RESUMEN
The axon initial segment (AIS) constitutes not only the site of action potential initiation, but also a hub for activity-dependent modulation of output generation. Recent studies shedding light on AIS function used predominantly post-hoc approaches since no robust murine in vivo live reporters exist. Here, we introduce a reporter line in which the AIS is intrinsically labeled by an ankyrin-G-GFP fusion protein activated by Cre recombinase, tagging the native Ank3 gene. Using confocal, superresolution, and two-photon microscopy as well as whole-cell patch-clamp recordings in vitro, ex vivo, and in vivo, we confirm that the subcellular scaffold of the AIS and electrophysiological parameters of labeled cells remain unchanged. We further uncover rapid AIS remodeling following increased network activity in this model system, as well as highly reproducible in vivo labeling of AIS over weeks. This novel reporter line allows longitudinal studies of AIS modulation and plasticity in vivo in real-time and thus provides a unique approach to study subcellular plasticity in a broad range of applications.
RESUMEN
Dysfunction in sodium channels and their ankyrin scaffolding partners have both been implicated in neurodevelopmental disorders, including autism spectrum disorder (ASD). In particular, the genes SCN2A, which encodes the sodium channel NaV1.2, and ANK2, which encodes ankyrin-B, have strong ASD association. Recent studies indicate that ASD-associated haploinsufficiency in Scn2a impairs dendritic excitability and synaptic function in neocortical pyramidal cells, but how NaV1.2 is anchored within dendritic regions is unknown. Here, we show that ankyrin-B is essential for scaffolding NaV1.2 to the dendritic membrane of mouse neocortical neurons and that haploinsufficiency of Ank2 phenocopies intrinsic dendritic excitability and synaptic deficits observed in Scn2a+/- conditions. These results establish a direct, convergent link between two major ASD risk genes and reinforce an emerging framework suggesting that neocortical pyramidal cell dendritic dysfunction can contribute to neurodevelopmental disorder pathophysiology.