RESUMEN
Manganese (Mn) toxicity is partially mediated by reduced ATP production. We have used oxidation rate assays--a measure of ATP production--under rapid phosphorylation conditions to explore sites of Mn(2+) inhibition of ATP production in isolated liver, brain, and heart mitochondria. This approach has several advantages. First, the target tissue for Mn toxicity in the basal ganglia is energetically active and should be studied under rapid phosphorylation conditions. Second, Mn may inhibit metabolic steps which do not affect ATP production rate. This approach allows identification of inhibitions that decrease this rate. Third, mitochondria from different tissues contain different amounts of the components of the metabolic pathways potentially resulting in different patterns of ATP inhibition. Our results indicate that Mn(2+) inhibits ATP production with very different patterns in liver, brain, and heart mitochondria. The primary Mn(2+) inhibition site in liver and heart mitochondria, but not in brain mitochondria, is the F1F0 ATP synthase. In mitochondria fueled by either succinate or glutamate+malate, ATP production is much more strongly inhibited in brain than in liver or heart mitochondria; moreover, Mn(2+) inhibits two independent sites in brain mitochondria. The primary site of Mn-induced inhibition of ATP production in brain mitochondria when succinate is substrate is either fumarase or complex II, while the likely site of the primary inhibition when glutamate plus malate are the substrates is either the glutamate/aspartate exchanger or aspartate aminotransferase.
Asunto(s)
Encéfalo/efectos de los fármacos , Manganeso/farmacología , Mitocondrias Cardíacas/efectos de los fármacos , Mitocondrias Hepáticas/efectos de los fármacos , Fosforilación Oxidativa/efectos de los fármacos , Adenosina Trifosfato/antagonistas & inhibidores , Adenosina Trifosfato/biosíntesis , Animales , Encéfalo/metabolismo , Femenino , Mitocondrias Cardíacas/metabolismo , Mitocondrias Hepáticas/metabolismo , Oxidación-Reducción/efectos de los fármacos , Ratas , Ratas Sprague-DawleyRESUMEN
Recent studies of speciation of manganese (Mn) in brain mitochondria, neuron-like cells, and astrocytes are reviewed. No evidence is found for oxidation of Mn(2+) complexes to a Mn(3+) complex. The only evidence for any Mn(3+) complex is found in a spectrum essentially identical to that of mitochondrial manganese superoxide dismutase (MnSOD). While this does not prove that no Mn(3+) is produced in these tissues by oxidation of Mn(2+), it does suggest that formation of an active Mn(3+) complex by oxidation of Mn(2+) probably does not play as important a role in Mn toxicity as has been suggested earlier. Since these results suggest that we should look elsewhere for the proximal causes of Mn neurotoxicity, we consider the possibilities that Mn(3+) may be transported into the cell via transferrin and that Mn(2+) may inhibit Ca(2+)-activation and control of the rate of ATP production by oxidative phosphorylation.
Asunto(s)
Células/ultraestructura , Magnesio/farmacocinética , Intoxicación por Manganeso/metabolismo , Intoxicación por Manganeso/patología , Mitocondrias/patología , Animales , Células/metabolismo , Células/patología , Humanos , Espectrometría por Rayos X/métodos , Superóxido Dismutasa/metabolismoRESUMEN
X-ray absorption near edge structure (XANES) spectroscopy can provide information on the oxidation state of metal ions within a biological sample and also the complexes in which it is found. This type of information could be of great use to toxicologists in understanding the mechanism of action of many toxic agents. The prospect of using a sophisticated physical technique such as XANES may be somewhat intimidating for those without a strong physical background. Here, we explain the concepts necessary to understand XANES spectroscopy at a level that can be easily understood by biological scientists without a strong physics background and describe useful sample preparation and data analysis techniques which can be adapted for a variety of applications. Examples are taken from an ongoing study of manganese in brain mitochondria and neuron-like cells.
Asunto(s)
Espectrometría por Rayos X/métodos , Toxicología/instrumentación , Toxicología/métodos , Animales , Química Encefálica , Humanos , Manganeso/análisis , Mitocondrias/química , Neuronas/química , Espectrometría por Rayos X/instrumentaciónRESUMEN
Manganese (Mn) is an essential dietary nutrient, but an excess or accumulation can be toxic. Disease states, such as manganism, are associated with overexposure or accumulation of Mn and are due to the production of reactive oxygen species, free radicals, and toxic metabolites; alteration of mitochondrial function and ATP production; and depletion of cellular antioxidant defense mechanisms. This review focuses on all of the preceding mechanisms and the scientific studies that support them as well as providing an overview of the absorption, distribution, and excretion of Mn and the stability and transport of Mn compounds in the body.
Asunto(s)
Intoxicación por Manganeso , Manganeso/metabolismo , Neuronas/metabolismo , Estrés Oxidativo , Adenosina Trifosfato/biosíntesis , Antioxidantes/metabolismo , Radicales Libres/metabolismo , Humanos , Manganeso/farmacología , Mitocondrias/metabolismo , Mitocondrias/patología , Neuronas/patología , Especies Reactivas de OxígenoRESUMEN
Excess brain manganese can produce toxicity with symptoms that resemble those of Parkinsonism and causes that remain elusive. Manganese accumulates in mitochondria, a major source of superoxide, which can oxidize Mn2+ to the powerful oxidizing agent Mn3+. Oxidation of important cell components by Mn3+ has been suggested as a cause of the toxic effects of manganese. Determining the oxidation states of intramitochondrial manganese could help to identify the dominant mechanism of manganese toxicity. Using X-ray absorbance near edge structure (XANES) spectroscopy, we have characterized the oxidation state of manganese in mitochondria isolated from brain, liver, and heart over concentrations ranging from physiological to pathological. Results showed that (i) spectra from different model manganese complexes of the same oxidation state were similar to each other and different from those of other oxidation states and that the position of the absorption edge increases with oxidation state; (ii) spectra from intramitochondrial manganese in isolated brain, heart and liver mitochondria were virtually identical; and (iii) under these conditions intramitochondrial manganese exists primarily as a combination of Mn2+ complexes. No evidence for Mn3+ was detected in samples containing more than endogenous manganese levels, even after incubation under conditions promoting reactive oxygen species (ROS) production. While the presence of Mn3+ complexes cannot be proven in the spectrum of endogenous mitochondrial manganese, the shape of this spectrum could suggest the presence of Mn3+ near the limit of detection, probably as MnSOD.