5 October 2007. A new technique that allows researchers to see the subcellular distribution of iron and other metals in neurons reveals that dopamine-containing neurotransmitter vesicles may function as iron storage depots. The implication is that perturbations in iron-dopamine interactions could contribute to the death of dopaminergic neurons in Parkinson disease, possibly by an increase in oxidative stress.
Consistent with a role for oxidative stress in the disease, another new study shows that the protein products of two different Parkinson-causing genes interact in a single pathway that protects cells against mitochondrial stress, and which appears altered in both sporadic and familial PD.
In the first paper, Richard Ortega and colleagues at Bordeaux University in Gradignan, France, tracked down iron in cells courtesy of a powerful, highly focused X-ray beam emanating from the European Synchrotron Radiation Facility in Grenoble, France. Because of the intense brightness of the beam and the high resolution of the images (down to 90 nm), the researchers were able for the first time to detect iron florescence in subcellular compartments of rat PC12 cells. They found iron mainly in dopamine-containing neurotransmitter vesicles. Overloading cells with iron increased the vesicular signal, particularly in neurite outgrowths and distal ends of the cells. Depleting the cells of dopamine caused a decrease in vesicular iron.
Dysfunctional iron metabolism has been implicated not only in Parkinson disease, but also in other neurodegenerative conditions. Published online in PLoS ONE on September 26, the new method will be generally useful for studying the distribution of iron and other metals in models of these diseases, the authors write. See open-access paper for X-ray fluorescence pictures of iron in dopamine cells.
Iron is essential for normal brain function. Local increases in brain or neuronal iron have been associated with PD, Alzheimer’s, and other neurodegenerative conditions (Oakley et al., 2007; Collingwood et al., 2005). Iron accumulation may even be useful as a biomarker for early PD: an ultrasound signal indicating high iron in the brain region affected by PD has been documented before cells begin to die off. The technique is now being tested as a clinical marker for early PD (Peng et al., 2007). Excess iron could cause increased oxidative damage, a pathway implicated in neurodegeneration generally. However, understanding exactly what iron is doing depends on knowing where it accumulates in cells.
In the new work, Ortega and colleagues first looked at iron distribution in normal dopamine-producing PC12 cells. They observed 200 nm cytosolic structures that contained most of the iron in the cell. Loading cells with additional iron increased the fluorescence associated with the vesicles. The scientists then went on to show that the vesicles also contained dopamine. When the researchers inhibited dopamine synthesis in the cells, the iron content of the vesicles decreased—results that are consistent with an iron-storage function for dopamine. The authors hypothesize that lowered dopamine or faulty storage of the neurotransmitter, both of which have been observed in PD, could raise levels of highly oxidizing cytosolic iron or iron-dopamine complexes, to the detriment of neurons. The researchers saw no such specific localization or changes in potassium or zinc distribution, which they measured with the same technique.
Blaming toxicity on excess iron or highly oxidizing iron-dopamine complexes fits with the idea that death of neurons in PD stems from oxidative stress. In further support of that notion, new work from the labs of Miguel Martins of the MRC in Leicester, UK, and Julian Downward at the Cancer Research UK London Research Institute, demonstrates that the protein product of PINK1, a putative mitochondrial kinase encoded at the PARK6 locus, associates with the apoptotic protease HtrA2 (encoded by the PARK13 gene). PINK1 appears necessary for the phosphorylation of HtrA2 after stimulation of the stress-activated p38 map kinase pathway, and loss of function of either protein sensitizes cells to oxidative stress. Thus, HtrA2, a protein previously proposed to be proapoptotic, appears to help protect cells from mitochondrial stress.
The work, published in the September 30 online issue of Nature Cell Biology, suggests that both proteins contribute resistance to apoptosis in response to mitochondrial stress. Thus, the protective effect recently discovered for PINK1 (Pridgeon et al., 2007 and see review by Abeliovich, 2007) may come at least in part from its ability to modulate HtrA2 activity.
PINK1 mutations cause rare, early-onset Parkinson disease, but the PINK1/HtrA2 pathway could play a role in the more frequent, sporadic form of PD as well, the authors claim. Looking at patient tissue, the investigators found that the phosphorylation of HtrA2 is decreased in patients with early-onset PD due to PINK1 mutations. However, the opposite occurs in sporadic PD, where HtrA2 phosphorylation is increased. Since phosphorylation appears to make HtrA2 more active, the results raise the possibility that HtrA2 is activated as a compensatory response to ongoing stress in sporadic PD, and that further understanding of this pathway could yield insights and novel therapeutic approaches to the most common form of the disease.—Pat McCaffrey.
Ortega R, Cloetens P, Deves G, Carmona A, Bohic S. Iron Storage within Dopamine Neurovesicles Revealed by Chemical Nano-Imaging. PLoS ONE. 2007 Sep 26;2(9):e925. Abstract
Plun-Favreau H, Klupsch K, Moisoi N, Gandhi S, Kjaer S, Frith D, Harvey K, Deas E, Harvey RJ, McDonald N, Wood NW, Martins LM, Downward J. The mitochondrial protease HtrA2 is regulated by Parkinson's disease-associated kinase PINK1. Nat Cell Biol. 2007 Sep 30; [Epub ahead of print] Abstract