CHARACTERISING MITOCHONDRIAL REMODELLING DURING NEUROGENESIS

Neurodifferentiation is an energy intensive process, driven by mitochondrial reprogramming that is critical for neuronal identity, development and function. This process is mediated by dynamic changes to mitochondrial networks that remain poorly understood. Here, we characterise the mitochondrial remodelling that drives neurogenesis in the presence and absence of metabolic stress induced by propionic acid (PPA). First, we established a model for neurogenesis in SH-SY5Y cells, using immunocytochemistry to examine molecular and morphological hallmarks of differentiation. We quantified length and branching of dendritic extensions and measured the expression of the mature dendritic marker, MAP2. Complementary confocal microscopy, qPCRs and image analysis workflows were employed to quantify mitochondrial content, morphology and connectivity. Lastly, we characterised how PPA influenced neuronal morphology and mitochondrial connectivity during neurodifferentiation. We demonstrated efficient differentiation on a population-wide and single-cell level, showing an increase in the B3T:Nestin ratio, as well as an increase in the length and complexity of MAP2+ dendritic extensions. Significant changes to mitochondrial content, morphology and connectivity were observed, indicating a change in the balance between fission and fusion dynamics. PPA exposure significantly disrupted neuronal morphology and decreased the production of tubular dendritic extensions. Concurrently, PPA altered both mitochondrial content and connectivity, increasing mitochondrial volume and fragmentation. Together, this work provides a quantitative assessment of mitochondrial remodelling during differentiation, demonstrating how metabolic stress reshapes mitochondrial networks. These data inform subsequent studies of neurogenesis in vitro, and yield insight into the changes to mitochondrial dynamics and structure that facilitate neuronal maturation.