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Quantitative Analysis of Supporting Cell Subtype Labeling Among CreER Lines in the Neonatal Mouse Cochlea

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ABSTRACT

Four CreER lines that are commonly used in the auditory field to label cochlear supporting cells (SCs) are expressed in multiple SC subtypes, with some lines also showing reporter expression in hair cells (HCs). We hypothesized that altering the tamoxifen dose would modify CreER expression and target subsets of SCs. We also used two different reporter lines, ROSA26 tdTomato and CAG-eGFP, to achieve the same goal. Our results confirm previous reports that Sox2 CreERT2 and Fgfr3-iCreER T2 are not only expressed in neonatal SCs but also in HCs. Decreasing the tamoxifen dose did not reduce HC expression for Sox2 CreERT2, but changing to the CAG-eGFP reporter decreased reporter-positive HCs sevenfold. However, there was also a significant decrease in the number of reporter-positive SCs. In contrast, there was a large reduction in reporter-positive HCs in Fgfr3-iCreER T2 mice with the lowest tamoxifen dose tested yet only limited reduction in SC labeling. The targeting of reporter expression to inner phalangeal and border cells was increased when Plp-CreER T2 was paired with the CAG-eGFP reporter; however, the total number of labeled cells decreased. Changes to the tamoxifen dose or reporter line with Prox1 CreERT2 caused minimal changes. Our data demonstrate that modifications to the tamoxifen dose or the use of different reporter lines may be successful in narrowing the numbers and/or types of cells labeled, but each CreER line responded differently. When the ROSA26 tdTomato reporter was combined with any of the four CreER lines, there was no difference in the number of tdTomato-positive cells after one or two injections of tamoxifen given at birth. Thus, tamoxifen-mediated toxicity could be reduced by only giving one injection. While the CAG-eGFP reporter consistently labeled fewer cells, both reporter lines are valuable depending on the goal of the study.

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Acknowledgments

We thank other members of the Cox lab for discussion. We thank Dr. William Richardson (University College London) for providing Fgfr3-iCreER T2 mice; Dr. Guillermo Oliver (St. Jude Children’s Research Hospital) for providing Prox1 CreERT2 mice; and Dr. Jeffery Robbins (Cincinnati Children’s Hospital) for providing CAG-eGFP mice. This work was supported by a grant from the Office of Naval Research (N000141310569). Southern Illinois University School of Medicine Research Imaging facility equipment was supported by Award Number S10RR027716 from the National Center for Research Resources-Health.

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  1. Department of Pharmacology, Southern Illinois University School of Medicine, Springfield, IL, 62711, USA

    Melissa M. McGovern, Joseph Brancheck, Auston C. Grant, Kaley A. Graves & Brandon C. Cox

  2. Department of Surgery, Division of Otolaryngology, Southern Illinois University School of Medicine, Springfield, IL, 62711, USA

    Brandon C. Cox

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  1. Melissa M. McGovern
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  2. Joseph Brancheck
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Correspondence to Brandon C. Cox.

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McGovern, M.M., Brancheck, J., Grant, A.C. et al. Quantitative Analysis of Supporting Cell Subtype Labeling Among CreER Lines in the Neonatal Mouse Cochlea. JARO 18, 227–245 (2017). https://doi.org/10.1007/s10162-016-0598-0

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