Fig. 5: Morphine reduces DMV-projecting PVN^Glu neuronal activity to induce constipation.

a Schematic diagram for the Cre-dependent retrograde trans-monosynaptic rabies virus (RV) tracing strategy. b Representative images for EGFP-labelled PVN neurons that co-localized with a MOR-specific antibody. Scale bars, 50 μm (overview) and 5 μm (zoom). c Schematic of microendoscopic calcium imaging in the PVN of freely moving mice. d Representative images showing a filed view from the GRIN lens (left) and the viral expression of GCaMP6m in the PVN (right). Scale bar, 50 μm. e,f Sample traces (e) and summary data (f) for calcium transients of GCaMP6m-expressing PVN neurons (n = 25 cells per group, P = 0.0031). g Schematic diagram for cannula implantation and morphine administration. h Summary data for the fecal weight in mice after administration of morphine into the PVN via cannulas (n = 10, F_1,18 = 16.89, P < 0.0001 for 2 h, P < 0.0001 for 4 h). i, j Sample traces (i), summary data showing the frequency and amplitude values for small intestinal motility ( j) in mice after morphine administration into the PVN (n = 10, P = 0.0021 for frequency, P < 0.0001 for amplitude). k Schematic diagram for cannula implantation and naloxone administration. l Summary data for the fecal weight in mice after administration of naloxone into the PVN via cannulas (n = 10, F_1,18 = 5.612, P = 0.0166 for 4 h). m, n Sample traces (m), summary data showing the frequency and amplitude values for small intestinal motility (n) in mice after naloxone administration into the PVN (n = 10, P < 0.0001 for frequency, P < 0.0001 for amplitude). Significance was assessed by two-tailed paired Student’s t test (f), two-way repeated-measures ANOVA with post hoc comparison between groups (h, l), Mann-Whitney U test and two-tailed unpaired Student’s t test (j, n). The data are presented as the mean ± SEMs. *P < 0.05, **P < 0.01, ***P < 0.001; n.s., not significant. See also Supplementary Table S1. Source data are provided as a Source Data file.