Changes in auditory brainstem response in rats chronically exposed to carbon disulfide

The chronic effect of carbon disulfide (CS₂) on the central nervous system (CNS) was studied by examining auditory brainstem responses (ABRs) in female rats (Jcl Wistar) exposed to 200 ppm or 800 ppm CS₂ by inhalation, 6 h a day, 5 days a week, for 15 weeks. Two modes of ABRs evoked by clicks at 61 and 96 dB sound pressure levels (61 dB-ABR and 96 dB-ABR) were recorded during the exposure and for 6 weeks afterwards. Three main components (I, III and V) of ABRs were analyzed from the latencies and differences between latencies of them (interpeak latencies, IPL I–III, IPL III–V and IPL I–V). The latencies of the three components and IPLs of 96 dB-ABR in rats group exposed to 800 ppm of CS₂ were significantly delayed during the exposure period. The delay of latency of component V and IPL III–V and I–V tended to increase with exposure time. At 61 dB-ABR, the changes in the latency of component V, IPL III–V and I–V resembled those at 96 dB-ABR. For the rats group exposed to 200 ppm CS₂, the latency of component I, IPL III–V and I–V at 96 dB-ABR were delayed significantly but transiently during the exposure period. For both groups, recovery from the latencies of the three components and IPLs of ABR was observed by the end of the recovery period. The delayed latencies of ABR observed in rats exposed to 800 ppm CS₂ suggested a conduction dysfunction in the brainstem due to CS₂ exposure. The transient delay of the parameters in the rats group exposed to 200 ppm CS₂ was considered to represent a slight conduction dysfunction.     

Studies on the rate of incorporation of tryptophan into nicotinamide-adenine dinucleotides in rats chronically exposed to carbon disulphide.

Previous experiments have revealed changes in the metabolism of nieotinic acid and nicotinamide-adenine dinucleotides due to CS₂ intoxication. In this study the incorporation of tryptophan into nicotinamide-adenine dinucleotides in the liver of rats chronically intoxicated with CS₂ has been measured. The concentration and radioactivity of NAD in the liver of rats were determined 5 hr after administration of a loading dose of tryptophan and 2 hr after injection of a trace dose of [¹⁴C]tryptophan. It was found that tryptophan was more extensively incorporated into nucleotides in the liver of rats intoxicated with CS₂. This indicates, that tryptophan, rather than nicotinic acid, is the main precursor of NAD synthesis in animals intoxicated with CS₂.     

Bone metabolism of male rats chronically exposed to cadmium

Recently, based on a female rat model of human exposure, we have reported that low-level chronic exposure to cadmium (Cd) has an injurious effect on the skeleton. The purpose of the current study was to investigate whether the exposure may also affect bone metabolism in a male rat model and to estimate the gender-related differences in the bone effect of Cd. Young male Wistar rats received drinking water containing 0, 1, 5, or 50 mg Cd/l for 12 months. The bone effect of Cd was evaluated using bone densitometry and biochemical markers of bone turnover. Renal handling of calcium (Ca) and phosphate, and serum concentrations of vitamin D metabolites, calcitonin, and parathormone were estimated as well. At treatment with 1 mg Cd/l, corresponding to the low environmental exposure in non-Cd-polluted areas, the bone mineral content (BMC) and density (BMD) at the femur and lumbar spine (L1-L5) and the total skeleton BMD did not differ compared to control. However, from the 6th month of the exposure, the Z score BMD indicated osteopenia in some animals and after 12 months the bone resorption very clearly tended to an increase. The rats' exposure corresponding to human moderate (5 mg Cd/l) and especially relatively high (50 mg Cd/l) exposure dose- and duration-dependently disturbed the processes of bone turnover and bone mass accumulation leading to formation of less dense than normal bone tissue. The effects were accompanied by changes in the serum concentration of calciotropic hormones and disorders in Ca and phosphate metabolism. It can be concluded that low environmental exposure to Cd may be only a subtle risk factor for skeletal demineralization in men. The results together with our previous findings based on an analogous model using female rats give clear evidence that males are less vulnerable to the bone effects of Cd compared to females.     

