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M Aschner

Publications and source records attributed to M Aschner.

At least 91 records · Page 5Linked to original sources

Manganese neurotoxicity: cellular effects and blood-brain barrier transport.

The observations by Couper in 1837 are acknowledged as the earliest description of the toxic syndrome associated with chronic manganese (Mn) exposure. Since that time, many of the neurotoxic aspects of manganism have been described, yet, the primary basis for its neurotoxicity remains unknown. Recent evidence corroborates the original hypothesis by Maynard and Cotzias (82) which invokes the mitochondrion as the target organelle for Mn cytotoxicity which is primarily expressed as a perturbation in Ca2+ homeostasis. Despite recognition that excessive Mn exposure culminates in Mn accumulation in the CNS and a clinical picture dominated by neurological disturbances, the role of the blood-brain barrier in the CNS uptake of Mn has received little attention. Accordingly, the first part of this review summarizes the current understanding of the interaction of Mn with biologically active sites in the induction of Mn cytotoxicity. The second part of this review summarizes what is known about Mn transport across the blood-brain barrier, a major regulator of the CNS milieu, with the contention that the rate and extent of Mn transport across the blood-brain barrier modulates its neurotoxicity.

Animals↗

The use of astrocytes in culture as model systems for evaluating neurotoxic-induced-injury.

The prevailing thought that astrocytes function predominantly as passive metabolic or even physical support for neurons has faded over the last 20 years. Today these stellar shaped cells are credited with an expanded role, playing key functions in CNS development, homeostasis, and pathology. In probing their expanded roles, primary astrocyte culture systems have proven to be an indispensable tool. Astrocytes have been implicated in both a defensive and facilitatory capacity for many toxic injuries. Evidence for a protective role of astrocytes in modulating CNS toxicity is afforded by observations that the toxicity of glutamate to cortical neurons is diminished upon astrocytic enrichment of the cell culture (Rosenberg and Aizenman, 1989). In cultures of rat cerebral cortex in which astrocyte proliferation is stringently suppressed, glutamate neurotoxicity occurs at low glutamate concentrations similar to those which are normally found in the extracellular space in the hippocampus. In the presence of excess astrocytes, concentrations of glutamate one-hundred fold higher are required to produce equivalent neurotoxicity (Rosenberg and Aizenman, 1989). Astrocytes can facilitate the action of neurotoxins via a modulating process which takes place within the astrocyte or by a direct cytotoxic effect. Whereas primary astrocyte cultures remain unaffected by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP; Marini et al., 1989), they function prominently in the selective destruction of dopaminergic neurons of the nigrostriatal pathway in humans, other primates and rodents (Davis et al. 1979; Langston et al., 1983; Burns et al., 1983; Langston et al., 1984; Heikkila et al., 1984; Jarvis and Wagner, 1985). Thus, while MPTP by itself is not toxic to cerebellar cells in co-culture with cerebellar astrocytes, MPTP is toxic to the granule cells (Marini et al, 1989). This is thought to be due to an astrocyte-mediated conversion of MPTP to its highly polar and toxic metabolite, 1-methyl-4-phenylpyridinium ion (MPP+; Chiba et al. 1984). There is compelling evidence that astrocytes respond directly or indirectly to a number of other neurotoxins. Direct cytotoxic effects on astrocytes constitute the major morphologic feature in hyperammonemia (Norenberg, 1981), a condition implicated as an etiologic factor in several CNS disorders. In addition, a predisposition of astrocytes for methylmercury uptake (Aschner et al., 1990 a,b) offers a possible explanation for the observed neurotoxicity of this heavy metal, since a direct toxic effect on astrocytes would result in failure of astrocyte homeostatic functions, indirectly resulting in neuronal impairment, injury and death.

Ammonia↗

Interactions of methylmercury with rat primary astrocyte cultures: inhibition of rubidium and glutamate uptake and induction of swelling.

