Comparison of VSVΔG and VSV(M51R)ΔG for TRAS labeling. (A) Schematic of TRAS labeling. G-deleted VSV (VSVΔG) and VSV-G pseudotyped lentivirus were co-injected into the temporal retina. This results in VSV infection and reporter gene expression in the RGCs in the retina and subsequent transsynaptic spread to retinorecipient cells in the contralateral brain (green circles). The red boxed area is shown in panels (B,C). OT: optic tectum. AF10: RGC arborization field (AF) 10. (B) TRAS labeling with wild-type VSVΔG-GFP. The same animals were imaged at 2 and 6 days post-infection (dpi) [5 and 9 days post fertilization (dpf)], as indicated. Dorsal views are shown in the top row, and lateral views are shown in the bottom row. The number of VSVΔG infected cells (green cell bodies) was significantly diminished between 2 and 6 dpi. Labeling of RGC arbors within the optic tectum (fibers within the dashed outline) was also significantly reduced. (C) TRAS-M51R labeling with VSV(M51R)ΔG-GFP. The same animals were imaged at 2 and 6 dpi and shown in dorsal and lateral views, as indicated. The number of infected cells and the labeling of RGC arbors were similar between 2 and 6 dpi. (D,E) Estimation plots for changes in retinorecipient cell number between 2 and 6 dpi for TRAS and TRAS-M51R. The left part of the graph side shows the cell number for each fish at 2 and 6 dpi. Dashed lines indicate the means of each time point. The right side of the graph shows the effect size (mean of differences between 2 and 6 dpi). Error bars show the mean and 95% confidence interval (CI). The CI in (D) and (E) do not overlap with 0, indicating a significant decrease in cell number. p values from the two-tailed paired t-test are shown on graphs. Scale bars are 100 μm.
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