Effect of exposure to carbon disulfide on tryptophan metabolism and the tissue vitamin B6 contents of rats

Female Wistar rats were exposed to 100 ppm or 600 ppm carbon disulfide for 6 hours a day, 5 days a week for 12 weeks. During the exposure period, their urinary excretion of 4-pyridoxic acid was determined. The urinary excretions of xanthurenic acid and kynurenic acid after tryptophan loading were also determined. At the end of the exposure period, the rats were sacrificed and the levels of the five major forms of vitamin B₆, pyridoxine, pyridoxal, pyridoxamine, pyridoxal phosphate and pyridoxamine phosphate, in the liver, kidneys and brain were determined by HPLC fluorometry. During the exposure the urinary excretions of xanthurenic acid and kynurenic acid after i. p. administration of tryptophan increased significantly in both experimental groups. However, urinary excretion of 4-pyridoxic acid decreased only slightly in the group exposed to carbon disulfide at 600 ppm and did not decrease in the group exposed to 100 ppm carbon disulfide. Furthermore, no significant changes were observed in the contents of vitamins B₆ in any tissues examined. These results indicate that carbon disulfide does not cause vitamin B₆ deficiency and thus that the disorders of tryptophan metabolism induced by carbon disulfide intoxication are not due to vitamin B₆ deficiency.     

Disorders in bone metabolism of female rats chronically exposed to cadmium

The effect of cadmium (Cd) on bone metabolism during skeletal development and maturity was investigated on a rat model of human exposure. Young female Wistar rats were exposed to 1, 5, or 50 mg Cd/l in drinking water for 3, 6, 9, and 12 months. Total bone mineral density (T-BMD), bone mineral content (BMC), density (BMD), and bone area at the femur and lumbar spine (L1-L5) were measured densitometrically. Alkaline phosphatase (ALP) and osteocalcin (OC) as bone formation markers, and carboxy-terminal cross-linking telopeptides of type I collagen (CTX) in bone (trabecular and cortical) or serum as bone resorption markers were measured. Renal calcium (Ca) handling and Cd body burden were evaluated as well. At the stage of intensive skeletal development (the first 6 months of the experiment), at all exposure levels, Cd inhibited the processes of bone formation and as a result disturbed the accumulation of bone mass leading to osteopenia (- 1 > Z score/T score BMD > -2.5) and at 5 and 50 mg Cd/l even to more advanced disorders in the BMD. Continuation of the exposure up to skeletal maturity led to high bone turnover with increased resorption enhancing the prevalence of osteopenia or the BMD values having the Z score/T score < -2.5. The results allow for the conclusion that chronic, even low-level exposure to Cd disturbs bone metabolism during skeletal development and maturity by affecting bone turnover most probably through a direct influence on bone formation and resorption, and indirectly via disorders in Ca metabolism. Our findings confirm the hypothesis that environmental exposure to Cd may be a risk factor for low BMD.     

Effect of zinc supplementation on bone metabolism in male rats chronically exposed to cadmium

The aim of the present study is to investigate, based on the rat model of moderate and relatively high human exposure to cadmium (Cd), whether zinc (Zn) supplementation may prevent Cd-induced disorders in bone metabolism. For this purpose, male Wistar rats received Cd (5 and 50mg/l) or/and Zn (30 and 60mg/l) in drinking water for 6 and 12 months. Bone densitometry and biochemical markers of bone turnover were used to assess the effects of Cd or/and Zn. Bone mineral content (BMC) and density (BMD) were measured in the femur. Serum osteocalcin (OC) and alkaline phosphatase in trabecular (bT-ALP) and cortical (bC-ALP) bone were determined as bone formation markers, and carboxy-terminal cross-linking telopeptides of type I collagen (CTX) in serum were measured as bone resorption marker. Serum concentration of calcium (Ca) and its renal handling, as well as Zn and Cd concentrations in the serum/blood, urine and femur were evaluated as well. The exposure to 5 and 50mg Cd/l (0.340+/-0.026 and 2.498+/-0.093mg Cd/kg body wt/24h, respectively), in a dose and duration dependent manner, affected bone turnover (inhibited bone formation and stimulated its resorption) and disturbed bone mineralization (decreased BMC, BMD and Zn concentration). Zn supply at the concentration of 30 and 60mg/l (1.904+/-0.123 and 3.699+/-0.213mg/kg body wt/24h, respectively) during Cd exposure influenced the Cd-induced disorders in bone metabolism. Zn administration to the Cd-exposed rats enhanced the bone ALP activity and prevented Cd-induced bone resorption, but had no statistically significant effect on BMC and BMD; however, mean values of the densitometric parameters in the rats receiving both Cd and Zn were higher than in those treated with Cd alone. Moreover, Zn supplementation at both levels of Cd exposure was found to prevent Cd accumulation in the femur and the Cd-induced decrease in bone Zn concentration. The results of the present study allow the conclusion that Zn supplementation during Cd exposure may partly protect from disorders in bone metabolism. The influence of Zn may be accompanied by its ability to prevent Cd-induced Zn deficiency and to decrease Cd accumulation in bone tissue. The findings seem to indicate that enhanced dietary intake of Zn in subjects chronically exposed to moderate and relatively high Cd levels may have a protective influence on the skeleton.     

