Characterizing a novelvGlut3-P2A-iCreERknockin mouse strain in cochlea

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Precise mouse genetic studies rely on specific tools that can label specific cell types. In mouse cochlea, previous studies suggest that vesicular glutamate transporter 3 (vGlut3), also known as Slc17a8, is specifically expressed in inner hair cells (IHCs) and loss of vGlut3 causes deafness. To take advantage of its unique expression pattern, here we generate a novel vGlut3-P2A-iCreER knockin mouse strain. The P2A-iCreER cassette is precisely inserted before stop codon of vGlut3, by which the endogenous vGlut3 is intact and paired with iCreER as well. Approximately, 10.7%, 85.6% and 41.8% of IHCs are tdtomato + when tamoxifen is given to vGlut3-P2A-iCreER/+; Rosa26-LSL-tdtomato/+ reporter strain at P2/P3, P10/P11 and P30/P31, respectively. Tdtomato + OHCs are never observed. Interestingly, besides IHCs, glia cells, but not spiral ganglion neurons (SGNs), are tdtomato+, which is further evidenced by the presence of Sox10+/tdtomato+ and tdtomato+/Prox1(Gata3 or Tuj1)-negative cells in SGN region. We further independently validate vGlut3 expression in SGN region by vGlut3 in situ hybridization and antibody staining. Moreover, total number of tdtomato + glia cells decreased gradually when tamoxifen is given from P2/P3 to P30/P31. Taken together, vGlut3-P2A-iCreER is an efficient genetic tool to specifically target IHCs for gene manipulation, which is complimentary to Prestin-CreER strain exclusively labelling cochlear outer hair cells (OHCs).HighlightsvGlut3-iCreER specifically labels cochlear inner hair cells, but not outer hair cells at all postnatal ages.vGlut3-iCreER also efficiently targets cochlear glia cells, especially at neonatal ages.Cochlear spiral ganglion neurons do not express vGlut3.vGlut3 is weakly and transiently expressed in cochlear outer hair cells.

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