The ability of astrocytes to sequester MeHg may indicate an astrocyte-mediated role in MeHg's neurotoxicity. Hence, studies were undertaken to assess the effects of MeHg on metabolic functions in cultured astrocytes. MeHg (10(-5) M) significantly inhibited the initial rate (5 min) of uptake of 86RbCl, used as a tracer for K+. 86RbCl uptake was also sensitive to the omission of medium Na+. MeHg (10(-5) M) also markedly inhibited the initial rate of uptake (1 min) of the Na(+)-dependent uptake of [3H]L-glutamate. A second neurotoxin, MnCl2 (0-5 x 10(-4) M), did not alter [3H]glutamate or 86RbCl uptake. MeHg, but not MnCl2, also stimulated the release of intracellular 86Rb+ in a dose-dependent fashion. This effect could be prevented by the administration of MeHg as the glutathione conjugate. These observations support the hypothesis that the astrocyte plasma membrane is an important target for MeHg's toxic effect and specifically that small concentrations of this organometal inhibit the ability of astrocytes to maintain a transmembrane K+ gradient. This would be expected to compromise the ability of astrocytes to control extracellular K+ either by spatial buffering or active uptake, resulting in cellular swelling. We therefore studied volume changes in astrocytes using uptake of [14C]3-O-methyl-D-glucose, in attached cells in response to exposure to MeHg. Exposure to MeHg (0-5 x 10(-4) M) caused a marked increase in the cell volume that was proportional to concentrations of MeHg.

3-O-Methylglucose↗

Methylmercury uptake in rat primary astrocyte cultures: the role of the neutral amino acid transport system.

The significance of the dense labeling pattern of methylmercury (MeHg) over astrocytes in areas of damaged cortex remains obscure, and the extent to which individual neurons are altered by MeHg accumulation in astrocytes is unknown. As a first step in understanding the relationship between the astrocyte and the mechanisms of MeHg's neurotoxicity, studies were directed at how MeHg is transported into cultured astrocytes. Uptake of [203Hg]MeHg in primary astrocyte cultures from neonatal rat cerebral cortex following incubations with MeHgCl conformed to a simple diffusion process. Uptake of [203Hg]MeHg by astrocytes exhibited the kinetic criteria of a specific transport system when added to the media as the L-cysteine conjugate. Saturation kinetics, substrate specificity and inhibition, and trans-stimulation were demonstrated in the presence of this SH-containing amino acid. Cysteine-mediated uptake of MeHg was inhibited by the coadministration of L-methionine, and 2-aminobicyclo-[2,2,1]-heptane-2-carboxylic acid. 2-Methylaminoisobutyric acid was ineffective in inhibiting the uptake of the MeHg-cysteine conjugate. Preloading of the astrocytes with glutamate was moderately effective in trans-stimulating the uptake of MeHg-cysteine conjugates, while in the absence of cysteine, uptake of [203Hg]MeHg was unchanged. These results indicate the presence in astrocytes of a neutral amino acid carrier transport System L, capable of selectively mediating cysteine-MeHg uptake. The substrate specificity and high affinity of this transport system resemble the properties of the System L neutral amino acid transport across the blood-brain barrier in the rat. Cellular uptake of MeHg-cysteine conjugates was not inhibited by preincubation of astrocytes with 100 microM N-ethylmaleimide or NaF.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acids↗

Manganese transport across the blood-brain barrier: relationship to iron homeostasis.

The binding characteristics of manganese (Mn) to transferrin (Tf) were examined on G-75 Sephadex gel columns. When 54MnCl2 was combined with Tf and immediately fractionated on the Sephadex column, 49% of 54Mn was found to Tf. The fraction of 54Mn which was Tf-bound was dependent upon the incubation period, and increased in a time-dependent fashion. In vivo, 6 hr of intravenous administration of ferric-hydroxide dextran complex significantly inhibited (p less than 0.05) 54Mn brain uptake as compared to its uptake in iron-free dextran-treated rats. These results suggest that iron (Fe) homeostasis may play an important role in the regulation of Mn transport across the blood-brain barrier (BBB).

Animals↗

Mercury neurotoxicity: mechanisms of blood-brain barrier transport.

