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Abstract

Neuroscience

In Vivo Two-photon Imaging of Cortical Neurons in Neonatal Mice

Published: October 18th, 2018

DOI:

10.3791/58340

1International Research Center for Medical Sciences (IRCMS), Kumamoto University, 2Division of Neurogenetics, National Institute of Genetics, 3Department of Genetics, SOKENDAI (The Graduate University for Advanced Studies)

Abstract

Two-photon imaging is a powerful tool for the in vivo analysis of neuronal circuits in the mammalian brain. However, a limited number of in vivo imaging methods exist for examining the brain tissue of live newborn mammals. Herein we summarize a protocol for imaging individual cortical neurons in living neonatal mice. This protocol includes the following two methodologies: (1) the Supernova system for sparse and bright labeling of cortical neurons in the developing brain, and (2) a surgical procedure for the fragile neonatal skull. This protocol allows the observation of temporal changes of individual cortical neurites during neonatal stages with a high signal-to-noise ratio. Labeled cell-specific gene silencing and knockout can also be achieved by combining the Supernova with RNA interference and CRISPR/Cas9 gene editing systems. This protocol can, thus, be used for analyzing the developmental dynamics of cortical neurons, molecular mechanisms that control the neuronal dynamics, and changes in neuronal dynamics in disease models.

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Keywords In Vivo

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