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Heterogeneous responses to antioxidants in noradrenergic neurons of the Locus coeruleus indicate differing susceptibility to free radical content.

de Oliveira RB, Gravina FS, Lim R, Brichta AM, Callister RJ, van Helden DF - Oxid Med Cell Longev (2012)

Bottom Line: In current clamp experiments, most neurons (55%; 6/11) did not respond to the antioxidants.Calcium and JC-1 imaging demonstrated that these effects did not change intracellular Ca(2+) concentration but may influence mitochondrial function as both antioxidant treatments modulated mitochondrial membrane potential.If this is the case, there may be a protective role for antioxidant therapies.

View Article: PubMed Central - PubMed

Affiliation: School of Biomedical Sciences and Pharmacy, The University of Newcastle and Hunter Medical Research Institute, Newcastle, NSW 2308, Australia.

ABSTRACT
The present study investigated the effects of the antioxidants trolox and dithiothreitol (DTT) on mouse Locus coeruleus (LC) neurons. Electrophysiological measurement of action potential discharge and whole cell current responses in the presence of each antioxidant suggested that there are three neuronal subpopulations within the LC. In current clamp experiments, most neurons (55%; 6/11) did not respond to the antioxidants. The remaining neurons exhibited either hyperpolarization and decreased firing rate (27%; 3/11) or depolarization and increased firing rate (18%; 2/11). Calcium and JC-1 imaging demonstrated that these effects did not change intracellular Ca(2+) concentration but may influence mitochondrial function as both antioxidant treatments modulated mitochondrial membrane potential. These suggest that the antioxidant-sensitive subpopulations of LC neurons may be more susceptible to oxidative stress (e.g., due to ATP depletion and/or overactivation of Ca(2+)-dependent pathways). Indeed it may be that this subpopulation of LC neurons is preferentially destroyed in neurological pathologies such as Parkinson's disease. If this is the case, there may be a protective role for antioxidant therapies.

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Effect of Trolox and DTT treatment in [Ca2+]c and Ψm in control and impaired conditions. (a) Graph demonstrating that both antioxidants did not impact on [Ca2+]c in control conditions (n = 5 for both). (b) Graph demonstrating that 100 μM Trolox co-treatment with 1 μM CCCP increased [Ca2+]c and 1 mM DDT did not effect the normal CCCP-induced increase in [Ca2+]c (n = 6 for Trolox n = 5 for DTT cotreatments, and n = 7 for CCCP by itself). (c) JC-1 fluorescence demonstrating the impact of Trolox and DTT in Ψm in control and impaired (1 μM CCCP cotreatment) conditions (n = 8 for Trolox and n = 9 for DTT by themselves; n = 12 for Trolox and n = 10 for DTT cotreatments; n = 9 for CCCP by itself). All graphs show mean ± SEM.
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fig6: Effect of Trolox and DTT treatment in [Ca2+]c and Ψm in control and impaired conditions. (a) Graph demonstrating that both antioxidants did not impact on [Ca2+]c in control conditions (n = 5 for both). (b) Graph demonstrating that 100 μM Trolox co-treatment with 1 μM CCCP increased [Ca2+]c and 1 mM DDT did not effect the normal CCCP-induced increase in [Ca2+]c (n = 6 for Trolox n = 5 for DTT cotreatments, and n = 7 for CCCP by itself). (c) JC-1 fluorescence demonstrating the impact of Trolox and DTT in Ψm in control and impaired (1 μM CCCP cotreatment) conditions (n = 8 for Trolox and n = 9 for DTT by themselves; n = 12 for Trolox and n = 10 for DTT cotreatments; n = 9 for CCCP by itself). All graphs show mean ± SEM.

Mentions: Due to the heterogeneous nature of the responses produced by the two antioxidant treatments, we next investigated if the activation of different currents was related to the cytosolic Ca2+ concentration ([Ca2+]c) in both control conditions and those where mitochondrial function was impaired. Figure 6(a) demonstrates that under control conditions, neither Trolox or DTT treatments were able to increase [Ca2+]c, suggesting that Ca2+-activated pathways are not involved in the induction/modulation of ionic currents by these antioxidants. When CCCP was added to impair mitochondrial function, Trolox (100 μM) caused a further enhancement of the CCCP-induced increase in [Ca2+]c (Figure 6(b)). In contrast, cotreatment with DTT (1 mM) had no impact on the CCCP-induced increase in [Ca2+]c (Figure 6(b)). The consequences of the Trolox-induced increase in [Ca2+]c are unclear given that Trolox was less effective than DTT in modulating CCCP-related voltage dependent currents (Figure 5(a)).


