Abstract
The axon initial segment (AIS) is a specialized region within the proximal portion of the axon that initiates action potentials thanks in large part to an enrichment of sodium channels. The scaffolding protein ankyrinG (AnkG) is essential for the recruitment of sodium channels as well as several other intracellular and extracellular proteins to the AIS. In the present study, we explore the role of the cell adhesion molecule (CAM) neurofascin-186 (NF-186) in arranging the individual molecular components of the AIS in cultured rat hippocampal neurons. Using a CRISPR depletion strategy to ablate NF expression, we found that the loss of NF selectively perturbed AnkG accumulation and its relative proximal distribution within the AIS. We found that the overexpression of sodium channels could restore AnkG accumulation, but not its altered distribution within the AIS without NF present. We go on to show that although the loss of NF altered AnkG distribution, sodium channel function within the AIS remained normal. Taken together, these results demonstrate that the regulation of AnkG and sodium channel accumulation within the AIS can occur independently of one another, potentially mediated by other binding partners such as NF.
Highlights
Neurons are the most polarized electrically excitable cells, allowing for the rapid transfer of information throughout the nervous system
We evaluated protein enrichment using immunostaining against AnkG, panNav, and NF
Accumulation of AnkG staining at the axon initial segment (AIS) was rapid, with discernable enrichment compared to the soma found in 10 ± 2% of neurons at DIV1 (n = 39 fields of view; an average of five cells were contained in each field of view), and appeared in a majority of neurons on DIV3 (Figures 1A,B)
Summary
Neurons are the most polarized electrically excitable cells, allowing for the rapid transfer of information throughout the nervous system. A second important factor that modulates firing of the action potential is the isolation of the AIS from the somatodendritic compartment, NF-186 Modulates AnkG AIS Enrichment which generates a capacitive and conductive load that acts as a Na+ current sink and inhibits excitation (Brette, 2013; Eyal et al, 2014). The degree of this Na+ current sink is influenced by the morphology of the somato-dendritic compartment (Gulledge and Bravo, 2016; Hamada et al, 2016; Jamann et al, 2018; Kole and Brette, 2018). Factors controlling the location and function of Nav within the AIS are critical influences on excitability, though this influence will vary across cell types due to their respective morphology
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