Mercury exists in a wide variety of physical and chemical states, each of which has unique characteristics of target organ toxicity. The classic symptoms associated with exposure to elemental mercury vapor (Hg0) and methylmercury (CH3Hg+; MeHg) involve the central nervous system (CNS), while the kidney is the target organ for the mono- and divalent salts of mercury (Hg+ and Hg++, respectively). Physical properties and redox potentials determine the qualitative and quantitative differences in toxicity among inorganic mercury compounds, while the ability of MeHg to cross the blood-brain barrier accounts for its accumulation in the CNS and a clinical picture that is dominated by neurological disturbances. This review gives an up-to-date account of mercury's physical and chemical properties and its interaction with biologically active sites pertinent to transport across the blood-brain barrier, a major regulator of the CNS millieu.

Animals↗

Enzymatic activity is necessary for thrombin-mediated increase in endothelial permeability.

alpha-Thrombin causes a dose-dependent increase in endothelial permeability as measured by the clearance rate of 125I-albumin across a monolayer of bovine pulmonary artery endothelial cells. We determined if an active catalytic site is necessary for the thrombin-mediated increase in endothelial permeability. alpha-Thrombin was reacted with 10-fold excess D-phenylalanyl-prolyl-arginine chloromethyl ketone (PPACK), an irreversible inhibitor that forms a covalent bond with thrombin's active site, producing an enzymatically inactive thrombin. PPACK completely inhibited the alpha-thrombin-mediated increase in 125I-albumin permeability. Similar results were obtained with gamma-thrombin, an enzymatically active alpha-thrombin form with an altered fibrinogen recognition domain. PPACK alone and the active site-inhibited PPACK-alpha-thrombin had no effect on permeability. Diisopropylphospho (DIP)-alpha-thrombin was effective only in very high concentrations (10(-6)M), and this effect was abolished by the addition of PPACK. These studies demonstrate that binding alone is insufficient for the thrombin-mediated increase in endothelial monolayer permeability. Thrombin's active catalytic site is a requirement for the increase in transendothelial albumin permeability.

Amino Acid Chloromethyl Ketones↗

Methyl mercury uptake across bovine brain capillary endothelial cells in vitro: the role of amino acids.

Previous studies in the rat in vivo have demonstrated that co-injection of methyl mercury (MeHg) with L-cysteine into the common carotid artery enhances brain Hg levels following a single capillary pass through the CNS vasculature. In order to elucidate the relationship between MeHg transport and the neutral amino acid transport carrier system, regulatory aspects of MeHg transport across the bovine blood-brain barrier were investigated in isolated brain microvessel preparations. Following 1 hour co-incubations of 203Hg-MeHgCl with 0.1 mM L-cysteine at 37 degrees, 203Hg uptake by suspended microvessels was significantly increased (P less than 0.05) compared with controls. This enhanced capillary uptake of 203Hg was abolished by co-incubations of microvessels with 0.1 mM L-cysteine-L-methionine, or 0.1 mM L-cysteine plus AT-125 (alpha S, 5S)-alpha-amino-3-chloro-4,5-dihydro-5-isoxazolacetic acid), an irreversible inhibitor of gamma-glutamyl-transpeptidase. One hr co-incubations of bovine capillaries with 203Hg-MeHgCl and 0.1 mM D-cysteine at 37 degrees or 0.1 mM L-cysteine at 0 degrees did not increase rat of 203Hg uptake compare with controls. These results indicate that L-cysteine enhances the rate of capillary MeHg uptake. The accumulation of 203Hg in the bovine microvessels appears to be a carrier-mediated process. It is inhibited by L-methionine, a competitive substrate for neutral amino acid transport, and by AT-125. Capillary uptake of 203Hg is stereospecific to the L-enantiomorph of cysteine, suggesting selective uptake of MeHg across the blold-brain barrier. The data emphasize the relationship between the L-enantiomorph neutral amino acid carrier system and MeHg transport across the capillaries.

Amino Acids↗

Brain, kidney and liver 203Hg-methyl mercury uptake in the rat: relationship to the neutral amino acid carrier.