Blood-bound carbon disulfide: An indicator of carbon disulfide exposure, and its accumulation in repeatedly exposed rats

Carbon disulfide is present in exposed subjects in free and bound or acid-labile forms. Sensitivities of the blood acid-labile CS2 (AL CS2) concentration and the modified iodine-azide test (IAT) were compared as indicators of CS2 exposure. Rats were exposed to 15 (approximately 5 ppm), 30, 60, or 120 mg/m3 of CS2. Exposure to 15 or 30 mg/m3 of CS2 could not be detected by the modified IAT. However, a linear relationship between blood CS2 (free or AL CS2) concentrations and these exposure levels was observed. Free CS2 is eliminated rapidly, while AL CS2 is eliminated very slowly from the exposed subjects. Repetitive daily exposures (8 hr/day) to 120 mg/m3 of CS2 were carried out in rats. Blood AL CS2 concentrations in exposed rats increased with each successive exposure while the free CS2 level remained relatively constant. By the sixth or seventh daily exposure the blood AL CS2 concentration was about 2.5 times that of the first 8-hr exposure and about 3 times the level of free CS2. These results indicated an appreciable accumulation of CS2 in subjects repeatedly exposed to low concentrations of the solvent. Rats were also exposed to CS2 8 hr/day for 5 days. After a 2-day nonexposure period (Days 6 and 7), the animals were reexposed on Day 8. The blood AL CS2 concentration in animals exposed on Day 8 was substantially higher than in those that received a single 8-hr exposure (Day 1), despite the hiatus on Days 6 and 7. These results indicated that blood AL CS2 was not totally eliminated during the 2-day nonexposure period. In in vitro experiments, the binding profile of CS2 to human blood was remarkably similar to that of rats exposed to CS2 by inhalation.     

The influence of simultaneous exposure to carbon disulfide and hydrogen sulfide on the peripheral nerve toxicity and metabolism of carbon disulfide in rats

Three groups of 10 male Sprague-Dawley rats were exposed daily, 5 days a week for 25 weeks, either to 500 ppm carbon disulfide (CS2), 50 ppm hydrogen sulfide (H2S), or to both of them as a mixture and were periodically examined for sensory and motor tail nerve conduction velocity (SNCV, MNCV). A concomitant control group of 10 rats was used. In addition, rats exposed to 500 ppm CS2, and those simultaneously exposed to 500 ppm CS2 and 50 ppm H2S, were twice examined for 24-h urine excretion of 2-thio-thiazolidine-4-carboxylic acid (TTCA) in the course of the experimental period. Simultaneous exposure to CS2 and H2S had no significant interactive effect on nerve conduction velocities. A significant time-dependent slowing down of MNCV and SNCV occurred as the result of chronic exposure to CS2, including exposure to 500 ppm CS2 and to the mixture of 500 ppm CS2 and 50 ppm H2S, but did not occur after chronic exposure to 50 ppm H2S. With combined exposure to 500 ppm CS2 and 50 ppm H2S, the quantity of TTCA excreted in 24-h urine was not significantly different from that occurring in response to CS2 exposure alone. On the basis of these results it is suggested that chronic exposure to H2S would neither influence CS2-induced peripheral nerve toxicity nor obscure the interpretation of the measurement of urinary TTCA as a biological indicator of CS2 exposure.     

