmental defects. Embryos reared in 10 −5 M Ce(IV) resulted in 100% embryonic mortality, whereas 10 −5 M La(III) induced 100% developmental defects, without causing any embryonic mortality. A significant concentration-related mitotoxic effect and induction of mitotic aberrations were observed in Ce(IV)-exposed, but not in La(III)-exposed embryos, at concentrations ranging from 10 −7 M to 3 × 10 −6 M. Following sperm exposure, both Ce(IV) and La(III) induced a decrease in sperm fertilization success at the highest tested concentration (10 −5 M). The offspring of Ce(IV)-exposed, but not of La(III)-exposed sperm displayed a significant concentration-related increase in developmental defects. The results may suggest adverse impacts in REE-exposed biota and warrant further studies of a more extended REE series.

Micronuclei induced by airborne particulate matter from Mexico City

Particulate air pollution is an important environmental health risk. In the present study, we have investigated the ability of chemically characterized water and organic-soluble extracts of PM 10 from two different regions of Mexico City to induce micronuclei in a human epithelial cell line. We also evaluated the association between the chemical characteristics of the PM and its genotoxicity. The airborne particulate samples were collected from an industrial and a residential region; a Hi-Vol air sampler was used to collect PM 10 on glass fiber filters. PM mass was determined by gravimetric analysis of the filters. One section of each PM 10 filter was agitated either with deionized water to extract water-soluble compounds or with dichloromethane to prepare organic-soluble compounds. The chemical composition of the extracts was determined by ion and gas chromatography and atomic adsorption spectroscopy. A549-human alveolar epithelial cells were exposed to different concentrations of PM 10 extracts and the cytokinesis blocked micronucleus assay was performed to measure DNA damage. Even though the industrial region had a higher PM concentration, higher amounts of metals and PAHs were found in the residential area. Both industrial and residential extracts induced a significant concentration-related increase in the micronuclei frequency. The PM 10 water-soluble industrial extract induced significantly more micronuclei than the one of the residential region; inversely, the organic residential extract induced more micronuclei than the one from the industrial region. The association between the induction of micronuclei and the chemical components obtained by the comparative analysis of standardized regression coefficients showed that cadmium and PAHs were significantly associated with micronuclei induction. Data indicate that water-soluble metals and the organic-soluble fraction of PM 10 are both important in the production of micronuclei. Effects observed, point to the risk of PM exposure and shows the need of integrative studies.

N-nitrosodiethylamine initiation of carcinogenesis in Japanese medaka (Oryzias latipes): hepatocellular proliferation, toxicity, and neoplastic lesions resulting from short term, low level exposure.

To investigate relationships among carcinogen exposure, cell proliferation, and carcinogenesis, 14-day post-hatch Japanese medaka (Oryzias latipes) were exposed to 0, 10, 25, 50, or 100 ppm N-nitrosodiethylamine (DEN) for 48 h under static renewal conditions. They were then held in clean water until sampling at 3 and 6 months. The frequencies of hepatic lesions and neoplasms were determined from hematoxylin/eosin-stained paraffin sections. A significant (p < 0.0001) concentration-related increase in hepatic vacuolated foci occurred in 3- and 6-month samples, with males having a significantly (p = 0.02) higher incidence than females. Concentration-related increases in degenerative lesions were documented for spongiosis hepatis at 3 months (p = 0.053) and hepatic vacuoles at 6 months (p = 0.005). There was a significant (p = 0.0001) concentration-related increase in macrophage aggregates at 6 months. Basophilic foci were significantly related (p < 0.0001) to DEN concentration at 3 months post-exposure and were unaffected by gender or age. At both 3 and 6 months, there were significant concentration-related increases in hepatocellular carcinoma (p < or = 0.02). Hepatocyte proliferation in 3-month whole specimens was quantified using an immunohistochemical assay for proliferating-cell nuclear antigen. Trend tests and a probit dose-response model showed a significantly positive correlation (p = 0.015) between proliferating hepatocytes and DEN concentrations. These results confirm that short-term exposure to low and moderate levels of DEN initiates concentration-dependent carcinogenic effects in medaka that are apparent at 3 months postexposure. DEN could be an effective initiator in an initiation/promotion assay for medaka using a 48-h exposure period, DEN concentrations < or = 10 ppm, and a 6-month sampling period.