To investigate the effect of L-neutral amino acids on tissue levels of methyl mercury in the adult animal, rats were infused into the external jugular vein with solutions containing a) 0.05 mM 203Hg-MeHgCl and saline, b) 0.05 mM 203Hg-MgHgCl-0.1 mM L-cysteine, c) 0.05 mM 203Hg-MeHgCl-0.1 mM L-cysteine-0.1 mM L-cysteine-0.1 mM L-methionine, d) 0.05 mM 203Hg-MeHgCl-0.1 mM L-leucine, or e) 0.05 mM 203Hg-MeHgCl-0.1 mM L-cysteine-0.1 mM L-leucine, Groups of animals were sacrificed at 3 min. 7 hr, and 96 hr. Brain, kidney, and liver 203Hg radioactivity was measured by means of gamma-scintillation spectrometry. Brain 203Hg concentrations L-cysteine treated animals were significantly higher compared with saline treated animals (P less than 0.05) at 3 min., 7 hr and 96 hr. The coinjection or coinfusion of methyl mercury with L-cysteine and L-methionine abolished the L-cysteine-mediated brain 203Hg uptake (P less than 0.05), at each sacrifice time. Kidney and liver 203Hg concentrations were not significantly different in any of the treatment groups compared with controls, irrespective of the sacrifice time. Furthermore, the percentage of diffusible 203Hg (non-protein bound) at each sacrifice time was not statistically different irrespective of the treatment assigned. These results suggest that methyl mercury L-cysteine conjugates in the plasma may share a common transport step with the L-neutral amino acid carrier transport system and indicate the presence in brain capillaries of a transport system capable of selectively mediating methyl mercury uptake across the capillary endothelial cell membrane.

Amino Acids↗

Uptake of methylmercury in the rat brain: effects of amino acids.

Aspects of the regulation of methylmercury (MeHg) transport across the blood-brain barrier (BBB) were investigated in the in vivo Long-Evans female rat. Fifteen s after intracarotid injection, brain 203Hg concentration was significantly increased in animals injected with 0.05 mM [203Hg]MeHgCl plus 0.1 mM L-cysteine compared with controls (P less than 0.05). This L-cysteine-enhanced 203Hg brain uptake was abolished by coinjections of [203Hg]MeHgCl with 0.1 mM L-cysteine-L-methionine, or 0.1 mM L-cysteine plus AT-125 (alpha S, 5S-alpha-amino-3-chloro-4,5-dihydro-5-isoxazolacetic acid), an irreversible inhibitor of gamma-glutamyl transpeptidase. Coinjections of rats with [203Hg]MeHgCl and 0.1 mM D-cysteine did not result in an increase in the rate of 203Hg uptake compared with controls. Furthermore, [203Hg]MeHg uptake at 15 s after intracarotid injections in the rat was stereospecific to the neutral amino acid carrier, as 203Hg uptake across the BBB was not inhibited by coinjections of [203Hg]MeHgCl with aspartic acid, an acidic amino acid. These results indicate the presence in brain capillaries of a transport system capable of selectively mediating MeHg uptake across the brain capillary endothelial cell membrane.

Amino Acids↗

Distribution of mercury 203 in pregnant rats and their fetuses following systemic infusions with thiol-containing amino acids and glutathione during late gestation.

To investigate the effect of amino acids and the tripeptide glutathione (GSH) on tissue uptake of methylmercury (MeHg) in the developing rat fetus in utero, pregnant rats were continuously infused into the external jugular vein with 0.1 mM L-cysteine, 0.1 mM L-leucine, 0.1 mM GSH or saline commencing on day 17 of gestation. This was followed at 24, 48, and 72 hours by external jugular infusion of 50 microM [203Hg]-MeHgCl administered in 1 ml over 1 hour. Pups were surgically removed from the uterus on gestational day 21. Whole body, brain, kidney, liver, and placental 203Hg radioactivity was measured by means of gamma-spectrometry. Brain 203Hg concentration in pups exposed in utero to L-cysteine was significantly higher compared with pups exposed to saline (P less than 0.05). Brain 203Hg concentration in pups exposed in utero to L-leucine and GSH was significantly depressed compared with pups exposed to saline (P less than 0.05). Kidney 203Hg concentration was not significantly changed in all treatment groups compared with controls. Liver 203Hg concentration was significantly depressed in L-leucine- and GSH-treated pups compared with controls (P less than 0.05). Placental 203Hg concentration was not affected by any treatment compared with controls. These effects occurred despite no difference in total 203Hg body burden among pups, irrespective of the treatment. In addition, infusion with L-cysteine resulted in a significant increase in 203Hg brain concentration in dams compared with controls, and 203Hg brain concentration in L-leucine- and GSH-treated dams was significantly depressed compared with controls. Thus 203Hg distribution in both adult and developing animals is altered by chronic amino acid or GSH infusions and suggests that MeHg uptake may be mediated through the formation of a cysteine-MeHg complex which is transported across the blood-brain barrier by the neutral amino acid carrier transport system.

