| Summary: | The axon initial segment (AIS) is a highly specialised neuronal subcompartment located within the proximal axon that has a crucial role in the generation of action potentials. Recent research has shown that this structure is plastic and can undergo changes in its length or location over a period of days, as a homeostatic response to perturbation in input. Here, I use a dissociated hippocampal culture model to investigate plasticity at the AIS in more depth. In order to reduce the variability brought about by cell-subtype heterogeneity, I focussed on a single excitatory subtype, the dentate granule cell (DGC), which I show reliably undergoes activity-dependent AIS relocation over a period of 48 h. I then use this model in combination with pharmacological, electrophysiological and genetic approaches to show that AIS relocation requires signalling through the intracellular phosphatase calcineurin, which acts downstream of L-type (Cav1) voltage-gated calcium channel activation. Moreover, calcineurin signalling is sufficient for AIS relocation. Whilst studying the timecourse of AIS relocation, I also discovered that the AIS is subject to a much more rapid form of plasticity. In response to 3 h depolarisation, AIS length shortened by approximately 25% as defined by immunostaining targeting important constituent AIS proteins. Interestingly, this process is largely reliant on the same signalling pathways as AIS relocation. Finally, to investigate whether this rapid structural change at the AIS has any direct consequences for DGC function, I combined targeted patch clamping of DGCs with a live label for AIS length. DGCs treated with +15 mM KCl had reduced excitability in multiple spiking, as well changes in AP waveform. Together, these findings demonstrate that calcineurin signalling is vital for AIS relocation and that plasticity at the AIS can occur over a much more rapid timescale than previously thought.
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