Investigating mitochondrial trafficking with dopaminergic neurons of the nigrostriatal pathway

Abstract

Parkinson’s disease (PD) is an age-related neurodegenerative disorder affecting more than 1% of the population aged over 60 years of age. PD is characterised by the loss of dopaminergic (DAergic) neurons from the substantia nigra pars compacta (SNpc), with mitochondrial dysfunction believed to be a contributor to this degeneration. The finding that complex I deficits can trigger Parkinsonism in the 1980s, has resulted in growing evidence of mitochondrial impairment in PD and ageing. Alongside the finding that SNpc neurons exhibit increased mtDNA deletion levels, these data suggest mitochondrial defects render them highly vulnerable in PD. These large scale mtDNA deletions have also been associated with the DAergic neuronal loss seen in normal ageing. Mitochondria are dynamic organelles, whose distribution is regulated by organelle transport to balance the energy needs of neurons. Axonal mitochondria travel long and short distances and genetic mutations in the machinery which controls these movements have been identified in early onset PD. Fundamental insight into the mitochondrial transport system has arisen from yeast and fly models, the effects however of altered mitochondrial movement on neuronal function still remain unanswered with a mammalian PD model. Hence, this study investigated mitochondrial trafficking within neurons via two disease models. The first involved the optimisation of a method to observe the movements of fluorescently tagged mitochondria within DAergic neurons in ex vivo nigrostriatal brain slices. Young and old mice were studied to understand the effects of age on these movements. Changes in these movements in the presence of PD pathology were studied in human wild type alpha-synuclein expressing mice. The second study utilised induced pluripotent stem cells (IPSCs) to compare modifications in mitochondrial trafficking in SNpc neurons from IPSCs derived from a mitochondrial disease patient harbouring a large scale mtDNA deletion. Mitochondrial movement was first observed within low and high mtDNA deletion cell lines, to ascertain the effect of mitochondrial dysfunction on the movement of these organelles. Following this initial study, the effect of mitochondrial Ca2+ extrusion into the cytoplasm was investigated, alongside how this mediated mitochondrial motility. This study successfully managed to obtain videos of mitochondrial motility, from cell body to synapse, allowing the observation of speed, directionality and membrane potential of mitochondria within neurons of these disease models. These models have collectively provided information on the movements and trafficking of mitochondria in both culture and in a mouse model, presenting a promising method to comprehend the consequences of mitochondrial dysfunction in PD.