Table 4.
Injection parameters of H129-dTK-tdT monosynaptic tracer
Animal | Brain region | Helper AAV | H129-dTK-tdT | Perfusion time (day)b | |||||
---|---|---|---|---|---|---|---|---|---|
AAVs (1 × 1012 vg/mL) | Vol.a (μL) | Inject. time | Dose (pfu) | Vol.a (μL) | Inject. time | For starter neuron | For postsynaptic neuron | ||
C57BL/6 | MOB [16] | AAV2/9-TK-GFP | 0.45 | Day 1 | 2.00 × 105 | 0.40 | Day 22 | Day 25 | Day 29 |
C57BL/6 | αBLA [22] | AAV2/9-TK-GFP | 0.20 | Day 1 | 5.00 × 104 | 0.10 | Day 22 | Day 25 | Day 27 |
C57BL/6 | VPM [16] | AAV2/9-TK-GFP | 0.20 | Day 1 | 1.00 × 105 | 0.20 | Day 22 | Day 25 | Day 32 |
C57BL/6 | LGN [16] | AAV2/9-TK-GFP | 0.20 | Day 1 | 1.00 × 105 | 0.20 | Day 22 | Day 25 | Day 32 |
C57BL/6 | pBLA [22] | AAV2/9-TK-GFP | 0.20 | Day 1 | 5.00 × 104 | 0.10 | Day 22 | Day 25 | Day 27 |
PV-Cre | nRT [16] | AAV2/9-DIO-TK-GFP | 0.35 | Day 1 | 1.25 × 105 | 0.25 | Day 22 | Day 25 | Day 32 |
DAT-Cre | VTA [16] | AAV2/9-DIO-TK-GFP | 0.35 | Day 1 | 1.25 × 105 | 0.25 | Day 22 | Day 25 | Day 32 |
aInjected volume should not exceed 0.5 μL for brain regions.
bExtending the waiting time before perfusion normally results in higher labeling intensity and more labeled postsynaptic neurons. However, the starter neurons are also more severely damaged due to H129-dTK-tdT toxicity when conditional replication occurs.