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Media formulation influences chemical effects on neuronal growth and morphology
Citation:
Harrill, J., B. Robinette, T. Freudenrich, AND W. Mundy. Media formulation influences chemical effects on neuronal growth and morphology. IN VITRO CELLULAR & DEVELOPMENTAL BIOLOGY - ANIMAL. Society for In Vitro Biology-Animal, 51(6):612-29, (2015).
Impact/Purpose:
In vitro test methods can provide a rapid approach for the screening of large numbers of chemicals for their potential to produce toxicity (hazard identification), including DNT. Such screening could facilitate prioritization of resources needed for in vivo toxicity testing towards those chemicals most likely to result in adverse health effects. Cell cultures derived from nervous system tissue can be used in automated, high content imaging (HCI) assays to examine the chemical effects on neurite outgrowth. In order to determine whether HCI-based DNT assays can be affected by differences in media formulation, a systematic comparison of serum-supplemented (DMEM + 10 % serum) and serum-free (Neurobasal A + B27) culture media on neuronal growth morphology was performed.
Description:
Abstract Screening for developmental neurotoxicity (DNT) using in vitro, cell-based test systems has been proposed as an efficient and cost-effective alternative to performing in vivo DNT studies. One of the principle tools currently used for DNT screening is automated high-content imaging (HCI) of neuronal morphology. While 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, including culture media formulation. In order to determine whether HCI-based DNT assays can be affected by differences in media formulation, a systematic comparison of serum-supplemented (DMEM + 10 % serum) and serum-free (Neurobasal A + B27) culture media on neuronal growth morphology was performed. Primary rat cortical neurons were used as the test system. Concentration-response assays for neuritogenesis, axon and dendrite outgrowth and synaptogenesis were performed in each media type using a chemical test set. 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 formulaion 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 DNT 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 cultures maintained Neurobasal A + B27 media had several features advantageous for HCI applications including less basal cell death, less cell clustering and neurite fasciculation as well as a tendency towards increased sensitivity and selectivity in chemical concentration-reponse studies.
