Fig. 1
From: Inhibitory insula-ACC projections modulate affective but not sensory aspects of neuropathic pain
Insula-pACC PV+ projections modulate CPP responses without mechanical threshold effects. a Confocal microscopy images from pIC slices from SHAM and SNI operated PV Cre:tdTomato mice subjected to CTB488 (green) injection in the pACC (PVCre n = 7) followed by SNI or SHAM procedures a week later. Slices were immunostained for c-Fos activity (shown in blue). White arrows highlight examples of cells in which CTB488 and tdTomato label overlap, circles highlight projecting cells that are positive for c-Fos. Dotted squares show only PV+ cells. b Verification of injection of CTB488 injection sites in the pAAC region. c Quantification of the percentage overlap between tdTomato positive cells that are also positive for CTB488. d Percentage overlap between pIC-pACC projecting GABAergic cells (i.e., cells that are both red and green) and those positive for c-Fos (blue). Data are presented as mean ± S.E.M. e Diagram showing that AAV (AAV9-EF1a-DIO-ChR2-eYFP) was injected in the pIC of PV Cre transgenic mice. f Voltage clamp recording in brain slices of the pACC. Left: percentage of cells that responded with oIPSCs (12.7%, upper) and with oEPSCs (0%, lower). Right: sample traces showing responses at − 70 mV (no oEPSC) and at 0 mV (with oIPSC) and that TTX blocked the oIPSC response. g Location of recording sites in pACC slices. Each recorded neuron in the pACC was mapped onto a brain section of pACC from a mouse brain atlas showing the location of responding neurons (responder, solid dots) and non-responding neurons (non-responder, open dots). h Amplitude and latency of oIPSCs from responding neurons. Error bars are S.E.M. i Mechanical withdrawal threshold of SHAM and SNI operated PVCre mice with and without optoactivation of ChR2 (Light and No Light-473 nm, pulsed light, 5mW, 40 Hz) and mechanical threshold was accessed using a DPA test with the animal under stimulation. Two-way ANOVA followed by Bonferroni post-test. ***p = 0.0001. j Optogenetic stimulation of the GABAergic pIC to pACC pathway was able to induce place preference in SNI mice (PVCre SNI (n = 5) or SHAM (n = 4 to 5)). The paradigm consists of one day conditioning, with lights off for 15 min in the morning in one chamber and opto-light stimulation for 15 min in the afternoon in the opposite chamber. Two-way ANOVA followed by Bonferroni post-test, *p = 0.0331 SHAM vs SNI Light paired chamber; **p = 0.0082 Pre (pre-test) vs SNI Light paired chamber; ####p < 0.0001 Light vs No Light SNI group. k CPP score (difference of time spent in the conditioning chamber during post- and pre-conditioning); One-way ANOVA followed by Bonferroni post-test, *p = 0.018 SNI No Lght vs Light. All data are presented as mean ± S.E.M