Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2021 Mar;1(3):e64.
doi: 10.1002/cpz1.64.

Ultrafast Sodium Imaging of the Axon Initial Segment of Neurons in Mouse Brain Slices

Laila Ananda Blömer  1   2 , Marco Canepari  1   2   3 , Luiza Filipis  1   2
Affiliations

Ultrafast Sodium Imaging of the Axon Initial Segment of Neurons in Mouse Brain Slices

Laila Ananda Blömer et al. Curr Protoc. 2021 Mar.

Erratum in

Abstract

Monitoring Na+ influx in the axon initial segment (AIS) at high spatial and temporal resolution is fundamental to understanding the generation of an action potential (AP). Here, we present protocols to obtain this measurement, focusing on the AIS of layer 5 (L5) somatosensory cortex pyramidal neurons in mouse brain slices. We first outline how to prepare slices for this application, how to select and patch neurons, and how to optimize the image acquisition. Specifically, we describe the preparation of optimal slices, patching and loading of L5 pyramidal neurons with the Na+ indicator ING-2, and Na+ imaging at 100 µs temporal resolution with a pixel resolution of half a micron. Then, we present a data analysis strategy in order to extract information on the kinetics of activated voltage-gated Na+ channels by determining the change in Na+ by compensating for bleaching and calculating the time derivative of the resulting fit. In sum, this approach can be widely applied when investigating the function of Na+ channels during initiation of an AP and propagation under physiological or pathological conditions in neuronal subtypes. © 2021 Wiley Periodicals LLC. Basic Protocol 1: Preparation of cortical slices Basic Protocol 2: Selection, patching, and Na+ fluorescence recording of a neuron Support Protocol: Calibrating Na+ fluorescence Basic Protocol 3: Data analysis.

Keywords: action potential; axon; pyramidal neuron; sodium imaging; somatosensory cortex; voltage-gated sodium channels.

PubMed Disclaimer

References

Literature Cited

    1. Ait Ouares, K., Filipis, L., Tzilivaki, A., Poirazi, P., & Canepari, M. (2019). Two distinct sets of Ca2+ and K+ channels are activated at different membrane potentials by the climbing fiber synaptic potential in purkinje neuron dendrites. Journal of Neuroscience, 39, 1969-1981. doi: 10.1523/JNEUROSCI.2155-18.2018.
    1. Ait Ouares, K., Jaafari, N., & Canepari, M. (2016). A generalised method to estimate the kinetics of fast Ca2+ currents from Ca2+ imaging experiments. Journal of Neuroscience Methods, 268, 66-77. doi: 10.1016/j.jneumeth.2016.05.005.
    1. Alle, H., Roth, A., & Geiger, J. R. (2009). Energy-efficient action potentials in hippocampal mossy fibers. Science, 325, 1405-1408. doi: 10.1126/science.1174331.
    1. Andreone, B. J., Lacoste, B., & Gu, C. (2015). Neuronal and vascular interactions. Annual Review of Neuroscience, 38, 25-46. doi: 10.1146/annurev-neuro-071714-033835.
    1. Astman, N., Gutnick, M. J., & Fleidervish, I. A. (2006). Persistent sodium current in layer 5 neocortical neurons is primarily generated in the proximal axon. Journal of Neuroscience, 26, 3465-3473. doi: 10.1523/JNEUROSCI.4907-05.2006.

LinkOut - more resources