Mitochondria play a vital role in most eukaryotic cells. They are particularly influential in neurons as these cells are highly-energy consuming and postmitotic (and therefore susceptible to the accumulation of damage over a lifetime). Mitochondria are essential for ATP production along with other cellular aspects such as intracellular Ca2+ signalling that modulates synaptic activity, and the generation of reactive oxygen species. Neurons depend on mitochondrial ATP production to maintain excitability and neurotransmission. Mitochondrial dysfunction is associated with many neurodegenerative diseases, potentially resulting from Ca2+-overload, which restricts ATP production and disrupts ROS balance leading to neuronal cell death. For example, in the early stages of Alzheimer’s disease, a build-up of extracellular amyloid-β may inhibit mitochondrial motility which could lead to mis-localisation of the organelle and subsequent cellular decline. This altered mitochondrial motility in neurons has also recently been reported to be induced due to elevation of extracellular glucose concentration, such as that which can arise due to metabolic disorders. Indeed, diabetic patients have a higher risk of developing Alzheimer’s disease, and at a faster rate of onset, than the non-diabetic population.
We used epifluorescence microscopy to investigate whether either chronic or acute exposure of primary hippocampal cells to hyperglycaemia (25mM glucose) affected the motility of mitochondria. Primary cultures were maintained until synaptically-mature (3 weeks) in either “standard” neuronal media (containing 25 mM glucose) or were switched into a more-physiological media containing 3 mM glucose for the final 5 or 10 days prior to loading cells with MitoTracker Green and live-cell imaging mitochondrial motility. Image stacks were analysed in ImageJ to quantify global mitochondrial motility (the change in mitochondrial position across the imaging region, normalised to total mitochondrial area). There was no difference in the baseline amount of mitochondrial movement between cells grown in 25 mM glucose throughout, compared to those switched into 3 mM glucose for either 5 or 10 days (all cells imaged at same final age). Furthermore, an acute increase of extracellular glucose from 3 mM to 25 mM for 1 hour also did not alter global mitochondrial motility (measured before and after, cells maintained in situ on the microscope). These results are in contrast with similar investigations published by other research groups and may reflect differences in methods of quantifying mitochondrial motility, differences in whole-region versus axon-only analyses or differences in the experimental protocol such as cell age.
|Date of Award||10 Jun 2021|
- University Of Strathclyde
|Supervisor||Susan Chalmers (Supervisor)|