Comparison of Local and Systemic Ethanol Effects on Extracellular Dopamine Concentration in Rat Nucleus Accumbens by Microdialysis

To determine the site of action of systemic ethanol on dopaminergic function in the nucleus accumbens, we compared the effect of intraperitoneal (ip) and local administration of ethanol on interstitial dopamine concentration using microdialysis in freely moving rats. The ip administration of 1 g/kg of ethanol significantly increased the dialysate dopamine (DA) concentrations (∼40% above basal), compared with saline treatment. The concentration-time profile of DA and ethanol in dialysates was similar after two ethanol injections 4 hr apart. Local perfusion with several ethanol concentrations showed that 510 and 860 mM of ethanol caused a significant concentration-related increase in extracellular DA concentrations in the nucleus accumbens (510 mM, 28% increase; 860 mM, 62% increase). However, lower ethanol concentrations, 170 mM or below, failed to change basal DA concentrations. Stimulation with high potassium (50 mM) in artificial cerebrospinal fluid preceding local ethanol treatment increased dialysate DA concentrations to 523 ± 83% of basal levels, confirming that the DA terminals were responsive to pharmacological manipulation. Basal DA levels in dialysates were ∼70% calcium-dependent when tested at the end of the local perfusion experiments. Stereological examination of the nucleus accumbens revealed probe-induced damage, but did not detect additional damage by local perfusion of ethanol. When ethanol concentrations in the DA sampling area around the probe are taken into account in both systemic and local administration experiments, this study suggests that concentrations of ethanol associated with moderate intoxication do not directly affect the function of DA terminals in the nucleus accumbens. Therefore, the systemic effects of ethanol on nucleus accumbens DAergic function is more likely due to an interaction with sites other than the nucleus accumbens.

Comparison of Local and Systemic Ethanol Effects on Extracellular Dopamine Concentration in Rat Nucleus Accumbens by Microdialysis

To determine the site of action of systemic ethanol on dopaminergic function in the nucleus accumbens, we compared the effect of intraperitoneal (i.p.) and local administration of ethanol on interstitial dopamine concentration using microdialysis in freely moving rats. The i.p. administration of 1 g/kg of ethanol significantly increased the dialysate dopamine (DA) concentrations (approximately 40% above basal), compared with saline treatment. The concentration-time profile of DA and ethanol in dialysates was similar after two ethanol injections 4 hr apart. Local perfusion with several ethanol concentrations showed that 510 and 860 mM of ethanol caused a significant concentration-related increase in extracellular DA concentrations in the nucleus accumbens (510 mM, 28% increase; 860 mM, 62% increase). However, lower ethanol concentrations, 170 mM or below, failed to change basal DA concentrations. Stimulation with high potassium (50 mM) in artificial cerebrospinal fluid preceding local ethanol treatment increased dialysate DA concentrations to 523 +/- 83% of basal levels, confirming that the DA terminals were responsive to pharmacological manipulation. Basal DA levels in dialysates were approximately 70% calcium-dependent when tested at the end of the local perfusion experiments. Stereological examination of the nucleus accumbens revealed probe-induced damage, but did not detect additional damage by local perfusion of ethanol. When ethanol concentrations in the DA sampling area around the probe are taken into account in both systemic and local administration experiments, this study suggests that concentrations of ethanol associated with moderate intoxication do not directly affect the function of DA terminals in the nucleus accumbens. Therefore, the systemic effects of ethanol on nucleus accumbens DAergic function is more likely due to an interaction with sites other than the nucleus accumbens.

Investigations on the in vitro and in vivo genotoxic potential of 5-vinyl-2-norbornene.

5-Vinyl-2-norbornene (VNB: CAS no. 3048-64-4), an industrial chemical that produces hyaline droplet nephropathy in the male rat with associated alpha2u-globulin increases, was investigated in vitro and in vivo for its genotoxic potential. A Salmonella typhimurium reverse mutation assay (strains TA98, 100, 1535, 1537, 1538) was negative both without (dose range 0.003-0.3 mg per plate) and with (0.003-0.3 mg per plate) metabolic activation. A forward gene mutation test in Chinese Hamster Ovary (CHO) cells (HGPRT locus) did not show any significant concentration-related increases in mutation frequencies in the absence (0.01-0.1 mg ml[-1]) or presence (0.005-0.1 mg ml[-1]) of metabolic activation. In a sister chromatid exchange (SCE) test, VNB did not produce statistically significant or dose-related increases in the incidence of SCEs in the absence (0.02-0.06 mg ml[-1]) or presence (0.005-0.03 mg ml[-1]) of metabolic activation. A bone marrow chromosome aberration test was conducted in groups of 10 male and 10 female Sprague-Dawley rats exposed for 6 hr/day for 5 consecutive days to mean (+/- SD) VNB vapor concentrations of 0 (air control), 48.1 +/- 1.29, 146 +/- 9.2, or 336 +/- 8.5 ppm. Marrow was collected 6 and 24 hr after the final exposure. No statistically significant or dose-related increases in chromosomal aberrations occurred in the VNB-exposed animals. 5-Vinyl-2-norbornene did not show any potential for genotoxic activity with this in vitro-in vivo battery of tests.

