T Cell Calcium Dynamics Visualized in a Ratiometric tdTomato-GCaMP6f Transgenic Reporter Mouse
Calcium is an essential cellular messenger that regulates numerous functions in living organisms. Here we describe development and characterization of \"Salsa6f\", a fusion of GCaMP6f and tdTomato optimized for cell tracking while monitoring cytosolic Ca2+, and a transgenic Ca2+ reporter mouse with Salsa6f floxed and targeted to the Rosa26 locus for expression in specific cell types. Using CD4-Cre-Salsa6f mice, we report normal development and function of T cells expressing Salsa6f and demonstrate Ca2+ signaling dynamics during T cell receptor engagement in naive T cells, helper Th17 T cells and regulatory T cells. Salsa6f expression also revealed functional expression of mechanosensitive Piezo1 channels in T cells. Transgenic expression of Salsa6f enables ratiometric imaging of Ca2+ signals in complex tissue environments found in vivo. Deep tissue two-photon imaging of T cells in the steady-state lymph node revealed a highly localized Ca2+ signaling behavior (\"sparkles\") as cells migrate.