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. 2016 Aug 17;42(3):661–670. doi: 10.1038/npp.2016.131

Figure 1.

Figure 1

Viral vector-mediated sequestration of soluble tumor necrosis factor (TNF) in the ventrolateral periaqueductal gray (vlPAG) prevents the development of tolerance to systemic morphine and to lipopolysaccharide (LPS) microinfusion into the vlPAG. (a) Schematic of lenti-GFP and lenti-DN-TNF lentiviral vectors. Both lenti-GFP and lenti-DN-TNF vectors contain a chicken β-actin cytomegalovirus enhancer/promoter (CAG), β-globin intron, a central polypurine tract of HIV-1 (ppt), a woodchuck hepatitis virus post-transcriptional response element (WPRE), and a self-inactivating deletion in the 3′-long terminal repeat (3′-LTR). The lenti-DN-TNF vector contains an internal ribosome entry site for GFP expression following the sequence for pro-human DN-TNF (A145R/I97T). (b) Paw withdrawal latency (s) during nociceptive testing on day 1 (D1) and day 3 (D3) at baseline and 15 min postsaline or morphine injection in groups pretreated with lenti-GFP or lenti-DN-TNF; Lenti-GFP+Saline (n=4; black circles), lenti-DN-TNF+Saline (n=5; black squares) lenti-GFP+Morphine (n=10; red circles), and lenti-DN-TNF+Morphine (n=10; red squares). (c) Representative locations of lenti-GFP (circles) and lenti-DN-TNF (squares) microinfusions in the caudal vlPAG (bregma −8.03), and representative light and fluorescein isothiocyanate (FITC) images of injection location and GFP expression from the same section. Dotted lines delineate cerebral aqueduct. (d) Paw withdrawal latency (s) during cumulative morphine injections on day 4 to assess for tolerance (see Supplementary Table 1 for ED50s). (e) Representative locations of lenti-GFP (circles) and lenti-DN-TNF (squares) microinfusions in the vlPAG (bregma −8.04), and representative FITC photomicrograph of GFP expression in the vlPAG. Aq indicates the location of the aqueduct. (f) Paw withdrawal latency (s) during cumulative morphine injections in rats pretreated with lenti-GFP+PAG Saline (n=4; black circles), lenti-GFP+PAG LPS (n=4; red circles), lenti-DN-TNF+PAG Saline (n=6; black squares), and lenti-DN-TNF+PAG LPS (n=9; red squares). *p<0.0001. DN-TNF, dominant-negative TNF; ED50, effective dose 50% GFP, green fluorescent protein. A full color version of this figure is available at the Neuropsychopharmacology journal online.