Inhalation studies of the genotoxicity of trichloroethylene to rodents

Trichloroethylene (TCE) (CAS No. 79-01-6) is an industrial solvent used in degreasing, dry cleaning, and numerous other medical and industrial processes. Controlled inhalation studies were performed using male C57BL/6 mice and CD rats to determine if TCE can induce cytogenetic damage in vivo. Animals were exposed in groups of five to target concentrations of either 0, 5, 500, or 5000 ppm TCE for 6 h. Tissue samples were taken between 18 and 19 h post exposure. Peripheral blood lymphocytes (PBLs) in rats and splenocytes in mice were cultured and analyzed for the induction of sister-chromatid exchanges, chromosome aberrations, and micronuclei (MN) in cytochalasin B-blocked binucleated cells. Bone marrow polychromatic erythrocytes (PCEs) were analyzed for MN. The only positive response observed was for MN in rat bone marrow PCEs. TCE caused a statistically significant increase in MN at all concentrations, inducing an approximate fourfold increase over control levels at 5000 ppm. TCE was also cytotoxic in rats, causing a significant concentration-related decrease in the ratio of PCEs/normochromatic erythrocytes. This study indicates that there may be species-specific cytogenetic effects attributed to TCE inhalation exposure. In follow-up studies, CD rats were exposed for 6 h/day over 4 consecutive days to either 0, 5, 50, or 500 ppm TCE. No statistically significant concentration-related increases in cytogenetic damage were observed. While the MN frequencies in the 4-day study were comparable to those at the equivalent concentrations in the 1-day study, they were not significantly elevated due to an unusually high MN frequency in the controls. A subsequent replication of the 1-day 5000 ppm TCE exposure with rats again showed a highly significant increase in MN frequencies compared to concurrent controls.

Muscarinic receptor-mediated increase in extracellular inositol 1,4,5-trisphosphate levels in the rat hippocampus: an in vivo microdialysis study.

The extracellular concentration of inositol 1,4,5-trisphosphate (IP3) has been monitored in the ventral hippocampus of the anesthetized rat by using a microdialysis technique coupled to a radioreceptor assay. Three hours after the implantation of the cannula, basal extracellular concentration of IP3 (corrected for a 9% recovery) was 71 nM (0.39 pmol/60-microliters fraction) and remained stable for at least 5 h. Local infusion of carbachol for 60 min caused a significant concentration-related increase in extracellular IP3 levels (0, 24, and 57% at 1, 50, and 100 microM, respectively). Acetylcholine (100 microM) and muscarine (100 microM) increased IP3 outflow by 40 and 42%, respectively. The effect of carbachol was fully prevented by coinfusion of 10 microM pirenzepine and reduced by 1 microM tetrodotoxin indicating that the carbachol response is mediated by neuronal muscarinic receptors. These data demonstrate the feasibility of using microdialysis and a radioreceptor assay to measure IP3 in the extracellular space. This approach could prove useful for the study of the in vivo operation of muscarinic and, by extension, a number of receptors coupled to phosphoinositide turnover.

Neuromedin N is a potent modulator of dopamine D 2 receptor agonist binding in rat neostriatal membranes

In the concentration range of 1–10 nM, neuromedin N produced a significant concentration-related increase in the K d values of [ 3 H] l -(−)-N- propylnorapomorphine binding sites in rat neostriatal membranes with a peak action at 10 nM (36% increase versus the control group mean value). The B max values were not affected by neuromedin N. Neurotensin at 10 nM induced an increase in the K d values, which was not affected by a threshold concentration of neuromedin N (0.1 nM). In view of the higher potency of neuromedin N versus neurotensin to modulate neostriatal D 2 receptors in contrast to the higher potency of neurotensin versus neuromedin N to bind to the cloned neurotensin receptors, it seems possible that the neuromedin N activated neostriatal neurotensin receptors controlling the D 2 receptors represent a distinct subtype of neurotensin receptors.

