Single-Cell Redox Imaging Demonstrates a Distinctive Response of Dopaminergic Neurons to Oxidative Insults

Abstract

Aims: The study of the intracellular oxido-reductive (redox) state is of extreme relevance to the dopamine (DA) neurons of the substantia nigra pars compacta. These cells possess a distinct physiology intrinsically associated with elevated reactive oxygen species production, and they selectively degenerate in Parkinson's disease under oxidative stress conditions. To test the hypothesis that these cells display a unique redox response to mild, physiologically relevant oxidative insults when compared with other neuronal populations, we sought to develop a novel method for quantitatively assessing mild variations in intracellular redox state. Results: We have developed a new imaging strategy to study redox variations in single cells, which is sensitive enough to detect changes within the physiological range. We studied DA neurons' physiological redox response in biological systems of increasing complexity-from primary cultures to zebrafish larvae, to mammalian brains-and identified a redox response that is distinctive for substantia nigra pars compacta DA neurons. We studied simultaneously, and in the same cells, redox state and signaling activation and found that these phenomena are synchronized. Innovation: The redox histochemistry method we have developed allows for sensitive quantification of intracellular redox state in situ. As this method is compatible with traditional immunohistochemical techniques, it can be applied to diverse settings to investigate, in theory, any cell type of interest. Conclusion: Although the technique we have developed is of general interest, these findings provide insights into the biology of DA neurons in health and disease and may have implications for therapeutic intervention. Antioxid. Redox Signal. 15, 855-871.

Mitochondrial DNA damage: Molecular marker of vulnerable nigral neurons in Parkinson's disease

“…To test this possibility, we used a probe for mitochondrial H 2 O 2 , MitoPY1, and found a robust rotenone-induced H 2 O 2 signal in VMB neurons but not cortical neurons. Whether this reflects differences in production or degradation of ROS remains to be determined, but these results are consistent with other recent work showing that VMB neurons handle ROS differently than cortical neurons (53). In fact, rotenone-exposed cortical neurons compared with vehicle showed less mtDNA damage both in vitro and in vivo .…”

Impaired enzymatic defensive activity, mitochondrial dysfunction and proteasome activation are involved in RTT cell oxidative damage

“…Thus, the decreased activity of TrxR in RTT cells compared to controls adds further understanding of how oxidative stress occurs in RTT. Overall, these data confirm the relevance of mechanisms controlling thiol/disulfide equilibria in the pathogenesis RTT and, in this respect, extend to neurodevelopmental disorders previous findings obtained in chronic neurodegenerative diseases [42,43]. It should be also mentioned that in previous work we have detected a decreased levels of GSH in RTT cells [44] and this can indirectly affect TrxR by maintaining Trx in the reduced state during oxidative stress.…”

α-Synuclein binds to TOM20 and inhibits mitochondrial protein import in Parkinson’s disease

“…Consistent with this hypothesis, knockdown of endogenous α-synuclein in control rats ‘restored’ a TOM20:TOM22 PL signal in nigrostriatal neurons relative to the untransduced hemisphere (Figure 8A). Further, we have reported previously that, under basal conditions, nigrostriatal neurons exist in a more oxidized state than cortical or other dopaminergic neurons (31) – and results in figure 7 suggest that α-synuclein may contribute to this process. Here, in a separate cohort of animals, we found that AAV-sh[SNCA]-mediated knockdown of endogenous α-synuclein decreased basal levels of protein thiol oxidation in nigrostriatal neurons relative to the contralateral hemisphere, which received AAV-sh[control] (p<0.05; Figure 8B).…”