Hematological changes in rats chronically exposed to oral aluminum

This study was aimed to investigate the effects of the long-term oral exposure to aluminum sulfate on hematological parameters in rats. For this purpose, 24 adult female Wistar rats were divided in three groups with 8 animals each (control, citrate, and citrate plus aluminum groups). Rats from control and citrate groups had free access to tap water and to a sodium citrate solution (35 mM), respectively. Rats from citrate plus aluminum group received, as unique source of liquid, an aluminum sulfate solution (30 mM) diluted in the above-mentioned sodium citrate solution, ad libitum. After the treatment period (18 months), aluminum-exposed rats showed a significant decrease in the number of red blood cells, blood hemoglobin concentration and hematocrit when compared to rats from the control group. Serum iron levels were also significantly lower in citrate plus aluminum group, whereas total iron binding capacity did not change after citrate plus aluminum exposure. Erythrocyte thiobarbituric acid-reactive substances (TBARS) and nonprotein thiols (NPSH) levels, erythrocyte osmotic fragility and hepatic delta-aminolevulinic acid dehydratase (delta-ALA-D) activity did not change after treatment with citrate plus aluminum. Conversely, aluminum exposure increased delta-ALA-D activity in bone marrow. The present results indicate that long-term oral exposure to low doses of aluminum sulfate promotes alterations on erythrocyte parameters in rats, probably as a consequence of alterations in the iron status. In addition, although the details of the underlying mechanism remain unclear, our study reports, for the first time, a stimulatory effect of chronic aluminum exposure on bone marrow delta-ALA-D activity.     

Activation of brain tyrosine hydroxylase in rats exposed to carbon disulfide and sodium diethyldithiocarbamate.

A time-dependent activation of brain tyrosine hydroxylase (TH) was observed in rats exposed by inhalation daily for 4 hr to carbon disulfide (5 mg/liter). TH activation rose above control after the second day of exposure, reached 140% of control by the fourth day, and declined thereafter. Activation of TH was also observed in rats given sodium diethyldithiocarbamate (dtc), 400 mg/kg ip daily, followed by 4 hr of air gassing. Brain TH activity fell 17% in control rats gassed with air on the same schedule as CS₂-exposed rats. The decrease in brain TH activity was observed in both CS₂-exposed and dtc-treated rats after one treatment, but rose above control on the second day of exposure and remained elevated for 6 days.     

Evaluation of pteridine metabolism in battery workers chronically exposed to lead

Occupationally-exposed lead affects the neuromuscular junction and might cause disturbances in the locomotor activity. This study was undertaken to evaluate pteridine metabolism, in which neurotransmitters are synthesized in battery workers. Urinary neopterin, biopterin and creatinine were measured using high performance liquid chromatography. Serum neopterin concentrations were detected by enzyme-linked immunoassay. Blood dihydropteridine reductase (DHPR) activities and deltaaminolevulinic acid (delta-ALA) were measured spectrophotometrically. Blood and urinary lead were detected by atomic absorption spectroscopy. Significantly increased blood and urinary lead levels, urinary neopterin, biopterin and delta-ALA were found in workers, while DHPR activities were indifferent compared to control group. Urinary creatinine decreased. This is the first study to demonstrate that increased activity of the pteridine pathway results in the accumulation of the neurotransmitters that may be responsible for the neurological disorders.     

