Live-cell, label-free identification of GABAergic and non-GABAergic neurons in primary cortical cultures using micropatterned surface

Sho Kono, Hideaki Yamamoto, Takatoshi Kushida, Ayumi Hirano-Iwata, Michio Niwano, Takashi Tanii

    Research output: Contribution to journalArticle

    2 Citations (Scopus)

    Abstract

    Excitatory and inhibitory neurons have distinct roles in cortical dynamics. Here we present a novel method for identifying inhibitory GABAergic neurons from non-GABAergic neurons, which are mostly excitatory glutamatergic neurons, in primary cortical cultures. This was achieved using an asymmetrically designed micropattern that directs an axonal process to the longest pathway. In the current work, we first modified the micropattern geometry to improve cell viability and then studied the axon length from 2 to 7 days in vitro (DIV). The cell types of neurons were evaluated retrospectively based on immunoreactivity against GAD67, a marker for inhibitory GABAergic neurons. We found that axons of non-GABAergic neurons grow significantly longer than those of GABAergic neurons in the early stages of development. The optimal threshold for identifying GABAergic and non-GABAergic neurons was evaluated to be 110 μm at 6 DIV. The method does not require any fluorescence labelling and can be carried out on live cells. The accuracy of identification was 98.2%. We confirmed that the high accuracy was due to the use of a micropattern, which standardized the development of cultured neurons. The method promises to be beneficial both for engineering neuronal networks in vitro and for basic cellular neuroscience research.

    Original languageEnglish
    Article numbere0160987
    JournalPLoS One
    Volume11
    Issue number8
    DOIs
    Publication statusPublished - 2016 Aug 1

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    Neurons
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    neurons
    GABAergic Neurons
    cells
    Axons
    axons
    Neurosciences
    neurophysiology
    Cell Survival
    Fluorescence
    Labeling
    cell viability
    engineering
    methodology
    Cells
    fluorescence
    Research
    In Vitro Techniques
    Geometry

    ASJC Scopus subject areas

    • Medicine(all)
    • Biochemistry, Genetics and Molecular Biology(all)
    • Agricultural and Biological Sciences(all)

    Cite this

    Live-cell, label-free identification of GABAergic and non-GABAergic neurons in primary cortical cultures using micropatterned surface. / Kono, Sho; Yamamoto, Hideaki; Kushida, Takatoshi; Hirano-Iwata, Ayumi; Niwano, Michio; Tanii, Takashi.

    In: PLoS One, Vol. 11, No. 8, e0160987, 01.08.2016.

    Research output: Contribution to journalArticle

    Kono, Sho ; Yamamoto, Hideaki ; Kushida, Takatoshi ; Hirano-Iwata, Ayumi ; Niwano, Michio ; Tanii, Takashi. / Live-cell, label-free identification of GABAergic and non-GABAergic neurons in primary cortical cultures using micropatterned surface. In: PLoS One. 2016 ; Vol. 11, No. 8.
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