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[Preprint]. 2024 Jul 11:2024.07.08.602522. [Version 1] doi: 10.1101/2024.07.08.602522

Figure 5: VTAMORs mediate protracted withdrawal-induced low sociability.

Figure 5:

(a) Experimental schematic for VTAMOR re-expression in MOR KO mice through bilateral VTA injections of AAVDJ-hSyn1-FLEX-mCh-T2A-FLAG-hMOR, followed by the protracted morphine withdrawal paradigm and culminating in a morphine challenge injection (20 mg/kg, s.c.). (b) AAV-hSyn-EGFP viral control or AAVDJ-hSyn1-hMOR in the VTA injection site (scale, 200 μm). (c) MOR knockout in the MOR KO mice demonstrated by decreased mouse Oprm1 mRNA expression relative to the line’s littermate controls (n=8 Oprm1+/+ mice/4 MOR KO mice, unpaired t test, t=8.949, df=10, p<0.0001). (d) VTAMOR re-expression in MOR KO mice through increased expression of human OPRM1 mRNA relative to MOR KOs (n=4 MOR KO mice/3 MOR KO mice with VTAMOR bilateral re-expression, unpaired t test, t=8.551, df=5, p=0.0004). (e) Daily morphine sulfate intake across a 13-day forced drinking exposure paradigm. (f) At 24 hours following morphine cessation, neither the AAV-EGFP-injected control or AAV-hMOR-injected treatment group show alterations in physical somatic signs as assessed through global withdrawal scores (n=8 AAV-EGFP-injected + opioid naïve mice/8 AAV-hMOR-injected + opioid naïve mice/8 AAVEGFP-injected + morphine-treated mice/7 AAV-hMOR-injected + morphine-treated mice, two-way ANOVA + Bonferroni, virus F 1,27=1.207, p=0.2817; treatment F1,27=0.04160, p=0.8399; interaction F1,27=0.09251, p=0.7633). (g) Morphine-treated MOR KO mice with VTAMOR re-expression show decreased sociability relative to their opioid naïve counterparts during the 4-week protracted withdrawal period, while the morphine-treated AAV-EGFP-injected viral controls remain unaffected during protracted morphine (two-way ANOVA + Bonferroni, virus F1,27=0.0007256, p=0.9787; treatment F1,27=4.325, p=0.0472; interaction F1,27=4.250, p=0.0490). VTAMOR re-expression in MOR KO mice does not alter protracted morphine withdrawal behaviors related to (h) Elevated zero maze (two-way ANOVA + Bonferroni, virus F1,27=1.727, p=0.1999; treatment F1,27=1.075, p=0.3090; interaction F1,27=0.01890, p=0.8917), (i) Tail suspension (two-way ANOVA + Bonferroni, virus F1,26=0.2659, p=0.6105; treatment F1,26=0.2668, p=0.6098; interaction F1,26=0.9168, p=0.3471), and (j) Nociceptive hot plate test (two-way ANOVA + Bonferroni, virus F1,27=0.3573, p=0.5550; treatment F1,27=0.1854, p=0.6702; interaction F1,27=0.03435, p=0.8544). Data = mean ± SEM, *p<0.05, ***p<0.001, ****p<0.0001.