P 1(A 2/Ra)-Purinoceptors may Mediate the Stimulatory Effect of Adenosine and Adenosine Analogs on Acid Formation in Isolated Rabbit Parietal Cells

The effects of adenosine (ADO) and its analogs 2-chloroadenosine (2CADO) and 5′-N-ethyl-carboxamido adenosine (NECA) on acid production, as measured by the [ 14C]-aminopyrine (AP) accumulation method, were studied in rabbit gastric parietal cells prepared by enzymatic dispersion and enriched by counterflow elutriation. ADO (1 μM to 1 mM), 2CADO (1 μM to 1 mM) and NECA (0.1 μM to 100 μM) caused significant concentration-related increases on AP uptake by resting parietal cells. The order of potency was NECA⪢2CADO≥ADO. Histamine and dibutyryl-cAMP (db-cAMP), known secretagogues, stimulated AP accumulation. In both, histamine- and db-cAMP-stimulated parietal cells, ADO, 2CADO and NECA at the above concentration ranges induced concentrationrelated increases of the respective AP accumulation raised-rates. The rank order of potency in both stimulated conditions was: NECA>2CADO⪢ADO. Cimetidine, a well known H 2 histamine receptor antagonist, at concentration of 100 μM caused potent inhibition of histamine-stimulated AP accumulation without affecting the stimulatory responses to db-cAMP. The increases of AP accumulation elicited by NECA, the most potent purine tested, in both resting and histamine- or db-cAMP-stimulated parietal cells were not significantly modified by cimetidine 100 μM. Theophylline (0.1 μM to 10 μM), a non-specific P 1-purinoceptor antagonist, caused significant inhibition of the NECA responses in both resting and histaminestimulated parietal cells. The concentration order of potency of theophylline was: 0.1 μM>1 μM>10 μM. These results suggest that there are P 1(A 2/Ra)-purinoceptors on rabbit parietal cells which mediate the stimulatory effects of adenosine and analogs on gastric acid production.

Cytogenetic studies of mice exposed to styrene by inhalation

The data for the in vivo genotoxicity of styrene (STY) are equivocal. To evaluate the clastogenicity and sister-chromatid exchange (SCE)-inducing potential of STY in vivo under carefully controlled conditions, B 6C 3F 1 female mice were exposed by inhalation for 6 h/day for 14 consecutive days to either 0, 125, 250 or 500 ppm STY. One day after the final exposure, peripheral blood, spleen, and lungs were removed and cells were cultured for the analysis of micronucleus (MN) induction using the cytochalasin B-block method, chromosome breakage, and SCE induction. Peripheral blood smears were also made for scoring MN in erythrocytes. There was a significant concentration-related elevation of SCE frequency in lymphocytes from the spleen and the peripheral blood as well as in cells from the lung. However, no statistically significant concentration-related increases were found in the frequency of chromosome aberrations in the cultured splenocytes or lung cells, and no significant increases in MN frequencies were observed in binucleated splenocytes or normochromatic erythrocytes in peripheral blood smears.

Subchronic Vapor Inhalation Toxicity Studies on 3.1 Oil in Male Fischer 344 Rats

3.1 Oil is a chlorofluorocarbon oil consisting of C6 and C8 oligomers of chlorotriflu-oroethylene. Physical properties such as high thermal stability, good lubricity, and high dielectric strength make 3.1 oil promising for potential use in hydraulic fluid systems. A previous inhalation study indicated that the liver is the target organ of 3.1 oil toxicity. Cross liver hypertrophy and microscopic hepatocytomegaly were the prime lesions noted. To determine a no-effect concentration, male Fischer 344 rats were exposed to air only, or vapor concentrations of 10, 50, or 250 mg 3.1 oil/m3 for 13 wk. Mean body weights of the highest concentration test group were slightly, but not significantly, depressed during the final two-thirds of the study. Alkaline phosphatase and blood urea nitrogen values were increased in this group at sacrifice. Significant concentration-related increases in relative kidney and liver weights occurred in the 250 and 50 mg 3.1 oil/m3 groups. Hyaline droplet distribution and severity was greater in the high-exposure group of rats. Hepatocytomegaly was a common finding in all rats exposed at these concentrations; however, this lesion resolved by dmo postexposure. Examination of hepatocytic ultrastructure revealed slightly swollen mitochondria in the two highest exposure groups. The number of peroxisomes per visual field was significantly greater in the highest exposure group only. The associated activity of peroxisomal β-oxidation of palmitoyl coenzyme A (CoA) also showed a concentration-dependent increase. Although an increase in this enzyme was noted, 70 mg 3.7 oil/m3 was identified as a no-effect level based on light and electron microscopic, hematologic, and in-life data.

