Chiho Watanabe

In utero exposure to methylmercury and Se deficiency converge on the neurobehavioral outcome in mice.

Pregnant female ICR mice, maintained on torula-based diets containing various amounts of Se (0.02, 0.05, or 0.4 mg/kg diet), were given methyl-mercury (MeHg; 0, 5, or 9 mg Hg/kg in total) on the 12-14th days of gestation. The neurobehavioral function of the offspring born to these dams was evaluated with respect to reflex and motor development, thermal preference, and open-field activity. Se deficiency per se as well as exposure to MeHg exerted additive or synergistic effects on the neurobehavioral functions examined. The group of mice most affected was the group given the lowest amount of Se and the highest dose of MeHg. Thus, the neurobehavioral outcome of in utero MeHg exposure and Se deficiency converged. Although the dietary level of Se did not affect the Hg concentration in the fetal brain, the Se concentration and the activity of glutathione peroxidase, a selenoenzyme, were severely depressed by MeHg in the neural tissue. The possibility that functional Se deficiency by MeHg exposure partly accounts for the neurobehavioral toxicity of MeHg is discussed.

Effects of prolonged selenium deficiency on open field behavior and Morris water maze performance in mice.

Neurobehavioral effects of prolonged Se deficiency were evaluated using ICR mice. Dams were fed Se-deficient or Se-adequate diet starting at 4 weeks before conception through the suckling period. After weaning, offspring of both groups were given the same diet as their dams were given. The behavior of these offspring was evaluated with open field apparatus (OPF) and Morris water maze. In OPF, Se-deficient females exhibited less locomotor activity, more defecation, and less entry to the center square areas than did the control females. No such difference was found in males. In the Morris maze, Se-deficient females showed slight but significant impairment during the initial phase of the trials. The behavioral changes in OPF and the maze might have been due to an altered reactivity to a novel environment, although this possibility needs further confirmation. The obtained data showed that the altered behavior was unlikely to be due to the changes in thyroid hormones. Mechanism of these behavioral effects is discussed in relation to possible neurochemical changes induced by Se deficiency.

Milk transfer and tissue uptake of mercury in suckling offspring after exposure of lactating maternal guinea pigs to inorganic or methylmercury

Maternal guinea pigs were injected with mercuric chloride (HgCl2; 1 mg Hg/kg body weight) or methylmercury (MeHg; 1 mg Hg/kg) 12 h after parturition, and exposure of the offspring to mercury (Hg) via breast milk were studied on days 3, 5 and 10 postpartum. Milk Hg concentrations were lower than maternal plasma Hg concentrations regardless of the form of Hg given to the dams. Milk Hg was higher in HgCl2-treated dams than in MeHg-treated dams. In MeHg-treated dams, MeHg was separately determined. While the ratio of MeHg to T-Hg decreased in the dams’ plasma, it did not in the milk. There was a strong correlation between milk and plasma T-Hg concentrations in HgCl2 treated dams. In the milk of MeHg-treated dams, the plasma MeHg concentrations correlated better than did the plasma T-Hg concentrations. In the offspring, regardless of the chemical forms of Hg given to the dams, the highest Hg concentrations were found in the kidney, followed by the liver and the brain. Brain Hg concentrations were, however, significantly higher in the offspring of MeHg-treated dams than in those of HgCl2-treated dams. In addition, Hg levels in the major organs of the offspring of HgCl2-treated dams peaked on day 5 postpartum, while those of MeHg-treated dams did not show a significant decrease up to day 10 postpartum. These facts indicate that the two chemical forms of Hg were transferred to the offspring via the breast milk and were distributed differently, depending on the chemical form, to the offspring’s tissues.

Brain selenium status and behavioral development in selenium-deficient preweanling mice

The influence of Se deficiency on behavioral development in preweanling mice was evaluated. Female ICR mice were fed either Se-deficient or control diet (containing < 20 or 400 ng Se/g diet, respectively) from 4 weeks before conception to the end of the suckling period. In the offspring of Se-deficient dams, liver and brain Se levels were reduced to < 5% and 60% of those in the control offspring, respectively, from as early as the third postnatal day. At weaning, brain Se content exceeded the hepatic one in Se-deficient offspring, whereas in the control offspring the liver contained 10 times more Se than the brain did. Thus, tissue-specific metabolism of Se was already functioning during the neonatal period. When placed in a thermogradient and allowed to move along the gradient, Se-deficient offspring exhibited a preference for a significantly warmer environment than did the controls. They also showed slightly retarded development of walking ability. These results showed that Se-deficient offspring differed from the controls in behavioral development. Possible mechanisms of these alterations are discussed.

Effects of buthionine sulfoximine (BSO) on mercury distribution after Hg° exposure

Effects of buthionine sulfoximine (BSO), a glutathione (GSH) depletor, on mercury (Hg) distribution were studied after mercury vapor (Hg(o)) exposure (3.2 mg/m3) in mice. BSO (1.6 g/kg, i.p.) significantly decreased the GSH levels in the liver, kidney and lung. Pretreatment with BSO increased Hg concentrations in the lung, liver and plasma, but decreased Hg concentration in the kidney. These findings indicate that GSH functions as a determinant in the changes of Hg distribution after Hg(o) exposure. Possible causes of the alterations in Hg distribution are discussed, including the anticipated enhancement in the Hg(o) oxidation process in the tissues with decreased GSH levels as well as the transportation of Hg2+ derived from oxidation of Hg(o). Concentrations of thiobarbituric acid reactive substances (TBARS) were determined as an indicator of oxidative damage, but the increase in TBARS was not detected in any tissue examined.

