Live Imaging of Primary Cerebral Cortex Cells Using a 2D Culture System

Bruna Soares Landeira; J&#233;ssica Alves de Medeiros Ara&#250;jo; Timm Schroeder; Ulrich M&#252;ller; Marcos R. Costa

doi:10.3791/56063

Live Imaging of Primary Cerebral Cortex Cells Using a 2D Culture System

JoVE Journal
Neuroscience

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JoVE Journal Neuroscience

Live Imaging of Primary Cerebral Cortex Cells Using a 2D Culture System

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10:12 min

August 9, 2017

DOI: 10.3791/56063-v

Bruna Soares Landeira¹, Jéssica Alves de Medeiros Araújo¹, Timm Schroeder², Ulrich Müller³, Marcos R. Costa¹

¹Brain Institute,Federal University of Rio Grande do Norte, ²Department of Biosystems Science and Engineering,ETH Zurich, ³The Solomon H. Snyder Department of Neuroscience,Johns Hopkins University

Overview

This article outlines a detailed protocol for time-lapse video-microscopy aimed at observing cellular behaviors in primary cerebral cortex cells. The method enables researchers to investigate the lineage progression from neural stem cells to differentiated neurons and glial cells, providing insights into neurodevelopmental processes.

Key Study Components

Area of Science

Neurodevelopment
Cellular Behavior
Imaging Techniques

Background

Live imaging is an effective method to examine cellular dynamics.
The study focuses on neural stem cells transitioning into various cell types.
The method allows for long-term observation of cell lineage.
It helps in understanding key questions about cell division and growth rates.

Purpose of Study

To observe cell behaviors during lineage progression from neural stem cells to neurons or glial cells.
To investigate the implications of symmetric and asymmetric cell divisions.
To enable detailed tracking of neurogenesis and glial genesis.

Methods Used

The platform used is time-lapse video-microscopy of primary cerebral cortex cells.
Primary neural stem cells are isolated from e14 embryos following detailed dissection protocols.
The cells are cultured and observed under controlled temperature and CO2 conditions during imaging.
Images are captured at specified intervals to minimize phototoxicity, and software aids in tracking cell lineages.

Main Results

Cell lineage tracking reveals shifts from neurogenesis to glial genesis over time.
Quantitative data on cell viability and tracking outcomes are presented.
The increase in GFP expressing cells indicates successful tracking and differentiation observations.
Mechanistic insights into division, death, and lineage tracing are captured throughout the experiment.

Conclusions

This study demonstrates a robust method for live imaging to investigate neural lineage development.
The protocol enables researchers to answer critical questions in neurodevelopment, potentially aiding in the understanding of related diseases.
Findings from the study can inform future research on cell differentiation processes in the nervous system.

Frequently Asked Questions

What are the advantages of using live imaging?

Live imaging allows for real-time observation of cellular behaviors, enabling detailed analysis of lineage progression over extended periods.

How is the primary neural stem cell model implemented?

Neural stem cells are isolated from the embryonic telencephalon and cultured in a controlled environment to study their development.

What types of data are obtained from this method?

The method produces time-lapse images that track cell lineage, viability, and differentiation status, providing quantitative and qualitative insights.

Can this method be adapted for different cell types?

Yes, while this study focuses on cortical cells, the protocol can be modified to study other types of stem cells or progenitors by adjusting dissection and culture parameters.

What are some limitations of this approach?

Limitations may include the complexity of the dissection process and the potential for variabilities in cell culture conditions that can affect results.

Live imaging is a powerful tool to study cellular behaviors in real time. Here, we describe a protocol for time-lapse video-microscopy of primary cerebral cortex cells that allows a detailed examination of the phases enacted during the lineage progression from primary neural stem cells to differentiated neurons and glia.

The overall goal of this experiment is to observe cell behaviors during lineage progression from neural stem cells to neurons or glial cells. This method can help answering questions in the neurodevelopmental field such as the role of symmetric and asymmetric cell divisions, cell cycle lengthening and cell growth rates. The main advantage of this technique is that it allows the observation of cell lineage for long periods.

This permits the observation of switch from neurogenesis to glial genesis within a single lineage and a direct comporation of sublinear neural and glial progenitors. Begin by removing the brains from 5 to 10 e14 embryos in a Petri dish with cold dissection medium. Use forceps to carefully pull out the skin and skull and isolate the brains.

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