Animals↗

Increased axonal transport in the rat optic system after systemic exposure to methylmercury: differential effects in local vs systemic exposure conditions.

Axonal transport was studied by several techniques in the optic system of adult female Long-Evans rats following systemic exposure to methylmercury in 5 mM Na2CO3. Control rats were treated with the buffer alone. Four mg Hg/kg body weight for 4-6 days, or for 12 days, induced significant changes in the rate of protein synthesis in the retinal cells and in the rate of propagation of protein-bound radioactivity along the ganglion cell axons. Axonal transport of particulate material in both groups treated with methylmercury increased to a rate of 147 mm/day compared to 93 mm/day in controls. Methylmercury was distributed evenly throughout the retinogeniculate system. No clinical neuropathy was evident in either mercury-treated group. It is proposed that the increased rates of transport may represent an adaptive compensatory response to distal axonopathy caused by methylmercury. To investigate why systemic dosing produced effects opposite to those observed with local application of MeHg, various doses of MeHg were tested in the local and systemic paradigms, including doses which yielded equal concentrations of Hg in the retina. The results indicate that the differential response between the two treatment conditions is not a function of local dose, per se. Local and systemic application produce different dose-effect curves, which do not coincide at any dose.

Animals↗

Mercury 203 distribution in pregnant and nonpregnant rats following systemic infusions with thiol-containing amino acids.

Near-term pregnant (gestational day 17) and nonpregnant Long-Evans female rats were continuously infused into the external jugular vein with 0.1 mmole/hour L-cysteine, 0.1 mmole/hour L-leucine, or saline. At 24, 48, and 72 hours, 50 mumole/hour [203Hg]-MeHgCl was administered over 1 hour. Total 203Hg body burden, brain, kidney, liver, and blood 203Hg concentrations were determined at 96 hours by gamma scintillation spectrometry. Despite significantly greater 203Hg whole body retention in the pregnant animals 203Hg concentrations in blood, brain, kidney, and liver were higher in nonpregnant rats. In addition, brain 203Hg concentrations in both pregnant and virgin rats were significantly higher in L-cysteine-treated rats compared with controls. These results suggest that the fetus may act as a "sink" for MeHg, thus decreasing 203Hg concentrations in maternal blood, brain, kidney, and liver. Furthermore, the data indicate that brain uptake of methylmercury in both pregnant and nonpregnant rats is enhanced by chronic L-cysteine infusion, lending support to the hypothesis that methylmercury in the rat may be translocated across the blood-brain barrier by the neutral amino acid carrier transport system.

Animals↗

Changes in axonally transported proteins in the mature and developing rat nervous system during early stages of methyl mercury exposure.

It was established by means of SDS polyacrylamide gel electrophoresis that direct injections of methyl mercury (10 micrograms Hg) into the mature rat vitreous body of the eye decrease protein synthesis in the retina and optic nerve at 4 hours after injection. Although the global spectrum of polypeptides did not change, a specific decrease in the volume of polypeptides of 20-23 K daltons molecular weight was evident. Conversely, systemic exposure to methyl mercury resulted in increased protein synthesis of polypeptides of 20-23 K molecular weight both in adult (8 mgHg/kg/day for 8 days) and neonatal rats (2 mgHg/kg/day for 10 days). In addition, specific changes in the volume of polypeptides 75-90 K molecular weight were noted in sciatic nerves of neonatal rats. These data are consistent with a bimodal response in protein synthesis following MeHg treatment. Local presence of MeHg following direct injection into the eye causes a reduction in protein synthesis, while chronic systemic exposure results in increased synthesis and transport of proteins in both mature and developing optic nerves and neonatal sciatic nerves. Thus, these systems possess the capacity to attempt regenerative processes through induction of a small subset of proteins known as GAPs (Growth-Associated Proteins) during the early stages of systemic methyl mercury exposure. These wide spread and system-specific changes are consistent with growth-specific functions during the early stages of methyl mercury exposure.

