Identification of TPBG-Expressing Amacrine Cells in DAT-tdTomato Mouse

PURPOSE: Neurons are the bricks of the neuronal system and experimental access to certain neuron subtypes will be of great help to decipher neuronal circuits. Here, we identified trophoblast glycoprotein (TPBG)-expressing GABAergic amacrine cells (ACs) that were selectively labeled in DAT-tdTomato transgenic mice. METHODS: Retina and brain sections were prepared for immunostaining with antibodies against various biomarkers. Patch-sequencing was performed to obtain the transcriptomes of tdTomato-positive cells in DAT-tdTomato mice. Whole-cell recordings were conducted to identify responses to light stimulation. RESULTS: Tyrosine hydroxylase immunoreactive cells were colocalized with tdTomato-positive cells in substantia nigra pars compacta, but not in the retina. Transcriptomes collected from tdTomato-positive cells in retinas via Patch-sequencing exhibited the expression of marker genes of ACs (Pax6 and Slc32a1) and marker genes of GABAergic neurons (Gad1, Gad2, and Slc6a1). Immunostaining with antibodies against relevant proteins (GAD67, GAD65, and GABA) also confirmed transcriptomic results. Furthermore, tdTomato-positive cells in retinas selectively expressed Tpbg, a marker gene for distinct clusters molecularly defined, which was proved with TPBG immunoreactivity in fluorescently labeled cells. Finally, tdTomato-positive cells recorded showed ON–OFF responses to light stimulation. CONCLUSIONS: Ectopic expression occurs in the retina but not in the substantia nigra pars compacta in the DAT-tdTomato mouse, and fluorescently labeled cells in the retina are TPBG-expressing GABAergic ACs. This type of transgenic mice has been proved as an ideal tool to achieve efficient labeling of a distinct subset of ACs that selectively express Tpbg.