As many astrocytes also abnormally contain a-synuclein in PD-affected regions, the increased Ndfip1 expression appears directly associated with the abnormal accumulation of a-synuclein in PD. In rodent injury models Ndfip1 has been identified as a neuroprotectant due to its upregulation in surviving neurons following stress caused through stroke or traumatic brain injury. Ndfip1 can be upregulated in response to extracellular stress such as hypoxia or metal toxicity, and this activation results in the regulation of a number of critical proteins such as DMT1. Previously we have observed that in an environment where metal concentrations are elevated, Ndfip1 is upregulated by a GDC-0199 mechanism that does not require either HSP70 or HIF-1a response, and resulted in the binding and degradation of DMT1. It should be noted that DMT1 abundance can also be regulated transcriptionally and differential expression of DMT1 isoforms having been observed due to iron exposure, hypoxia and NFkB activation. In the present report, we observed a protective function for Ndfip1 in mouse dopaminergic neurons where loss of Ndfip1 was found to increase dopaminergic neuron susceptibility to iron induced death. These results suggested that in PD Ndfip1 was upregulated in the substantia nigra as a stress response to protect cells against rising metal toxicity due to failing metal homeostasis. Indeed, our biochemical results from whole tissue lysates confirmed raised Ndfip1 levels in PD brains compared to controls, supporting our overall hypothesis. However, immunohistochemical staining of dopaminergic neurons showed no overall increase in the number of neurons containing Ndfip1 compared to controls. Instead, we found a significant increase in dopaminergic neurons containing both a-synuclein deposits and upregulated Ndfip1, suggesting heterogeneous patterns of Ndfip1 expression in dopaminergic neurons of the substantia nigra. The accumulation of a-synuclein has been shown to be secondary to iron concentrations, resulting in the formation of Lewy bodies, suggesting a link between iron concentrations and Ndfip1 observed in neurons containing a-synuclein deposits. Thus neurons containing both a-synuclein deposits and Ndfip1 may represent defensive snapshots of neurons undergoing metal stress. Due to the important association between metal accumulation and protein aggregation in PD, we tried to identify the relationship between Ndfip1 and DMT1 in PD brains. Despite extensive efforts we were unable to obtain reliable fluorescent staining for DMT1 in human tissue and could only identify its expression in dopaminergic neurons using peroxidase-conjugated antibody techniques. This technical limitation restricts our current understanding for the role of Ndfip1 in regulating DMT1 levels in the PD brain. The pattern of Ndfip1 upregulation in the PD substantia nigra holds parallels with Ndfip1 activation in cortical injury. In rodents after traumatic brain injury or stroke, Ndfip1 upregulation is maximal in surviving neurons at the lesion periphery rather than the core where most cells are necrotic.