Heterogeneous responses to antioxidants in noradrenergic neurons of the Locus coeruleus indicate differing susceptibility to free radical content.

de Oliveira RB, Gravina FS, Lim R, Brichta AM, Callister RJ, van Helden DF - Oxid Med Cell Longev (2012)

Effect of Trolox and DTT treatment in [Ca2+]c and Ψm in control and impaired conditions. (a) Graph demonstrating that both antioxidants did not impact on [Ca2+]c in control conditions (n = 5 for both). (b) Graph demonstrating that 100 μM Trolox co-treatment with 1 μM CCCP increased [Ca2+]c and 1 mM DDT did not effect the normal CCCP-induced increase in [Ca2+]c (n = 6 for Trolox n = 5 for DTT cotreatments, and n = 7 for CCCP by itself). (c) JC-1 fluorescence demonstrating the impact of Trolox and DTT in Ψm in control and impaired (1 μM CCCP cotreatment) conditions (n = 8 for Trolox and n = 9 for DTT by themselves; n = 12 for Trolox and n = 10 for DTT cotreatments; n = 9 for CCCP by itself). All graphs show mean ± SEM.
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig6: Effect of Trolox and DTT treatment in [Ca2+]c and Ψm in control and impaired conditions. (a) Graph demonstrating that both antioxidants did not impact on [Ca2+]c in control conditions (n = 5 for both). (b) Graph demonstrating that 100 μM Trolox co-treatment with 1 μM CCCP increased [Ca2+]c and 1 mM DDT did not effect the normal CCCP-induced increase in [Ca2+]c (n = 6 for Trolox n = 5 for DTT cotreatments, and n = 7 for CCCP by itself). (c) JC-1 fluorescence demonstrating the impact of Trolox and DTT in Ψm in control and impaired (1 μM CCCP cotreatment) conditions (n = 8 for Trolox and n = 9 for DTT by themselves; n = 12 for Trolox and n = 10 for DTT cotreatments; n = 9 for CCCP by itself). All graphs show mean ± SEM.
Mentions: Due to the heterogeneous nature of the responses produced by the two antioxidant treatments, we next investigated if the activation of different currents was related to the cytosolic Ca2+ concentration ([Ca2+]c) in both control conditions and those where mitochondrial function was impaired. Figure 6(a) demonstrates that under control conditions, neither Trolox or DTT treatments were able to increase [Ca2+]c, suggesting that Ca2+-activated pathways are not involved in the induction/modulation of ionic currents by these antioxidants. When CCCP was added to impair mitochondrial function, Trolox (100 μM) caused a further enhancement of the CCCP-induced increase in [Ca2+]c (Figure 6(b)). In contrast, cotreatment with DTT (1 mM) had no impact on the CCCP-induced increase in [Ca2+]c (Figure 6(b)). The consequences of the Trolox-induced increase in [Ca2+]c are unclear given that Trolox was less effective than DTT in modulating CCCP-related voltage dependent currents (Figure 5(a)).

Bottom Line: In current clamp experiments, most neurons (55%; 6/11) did not respond to the antioxidants.Calcium and JC-1 imaging demonstrated that these effects did not change intracellular Ca(2+) concentration but may influence mitochondrial function as both antioxidant treatments modulated mitochondrial membrane potential.If this is the case, there may be a protective role for antioxidant therapies.

View Article: PubMed Central - PubMed

Affiliation: School of Biomedical Sciences and Pharmacy, The University of Newcastle and Hunter Medical Research Institute, Newcastle, NSW 2308, Australia.

ABSTRACT
The present study investigated the effects of the antioxidants trolox and dithiothreitol (DTT) on mouse Locus coeruleus (LC) neurons. Electrophysiological measurement of action potential discharge and whole cell current responses in the presence of each antioxidant suggested that there are three neuronal subpopulations within the LC. In current clamp experiments, most neurons (55%; 6/11) did not respond to the antioxidants. The remaining neurons exhibited either hyperpolarization and decreased firing rate (27%; 3/11) or depolarization and increased firing rate (18%; 2/11). Calcium and JC-1 imaging demonstrated that these effects did not change intracellular Ca(2+) concentration but may influence mitochondrial function as both antioxidant treatments modulated mitochondrial membrane potential. These suggest that the antioxidant-sensitive subpopulations of LC neurons may be more susceptible to oxidative stress (e.g., due to ATP depletion and/or overactivation of Ca(2+)-dependent pathways). Indeed it may be that this subpopulation of LC neurons is preferentially destroyed in neurological pathologies such as Parkinson's disease. If this is the case, there may be a protective role for antioxidant therapies.

Show MeSH
Related in: MedlinePlus