Disruption of the potential across the synaptosomal plasma membrane and mitochondria by neurotoxic agents (original) (raw)
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Toxicology and Applied Pharmacology, 1990
The effects of the neurotoxic metals methylmercury (MeHg) and trimethyltin (TMT) on oxygen reactive species formation within a crude synaptosomal fraction (P2), using the probe 2'. 7'-dichlorofluorescin diacetate (OCFH-DA), and intracellular calcium ([Ca 2 +];), with the fluorescent indicator fluo-3, have been investigated. Two and seven days after a single injection ofMeHg (1 mg/kg) the formation rate of cerebellar oxygen reactive species was significantly increased. Hippocampal and frontocortical oxygen reactive species were elevated 2 days after TMT injection (3 mg/kg). In vitro exposure to MeHg (10-20 µM) increased the formation rate of oxygen reactive species, while TMT (5-40 µM) was without effect. Levels of[Ca 2 +]i were unaltered in P2 fractions from cerebellum and hippocampus of animals treated with either organometal. The data demonstrate that oxygen reactive species are elevated in brain regions, cerebellum (MeHg) and hippocampus (TMT), believed to be selectively vulnerable to these toxic agents. Findings suggest that oxidative damage may be a mechanism underlying the toxicity of both organometals. The use ofOCFH-DA may have potential in the nervous system as an indicator of neurotoxic damage.
Biochemical Pharmacology, 2001
This study investigated the effects of C7 and C9 aliphatic (n-heptane, n-nonane), naphthenic (methylcyclohexane, 1,2,4-trimethylcyclohexane (TMCH)) and aromatic (toluene, 1,2,4-trimethylbenzene (TMB)) hydrocarbons on the production of reactive oxygen species (ROS) and reactive nitrogen species (RNS) in rat brain synaptosome fraction. Methyl mercury (MeHg) was included as a positive control. Exposure of the synaptosomes to the hydrocarbons produced a concentration-dependent linear increase in the formation of the fluorescence of 2Ј,7Ј-dichlorofluorescein (DCF) as a measure of the production of ROS and RNS. Formation of RNS was demonstrated by preincubation of the synaptosome fraction with the neuronal nitric oxide synthase (nNOS) inhibitor N-nitro-L-arginine methyl ester (L-NAME), which reduced the MeHg and TMCH-stimulated fluorescence by 51% and 65%, respectively. The naphthenic hydrocarbon TMCH showed the strongest potential for ROS and RNS formation in rat brain synaptosomes, followed by TMB, toluene, n-nonane, n-heptane, and methylcyclohexane, respectively. TMCH was selected for mechanistic studies of the formation of ROS. Both MeHg and TMCH induced an increase in intracellular calcium concentration [Ca 2ϩ ] i as measured with Fura-2. Blockade of voltage-dependent Ca 2ϩ channels with lanthanum prior to stimulation with MeHg and TMCH led to a reduction in the ROS/RNS formation of 72% and 70%, respectively. Furthermore, addition of cyclosporin A (CSA), a blocker of the mitochondrial permeability transition pore (MTP), lowered both the MeHg and TMCH-elevated DCF fluorescence by 72% and 59%. Preincubation of the synaptosome fraction with the protein tyrosine kinase inhibitor genistein lowered the MeHg and TMCH-stimulated fluorescence by 85% and 91%, respectively. Addition of the extracellular signal-regulated protein kinase (MEK)-1 and-2 inhibitor U0126 reduced the fluorescence stimulated by MeHg and TMCH by 62% and 63%. Furthermore, the protein kinase C inhibitor bisindolylmaleimide reduced the fluorescence stimulated by MeHg and TMCH by 52% and 56%. The compound 1-(6-[17beta-3-methoxyestra-1,3,5(10)-trien-17-yl]-aminohexyl)-1H-pyrrole-2,5-dione (U73122), which inhibits phospholipase C, was shown to decrease the ROS and RNS formation induced by MeHg and TMCH by 49% and 64%, respectively. The phospholipase A 2 (PLA 2) inhibitor 7,7-dimethyl eicosadienoic acid (DEDA) reduced fluorescence in response to MeHg and TMCH by 49% and 54%. Simultaneous addition of L-NAME, CSA, and DEDA to the synaptosome fraction totally abolished the DCF fluorescence. In conclusion, C7 and C9 aliphatic, naphthenic, and aromatic hydrocarbons stimulated formation of ROS and RNS in rat brain synaptosomes. The naphthenic hydrocarbon TMCH stimulated formation of ROS and RNS in the synaptosomes through Ca 2ϩ-dependent activation of PLA 2 and nNOS, and through increased transition permeability of the MTP. Exposure of humans to the naphthenic hydrocarbon TMCH may stimulate formation of free radicals in the brain, which may be a key factor leading to neurotoxicity.
