Fig. 4

The vlPAG is an anatomical upstream of WIM. a Retrograde tracing strategy of tracing the upstream of WIM through the transsynaptic retrograde PRV-EGFP vector injection into the unilateral WIM of mice with transected infraorbital branch of the maxillary nerve of the trigeminal ganglion. b The primary order of WIM, which was labelled by PRV-EGFP green signals was located in the lateral subnucleus of FN (lFN) 48–60 h after the virus injection. Scale bar, 500 μm. c Representative images of one side of the rostral to caudal vlPAG showed PRV-EGFP-labelled neurons 72 h later of PRV-EGFP infections (n = 3 mice per group). Scale bar, 500 μm. d Representative image of PRV-EGFP infectious neurons in the unilateral vlPAG that were co-labelled with CaMKII-positive neurons. Scale bar, 200 μm (left one); 50 μm (right four). e Percentage of total PRV-EGFP infectious neurons expressing CaMKII in the ipsilateral vlPAG of injection side (n = 7 brain sections from 3 mice). f Scheme of verifying vlPAG descending output fibres onto the ipsilateral wFMNs upon a monosynaptic anterograde virus vector infused into the unilateral vlPAG. g Typical image of unilateral vlPAG with starter cells labelled with AAV-hSyn-mGFP-2A-Synaptophysin-mRuby (yellow). Scale bar, 200 μm. h Representative image of Synaptophysin-mRuby-labelled synaptic fibres and terminals innervating ChAT positive neurons in the ipsilateral lFN. The right image was the magnified view of the left one. Scale bar, 500 μm (left); 50 μm (right). Aq, aqueduct; TG, trigeminal ganglion; lPAG, lateral PAG