posted on 2023-02-17, 13:06authored byMolly
J. Kirk, Arya Gold, Ashvin Ravi, Gabriella R. Sterne, Kristin Scott, Evan W. Miller
Visualizing neuronal anatomy often requires labor-intensive
immunohistochemistry
on fixed and dissected brains. To facilitate rapid anatomical staining
in live brains, we used genetically targeted membrane tethers that
covalently link fluorescent dyes for in vivo neuronal labeling. We
generated a series of extracellularly trafficked small-molecule tethering
proteins, HaloTag-CD4 (Kirk et al. Front. Neurosci.2021, 15, 754027) and SNAPf-CD4, which directly label transgene-expressing cells with commercially
available ligand-substituted fluorescent dyes. We created stable transgenic Drosophila reporter lines, which express extracellular HaloTag-CD4
and SNAPf-CD4 with LexA and Gal4 drivers. Expressing these
enzymes in live Drosophila brains, we labeled the
expression patterns of various Gal4 driver lines recapitulating histological
staining in live-brain tissues. Pan-neural expression of SNAPf-CD4 enabled the registration of live brains to an existing
template for anatomical comparisons. We predict that these extracellular
platforms will not only become a valuable complement to existing anatomical
methods but will also prove useful for future genetic targeting of
other small-molecule probes, drugs, and actuators.