J Neurochem, 2003
Six chemicals, 2-halopropionic acids, thiophene, methylhalides, methylmercury, methylazoxymethanol (MAM) and trichlorfon (Fig. 1), that cause selective necrosis to the cerebellum, in particular to cerebellar granule cells, have been reviewed. The basis for the selective toxicity to these neurones is not fully understood, but mechanisms known to contribute to the neuronal cell death are discussed. All six compounds decrease cerebral glutathione (GSH), due to conjugation with the xenobiotic, thereby reducing cellular antioxidant status and making the cells more vulnerable to reactive oxygen species. 2-Halopropionic acids and methylmercury appear to also act via an excitotoxic mechanism leading to elevated intracellular Ca 2+ , increased reactive oxygen species and ultimately impaired mitochondrial function. In contrast, the methylhalides, trichlorfon and MAM all methylate DNA and inhibit O 6-guanine-DNA methyltransferase (OGMT), an important DNA repair enzyme. We propose that a combination of reduced antioxidant status plus excitotoxicity or DNA damage is required to cause cerebellar neuronal cell death with these chemicals. The small size of cerebellar granule cells, the unique subunit composition of their N-methyl-D-aspartate (NMDA) receptors, their low DNA repair ability, low levels of calcium-binding proteins and vulnerability during postnatal brain development and distribution of glutathione and its conjugating and metabolizing enzymes are all important factors in determining the sensitivity of cerebellar granule cells to toxic compounds.
Biochemical and Biophysical Research Communications, 1986
l-methyl-4-phenylpyridine (MPP+), a major product of the oxidation of the neurotoxic amine l-methyl-4-phenyl-I ,2,3,6-tetrahydropyridine (MPTP) has been postulated to be the compound responsible for destruction of nigrostriatal neurons in man and primates and for inhibition of mitochondri~l NADH oxidation which leads to cell death..We have confirmed that 0.5 mM MPP-inhibits extensively the oxidation of NADt-linked substrates in intact liver mitochondria in State 3 and after uncoupling, while succinate oxidation is unaffected. However, in inverted mitochondria, inner membrane preparations, and Comple~ I NADH oxidation is not significantly affected at this concentration of MPP-, nor are malate and glutamate dehydrogenases or the carriers of t~ese substrates inhibited. We report here the discovery of an uptake system for MPP in mitochondria which is greatly potentiated by the presence of malate plus glutamate and inhibited by respiratory inhibj~tors, suggesting an energy-dependent carrier. A 40-fold concentration of MPP-in the mitochondria occurs in ten minutes. This might account for the inhibition of malate and glutamate oxidation in intact mitochondria.
