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Media formulation influences chemical effects on neuronal growth and morphology

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Abstract

Screening for developmental neurotoxicity using in vitro, cell-based systems has been proposed as an efficient alternative to performing in vivo studies. One tool currently used for developmental neurotoxicity screening is automated high-content imaging of neuronal morphology. While high-content imaging (HCI) has been demonstrated to be useful in detection of potential developmental neurotoxicants, comparison of results between laboratories or assays can be complicated due to methodological differences. In order to determine whether high-content imaging-based developmental neurotoxicity assays can be affected by differences in media formulation, a systematic comparison of serum-supplemented (Dulbecco’s modified Eagle’s media (DMEM) + 10% serum) and serum-free (Neurobasal A + B27) culture media on neuronal morphology was performed using primary rat cortical neurons. Concentration–response assays for neuritogenesis, axon and dendrite outgrowth, and synaptogenesis were performed in each media type using chemicals with previously demonstrated effects. Marked qualitative and quantitative differences in the characteristics of neurons cultured in the two media types were observed, with increased neuronal growth and less basal cell death in Neurobasal A + B27. Media formulation also affected assay sensitivity and selectivity. Increases in assay sensitivity were observed in Neurobasal A + B27 media as compared to serum-supplemented DMEM. In some instances, a greater difference between effective concentrations for cell death and neurodevelopmental-specific endpoints was also observed in Neurobasal A + B27 media as compared to serum-supplemented DMEM. These data show that media formulation must be considered when comparing data for similar endpoints between studies. Neuronal culture maintained in Neurobasal A + B27 media had several features advantageous for HCI applications including less basal cell death, less cell clustering and neurite fasciculation, and a tendency towards increased sensitivity and selectivity in chemical concentration–response studies.

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Acknowledgments

This manuscript has been reviewed by the National Health and Environmental Effects Research Laboratory, U.S. Environmental Protection Agency, and approved for publication. Approval does not signify that the contents reflect the views of the agency, nor does mention of trade names or commercial products constitute endorsement or recommendation for use.

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  1. Joshua A. Harrill

    Present address: Center for Toxicology and Environmental Health (CTEH), LLC, 5120 North Shore Drive, North Little Rock, AR, 72118, USA

Authors and Affiliations

  1. Integrated Systems Toxicology Division, National Health and Environmental Effects Research Laboratory (NHEERL), United States Environmental Protection Agency, 109 TW Alexander Drive, Research Triangle Park, NC, 27711, USA

    Joshua A. Harrill, Brian L. Robinette, Theresa M. Freudenrich & William R. Mundy

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  1. Joshua A. Harrill
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  2. Brian L. Robinette
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  3. Theresa M. Freudenrich
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Correspondence to William R. Mundy.

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Editor: T. Okamoto

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Harrill, J.A., Robinette, B.L., Freudenrich, T.M. et al. Media formulation influences chemical effects on neuronal growth and morphology. In Vitro Cell.Dev.Biol.-Animal 51, 612–629 (2015). https://doi.org/10.1007/s11626-015-9873-3

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  • DOI: https://doi.org/10.1007/s11626-015-9873-3

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