Growth and behavioral changes in mice prenatally exposed to methylmercury and heat

Postnatal and behavioral changes in mice exposed prenatally to methylmercury and heat were investigated. Pregnant ICR mice were immersed in water at 37 degrees C or 42 degrees C for 10 min once or twice daily from day 12 through day 15 of gestation. Two hours after the heat exposure on day 12 of age, mice were injected s.c. with 5 mg Hg/kg of methylmercury (MeHg, as chloride) or saline. Prenatal exposure to heat significantly induced inactivity in an open field test (OPF) in males and retarded walking ability in both males and females. Prenatal exposure to MeHg caused significant inactivity in the OPF in females. Although heat did not enhance the effect of MeHg on physical growth or the behavior of pups and vice versa, there were some interactions between the effects of these two agents. Thus, the difference in walking ability in both sexes caused by heat was more distinctive in the saline-treated groups than in the MeHg-treated groups; the difference in locomotion in OPF caused by MeHg in females was more distinctive in the normothermic group than among the hyperthermic groups. The mechanisms underlying these behavioral changes need to be further examined.

DETERMINATION OF SELENIUM IN THE HUMAN BRAIN BY GRAPHITE FURNACE ATOMIC ABSORPTION SPECTROMETRY

For the investigation of neurological disorders, a development of simple and accessible methods for determining selenium in human brain samples is required. We devised a method of determining selenium using graphite furnace atomic absorption spectrometry (GFAAS). An electrodeless discharge lamp provided the sufficient sensitivity to determine brain selenium. The matrix interferences were avoided by using high temperature, a prolonged pyrolysis step, and a palladium matrix modifier. The technique of standard addition was used to evaluate the sample concentrations. The accuracy of the method was confirmed by a bovine liver reference material. The detection limit of selenium was 0.04 ng. The determined selenium concentrations of human brain cortex and white matter were higher than those of putamen (115–155 and 206–222 ng/g wet wt, respectively). These GFAAS values agreed with those obtained by fluorometric analysis (r=0.91,n=10). Moreover, the GFAAS values were compatible to those reported by other researchers (99–274 ng/g wet wt), in which selenium concentrations in putamen also tended to be higher than the other two regions. We conclude that GFAAS is useful for selenium analysis in brain samples.

Tissue-specific modification of selenium concentration by acute and chronic dexamethasone administration in mice.

Several clinical reports have shown changes in plasma Se concentration with corticosteroid treatments, but the results have been inconsistent. Few experimental studies have been done on this subject. In the present study the effect of dexamethasone (DEX) treatment on Se concentrations and activities of Se-dependent glutathione peroxidase (EC 1.11.1.9; SeGPx) were examined in adult male ICR mice. In the first experiment, DEX was given via drinking water containing 5 or 50 mg DEX/l. At 1 or 3 weeks of DEX treatment, mice were dissected and the Se concentrations as well as SeGPx activities in various tissues, including plasma, were determined. At 1 week the DEX-treated groups had significantly lower hepatic Se concentrations and significantly higher plasma and cerebral concentrations than the control group. The DEX-treated groups showed lower SeGPx activities in the hepatic cytosol and higher SeGPx activities in the plasma than the saline (9 g NaCl/l)-treated group, in parallel with the changes in Se concentrations. At 3 weeks, neither hepatic nor plasma Se concentrations showed a significant change. In the second experiment, mice were injected subcutaneously with DEX and, thereafter, mice were food-deprived. The DEX-injected groups had higher plasma Se concentrations. A similar finding was obtained also when the DEX- or saline-injected mice were not food-deprived. Thus, the difference between the DEX-treated and control groups was possibly caused by redistribution of tissue Se. These results suggested that the effects of DEX on Se concentrations were tissue dependent and that the higher plasma Se observed in DEX-treated groups might be explained by the release of tissue Se into plasma as plasma SeGPx.

Inhibition of γ-glutamyltranspeptidase decreases renal deposition of mercury after mercury vapor exposure

The alteration of renal deposition of mercury (Hg) after mercury vapor (Hgo) exposure was studied in mice pretreated with acivicin, a potent and irreversible inhibitor of γ-glutamyltranspeptidase (GGT). Pretreatment with acivicin decreased renal Hg concentration by about 60% and significantly increased Hg concentration in the urine compared with the non-treated group. The results suggest that renal deposition of Hg after Hgo exposure depends on renal GGT, which plays an important role in the uptake of GSH or GSH conjugates filtered through the glomeruli. It seems that the mechanism of renal Hg deposition after Hgo exposure is similar to that after exposure to ionic Hg: a GGT-mediated incorporation of an Hg-GSH complex into renal tubular cells. The acivicin pretreatment after Hgo exposure did not affect Hg concentrations in the liver and erythrocytes.

Deficiency of selenium enhances the K+-induced release of dopamine in the striatum of mice.

To determine whether a selenium (Se) deficiency in the brain leads to a functional change in dopaminergic transmission in the striatum, in vivo microdialysis was conducted in mice fed a low-Se diet. After 11-13 weeks of the diet regimen, the activity of glutathione peroxidase (GPx) in the Se-deficient brain was reduced to 60% of the control brain. A high K+ perfusion (100 mM) increased the level of dopamine in the dialysate to 67 +/- 16 times the basal level; the increase was significantly greater than that observed in the control group (28 +/- 4 times). Such a between-group difference was not observed after 4-5 weeks of the Se-diet. These results indicated that prolonged Se deficiency altered the function of striatal dopaminergic neurons in mice. A possible contribution of enhanced oxidative stress due to the reduced GPx activity is discussed.