Toxicology in Vitro, 2008
Neurotoxicology considers that chemicals perturb neurological functions by interfering with the structure or function of neural pathways, circuits and systems. Using in vitro methods for neurotoxicity studies should include evaluation of specific targets for the functionalism of the nervous system and general cellular targets. In this review we present the neuronal characteristics of primary cultures of cortical neurons and of cerebellar granule cells and their use in neurotoxicity studies. Primary cultures of cortical neurons are constituted by around 40% of GABAergic neurons, whereas primary cultures of cerebellar granule cells are mainly constituted by glutamatergic neurons. Both cultures express functional GABAA and ionotropic glutamate receptors. We present neurotoxicity studies performed in these cell cultures, where specific neural targets related to GABA and glutamate neurotransmission are evaluated. The effects of convulsant polychlorocycloalkane pesticides on the GABA A , glycine and NMDA receptors points to the GABA A receptor as the neural target that accounts for their in vivo acute toxicity, whereas NMDA disturbance might be relevant for long-term toxicity. Several compounds from a list of reference compounds, whose severe human poisoning result in convulsions, inhibited the GABA A receptor. We also present cell proteomic studies showing that the neurotoxic contaminant methylmercury affect mitochondrial proteins. We conclude that the in vitro assays that have been developed can be useful for their inclusion in an in vitro test battery to predict human toxicity.
Methyl Mercury Injury to CNS: Mitochondria at the Core of the Matter
Methyl Mercury Injury to CNS: Mitochondria at the Core of the Matter, 2015
Methyl-mercury (MeHg) is one of the most hazardous environmental pollutants of great concern to public health and regulatory agencies because of its primary toxicity to the human central nervous system. The major source of MeHg exposure to the general population is through consumption of contaminated fish and other food products. MeHg, absorbed from the gastrointestinal tract, is easily transported across the blood-brain barrier (BBB). Cysteine-facilitated transport of MeHg into the brain has been demonstrated, and in particular a neutral amino acid transport system capable of mediating MeHg-cysteine uptake has been identified in astrocytes where MeHg accumulation induces cell swelling and inhibition of glutamate uptake. Elevation of glutamate levels in the extracellular space may, in turn, trigger or accelerate processes of excitotoxic neuro degeneration. The rising of extracellular glutamate levels is responsible for the sustained activation of glutamate receptors, hence enhancing Na + influx and Ca 2+ release from intracellular organelles that may trigger a biochemical cascade which promotes the reactive oxygen species (ROS) production. In this scenario, mitochondria may play a crucial role, as these organelles act as a buffer against cytosolic calcium and mediate ROS formation in cells. Herein, we summarize studies providing insights into the molecular and cellular mechanisms involved in MeHg-induced neuro degeneration with particular focus on the role of astrocytes and mitochondria. Indeed, mitochondria may be supposed to lie at the crossroads of a network of events (microtubule disorganization, Ca 2+ dyshomeostasis, ROS generation) leading to neuro degeneration, although it is difficult to establish the upstream mechanisms and downstream effectors in this cascade of events.
Dichlorvos is an organophosphate insecticide effectively used against mushroom flies, aphids, spiders, mites, caterpillars and thrips. Toxicity of dichlorvos is confirmed by air, water and food via inhalation, dermal, absorption and ingestion. Earlier studies velate the dichlorvos administration with the toxicity of the reproductive system, respiratory system, cardiovascular system and nervous system. Primarily, it affects the nervous system through cholinesterase inhibition or anticholinesterase effect and also leads to increased intracellular calcium level, oxidative stress and the rate of lipid peroxidation in the brain mitochondria. Mitochondria are pleomorphic organelles that generate the ATP supply for the cells. Disturbance in the electron transport chain (ETC) of mitochondria by the dichlorvos ultimately increases the ROS production, thereby leading to an increase of oxygen consumption and decrease of ATP synthesis which is the hallmark of the neuronal lesions. The increased calcium level is reported to be associated with neurodegenerative diseases such as Alzheimer’s disease, Parkinson’s disease and amyotrophic lateral sclerosis. Since, the brain is a controlling and coordinating organ in the body, therefore toxic effects of dichlorvos on it will also be deleterious to other organ-systems indirectly. Current review deals with possible implications of impairment of cholinergic circuit and brain mitochondrial functions carried by dichlorvos which may be the cause of potential neurotoxicity.