Serum gonadotropins and testosterone in men occupationally exposed to carbon disulfide

Serum levels of gonadotropins and testosterone were determined for 69 men exposed to CS2 in viscose rayon production and for 22 nonexposed male controls. Mean ages of the two groups were 40 and 39 yr, respectively. The duration of exposure to CS2 ranged from 1 to 36 yr (mean, 12.5). Follicle stimulating hormone (FSH) levels were significantly higher among the exposed men than among the controls (13.6 versus 10.0 IU/l). The level of sex hormone binding globulin (SHBG) was significantly lower among the exposed men than among the controls (44 versus 53 nmol/l). Luteinizing hormone (LH) values were significantly higher among the 24-31 yr old exposed men than the controls of the same age. Levels of SHBG, of the free testosterone index, and of FSH and LH in men under 39 yr who had been exposed to CS2 for 1-9 yr differed significantly from those of their controls, whereas only FSH differed significantly among those exposed for 10 or more years in the same age group. In men aged 40 yr or more, after at least 10 yr of exposure, only FSH and LH were significantly higher than in the controls. Although the observed alterations do not seem to have any overt clinical implications for the study groups, they indicate that overall levels of CS2 that are well below the current Finnish threshold limit value (30 mg/m3) may affect the hormonal balance in the pituitary-gonadal axis, favoring the view that there may be an increased risk of latent primary gonadal insufficiency.     

Biochemical and morphological studies of monkeys chronically exposed to methylmercury

This study was designed to correlated autopsy findings with the effects on cage behavior, laboratory values, and mercury clearance of long-term, low-dose exposure of primates to methylmercury. Six rhesus monkeys were given daily methylmercury hydroxide (MeHg) orally in apple juice on a preplanned dosage schedule. Three were sacrificed while receiving MeHg (group I) and the other 3 were sacrificed 2-5 mo after cessation of MeHg administration (group II). Whole-blood Hg levels (organic and inorganic) were assayed weekly, and major organ levels were assayed at autopsy. Whole-blood Hg levels were maintained between 1 and 2 micrograms/ml when the monkeys were given a MeHg dose of 80-125 micrograms/kg . d for up to 1 yr. The Hg burden of the major organs appeared to be dose- and duration-related. After periods of clearance (2.5-5 mo), intestinal wall Hg burden decreased to less than 1 microgram/g, and the hepatic Hg burden was still between 1.12 and 2.37 micrograms/g. However, the kidneys had a higher concentration of Hg, ranging from 10.34 to 29.54 micrograms/g. Whenever there was a high concentration of Hg, significant ultrastructural changes were observed. In the kidneys there were intracytoplasmic vacuoles and electron-dense inclusion bodies. In the small intestine of the animals cleared of mercury (group II), there were normal Paneth cells, as well as some degenerative cells characterized by dilation of endoplasmic reticulum and the presence of intracellular inclusion bodies. These findings suggest the long turnover time of Hg in these cell populations. During the period of study, weekly routine laboratory data including hematology, blood chemistry, and liver and kidney function tests did not reveal any significant changes.     

Elevation of hepatic metallothionein in rats chronically exposed to dietary ethionine

Following 30 weeks of exposure to 0.3% ethionine in the diet, male Fischer rats bearing liver cell tumors showed a marked increase (9.5 fold) in hepatic metallothionein (MT) concentrations relative to control, as assessed by the cadmium-hemoglobin assay. To confirm the presence of MT in the tumor-containing livers of ethionine-fed rats, biochemical analyses were performed. Spectral analysis of a purified protein from ethionine-treated livers revealed a UV absorption spectrum characteristic of MT. Gel filtration (Sephadex G-75) indicated a prominent copper (Cu) and zinc (Zn)-containing peak with a Mr (approx. 9 000) similar to that of MT. Concurrent with the elevation in hepatic MT in ethionine-fed rats there was an increase in the concentration of Cu and Zn in hepatic cytosol.     

Assessment of tail nerve function in rats chronically exposed to vinyltoluene

In male Sprague-Dawley rats, motor and sensory conduction velocities (MNCV and SNCV) of the tail nerve decreased significantly as a result of oral administration of 400 and 200 mg/kg of 2,5 hexanedione (2,5-HD) once daily, 5 days a week for up to 7 and 15 weeks, respectively; and of whole-body exposure to 300 ppm of vinyltoluene for 6 h daily, 5 days a week for up to 21 weeks. Exposure to 100 ppm of vinyltoluene did not cause any significant impairment of tail nerve function throughout the 21-week exposure period. Significant changes in MNCV and SNCV were consistently observed from weeks 4 and 2, respectively, in 2,5-HD-treated rats. Changes resulting from exposure to 300 ppm of vinyltoluene were reported intermittently from week 15. Significant linear relationships were established between the length of treatment with 2,5-HD, length of exposure to 300 ppm of vinyltoluene, and the extent of impairment of tail nerve function. Structural damage to the sciatic nerves was only seen in 2,5-HD-treated rats. It is concluded that vinyltoluene can be regarded as an airborne chemical which leads to the development of a borderline experimental neuropathy at a level of 300 ppm.     