Aging↗

Reduction of axonal transport in the rat optic system after direct application of methylmercury.

Fast axonal transport of proteins in the optic nerve and tract was quantified by scintillation counts of protein-bound radioactivity along the visual pathway after an intraocular injection of [3H]proline. In control rats the label traveled at a rate of about 60 mm/day, reaching the optic chiasm at 4 h and the lateral geniculate body at 8 h postinjection. When methylmercury was injected simultaneously with [3H]proline, the label traveled at a rate of about 30 mm/day. At 8 h postinjection, the labeled protein had reached the optic chiasm, but the more distal pathway was unlabeled. The same pattern was observed histologically by emulsion autoradiography of the pathway. Some label was detected in the lateral geniculate of methylmercury-treated animals at 8 h, but this may have resulted from local incorporation, as judged by a similar level of labeling in the contralateral visual pathway. Alternatively, it may be the case that a small fraction of the axons in the treated pathway continued to transport proteins in a normal fashion. The very heavy label observed throughout the pathway in controls was present only in the proximal half of the pathway in methylmercury-treated rats. Methylmercury significantly reduced incorporation of [3H]proline in the rat retina, but this reduction was not as great as the effect in the optic nerve. In contrast, cycloheximide, a potent protein synthesis inhibitor, reduced labeled protein in the optic nerve only to the same extent as it reduced incorporation. These results suggest that methylmercury's effect on transport is not dependent solely on its effects on protein synthesis, but represents a separate mechanism of neurotoxicity.

Animals↗

Cell proliferation in developing brain after brief exposure to nitrous oxide or halothane.

Several inhalant anesthetics, including nitrous oxide and halothane, are known to be antimitotic in a variety of developing tissues, but none has been tested for antimitotic activity in developing brain. Concern about the safety of these agents has centered around behavioral effects reported in humans and animals after early exposure. Because interference with cell production during CNS development is a sufficient cause for later behavioral abnormalities, it is important to know whether cell production in the nervous system is altered by these agents. Mice were exposed to either nitrous oxide (75% N2O and 25% O2) or halothane (0.5% halothane in 75% N2 and 25% O2) or a mixture of 75% N2 and 25% O2. Prenatal treatment groups were exposed for 6 h on the 14th day of gestation, while postnatal treatment groups were exposed for 4 h on the second day after birth. Treated and control animals were then killed immediately after exposure, or 12, 24, or 48 h later, to be evaluated for CNS mitotic activity. Each of the four anesthetic-exposed groups showed some deviations from normal mitosis, but only the postnatal nitrous oxide group showed the pattern of reduced cell proliferation followed by a rebound that is characteristic of many antimitotic teratogens. Although prenatal nitrous oxides' effects on the fetal brain were not clearly interpretable, it did delay development of blood, as has been reported by other investigators. Both nitrous oxide and halothane significantly reduced body weight of fetuses in utero, but did not reduce body weight of neonates. The pattern of the body-weight effects suggests that they occur by some mechanism other than reduced cell production.(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of Variance↗

Changes in axonally transported proteins in the rat visual system following systemic methyl mercury exposure.

In an effort to understand the effect of methylmercury on protein synthesis and axonal transport, we have analysed the composition and rate of axonally transported proteins in the retinal ganglion cells of the mature rat. By means of scintillation spectrometry and autoradiography, it was established that systemic exposure to 4 mg Hg/kg/day for four to six days, or twelve days, resulted in an increased rate and volume of transported protein-bound radioactivity in the visual system of the mature rat. In an effort to characterize these changes, the composition of transported polypeptides was analysed by means of SDS polyacrylamide gel electrophoresis. Selective changes in the composition of transported polypeptides were evident. These changes of a small subset of proteins known as GAPs (growth-associated proteins) are consistent with the suggestion that they may have been involved in growth-specific functions during the early stages of methyl mercury exposure. We concluded that, during this period, retinal ganglion cells may express growth-related genes and engage in regenerative processes.

Animals↗