The axonal initial segment (AIS) is a unique sub-domain that plays a central role in the physiology of the neuron, as it orchestrates both electrogenesis and the maintenance of neuronal polarity. The voltage-gated ion channels responsible for generating action potentials are concentrated at the plasma membrane of the AIS through interactions with the scaffolding protein ankyrin G (ankG). AnkG also binds to adhesion protein Neurofascin-186 and NrCAM, and links membrane proteins to the actin cytoskeleton through β4-spectrin, and to microtubules through End Binding proteins EB3 and EB1. The precise location and length of the AIS can vary from neuron to neuron and thus influence neuronal function. Moreover, recent studies revealed that the entire AIS is a dynamic structure, adapting its length and its position along development and in response to electrical activity.Since the cellular morphology of the AIS regulates neuronal excitability, it is crucial to understand the detailed molecular mechanisms accounting for AIS stabilization and positioning along the proximal axon. AIS maintenance depends on the stabilization of membrane and cytoskeletal protein complexes, but also on distal axon properties. We have recently discovered new AIS components and I will present their role in the proper AIS positioning and morphology.