Dose-related proximal tubular dysfunction in male rats chronically exposed to lead

Male Wistar rats were given 0.5 and 2% lead acetate in drinking water for 2 months, 1% lead acetate for 3 months and sodium acetate equimolar to 2% lead acetate for 3 months. Glucose, total proteins, lactate dehydrogenase (LDH), lysozyme and beta 2-microglobulin (beta 2-m) were measured in 24-h urine every month. Kidney weight and histology were also examined. At the three doses, lead exposure produced a significant elevation of the kidney weight. No significant change in urinary parameters was observed in rats given 0.5% lead acetate. Exposure to 1% lead acetate increased the urinary excretion of beta 2-m only. At the 2% lead acetate dose the elevation of beta 2-m excretion was accompanied by an increased urinary output of glucose, total proteins, lysozyme and LDH. Observations of the kidneys by light microscopy were in agreement with these biochemical findings. The nephrotoxic effect of acetate was excluded by the lack of biochemical or histological effects of sodium acetate on the kidney. It is concluded that a proximal tubular dysfunction is induced in rats chronically exposed to high doses of lead.     

Effect of acetaminophen administration to rats chronically exposed to depleted uranium

The extensive use of depleted uranium (DU) in both civilian and military applications results in the increase of the number of human beings exposed to this compound. We previously found that DU chronic exposure induces the expression of CYP enzymes involved in the metabolism of xenobiotics (drugs). In order to evaluate the consequences of these changes on the metabolism of a drug, rats chronically exposed to DU (40mg/l) were treated by acetaminophen (APAP, 400mg/kg) at the end of the 9-month contamination. Acetaminophen is considered as a safe drug within the therapeutic range but in the case of overdose or in sensitive animals, hepatotoxicity and nephrotoxicity could occur. In the present work, plasma concentration of APAP was higher in the DU group compared to the non-contaminated group. In addition, administration of APAP to the DU-exposed rats increased plasma ALT (p<0.01) and AST (p<0.05) more rapidly than in the control group. Nevertheless, no histological alteration of the liver was observed but renal injury characterized by incomplete proximal tubular cell necrosis was higher for the DU-exposed rats. Moreover, in the kidney, CYP2E1 gene expression, an important CYP responsible for APAP bioactivation and toxicity, is increased (p<0.01) in the DU-exposed group compared to the control group. In the liver, CYP's activities were decreased between control and DU-exposed rats. These results could explain the worse elimination of APAP in the plasma and confirm our hypothesis of a modification of the drug metabolism following a DU chronic contamination.     

Vulnerability of gastric mucosa to prednisolone in rats chronically exposed to cigarette smoke

We examined gastric mucosal vulnerability in a rat model of chronic obstructive pulmonary disease (COPD). Male Wistar rats were exposed to cigarette smoke for 12 weeks (CSE rats), and on the last 4 days of exposure, prednisolone was given to induce gastric mucosal injury. Histopathology, pulmonary function, arterial blood gases, and levels of lipid peroxides (LPO), prostaglandin E(2) (PGE(2)), hypoxia-inducible factor 1 alpha subunit (HIF-1alpha), and vascular endothelial growth factor (VEGF) in gastric mucosa were examined. We also tested the effect of rebamipide on prednisolone-induced gastric lesions. In CSE rats, although no gastric lesions were detected, LPO, PGE(2), HIF-1alpha, and VEGF levels were higher than in control rats. Prednisolone induced gastric hemorrhagic lesions more readily in CSE rats than controls, with concomitant decrease in PaO(2) and increased levels of LPO, HIF-1alpha, and VEGF. Rebamipide reversed gastric lesions without affecting any parameters examined. CSE rats were found to be a useful animal model of COPD, and COPD appeared to render the gastric mucosa vulnerable to prednisolone.