Characterization of the effects of adenosine, adenosine 5′-triphosphate and related purines on acid secretion in isolated rabbit gastric glands

The influence of adenosine, AMP, ADP, ATP, the adenosine analogue l-PIA and the ATP analogue β,γ-methylene ATP, on gastric acid secretion, as measured by the aminopyrine accumulation method, in resting and histamine-stimulated glands isolated from rabbit gastric mucosa was studied. In resting glands, adenosine and its analogue l-PIA (10 μ m-1 m m) caused significant concentration-related increases of the basal H + secretion, whereas no changes were obtained in response to the other purines tested. In histamine-stimulated glands, adenosine, l-PIA and AMP (10 μ m-1 m m) induced concentration-related increases of the H + secretory rate, whereas ATP, β,γ-methylene ATP and ADP (10 μ m-1 m m) produced concentration-related decreases of the H + raised rate. The rank order of potency of the purine compounds in increasing the stimulated H + secretion was: adenosine > l-PIA ≫ AMP, and in decreasing it was: ATP> ADP> β,γ-methylene ATP. The stimulatory responses to adenosine were inhibited by theophylline (10 μ m–100 μ m) and caffeine (10 μ m-1 m m); whereas, the inhibitory responses to ATP were significantly reduced by the well known prostaglandin synthesis inhibitor indomethacin (1 μ m–100 μ m). From the results it is concluded that in isolated rabbit gastric glands, purine compounds are effective modulators of the gastric H + secretory process. The pattern of purine activity obtained suggests that the stimulatory responses, inhibited by methylxanthines, may be mediated via P 1-purinoceptors, while the inhibitory responses, reduced by indomethacin, may be mediated via P 2-purinoceptors.

The effects of neomycin on membrane properties and discharge activity of an isolated sensory neuron

The effects of neomycin sulfate were examined upon the discharge activity and electrical membrane properties of an isolated invertebrate sensory neuron, the crayfish stretch receptor neuron. Neomycin depressed cell discharge activity in a concentration-dependent manner over the concentration range of 0.01-1.0 mM. Significant concentration-related increases were observed in the resting membrane potential and the width of the orthodromic action potential. There was a significant concentration-dependent decrease in the fast rising phase of the antidromic action potential. Significant changes also were observed in other electrical properties such as membrane resistance, but these were found not to be concentration related. The most significant change was membrane hyperpolarization, which could account for the depression of cell discharge activity. The observed changes are consistent with a neomycin-induced change in the membrane potassium conductance. It is proposed that the neural effect of neomycin is a selective interaction with the neuronal membrane phospholipids.

Induction of cytogenetic damage in rodents after short-term inhalation of benzene.

Experiments were designed to investigate both the induction of sister chromatid exchanges (SCEs) in peripheral blood lymphocytes (PBLs) and micronuclei (MN) in bone marrow polychromatic erythrocytes (PCEs) of mice and rats after inhalation of benzene (BZ). Male DBA/2 mice (17-19 weeks old) were exposed to target concentrations of either 0, 10, 100, or 1,000 ppm BZ for 6 hr. Male Sprague-Dawley rats (11-14 weeks old) were exposed to target concentrations of either 0, 0.1, 0.3, 1, 3, 10, or 30 ppm BZ for 6 hr. Blood was obtained by cardiac puncture 18 hr after exposure, and PBLs were cultured in the presence of lipopolysaccharide (mouse B cells, 60 micrograms/ml) or concanavalin A (rat T cells, 30 micrograms/ml) to stimulate blastogenesis for SCE analysis. Femoral bone marrow smears from both species were analyzed for MN in PCEs 18 hr after BZ exposure. Mouse PBLs revealed a significant concentration-related increase in the SCE frequency over controls at 10, 100, or 1,000 ppm BZ. Mouse bone marrow showed a significant concentration-dependent increase in MN over controls after exposure to 10, 100, or 1,000 ppm BZ. Rat PBLs showed a significant increase in the SCE frequency after exposure to 3, 10, or 30 ppm BZ. The statistical significance of the 1 ppm BZ result was borderline and dependent on the statistical test chosen. Rat cells revealed a significant concentration-related increase in MN after inhalation of either 1, 3, 10, or 30 ppm BZ. PBLs from treated mice showed significant concentration-dependent decreases in mitotic indices; however, cell cycle kinetics and leucocyte counts remained unaffected. Rat PBLs showed significant decreases in mitotic activity only after exposure to 3 and 30 ppm BZ, whereas cell cycle kinetics and leucocyte counts were unaffected. These results show that BZ can induce statistically significant cytogenetic effects in PBLs and PCEs of both mice and rats after a 6-hr inhalation of BZ at low concentrations.