Figure 3. Assessment of intermediate BA-NAc neuron population activity during social reward-fibre photometry testing.
(A) Experimental design. Handling; Surgery: stereotactic surgery for AAV vector injections for intermediate BA-NAc neuron GcaMP6 expression (or EGFP expression as control) and implantation of optic fibre in intermediate BA; rAAV + Recovery: post-operative care, AAV vector expression, habituation to photometry system; Stimulus: Two daily tests with social reward (SR) combined with fibre photometry recording; Ex-vivo: brain perfusion-fixation for target validation. (B) Schematic of social reward-fibre photometry test phases: each session comprised the phases, each of 20 min, no stimulus, distal-SR, proximal-SR, with BA-NAc neuron activity (n=10) or BA-NAc neuron EGFP signal (n=4) recorded in each phase. For the phases, distal-SR and proximal-SR (yellow background), fibre photometry data were analyzed in terms of Ca2+ activity (or EGFP signal) associated with social events. (C) Representative microscope image of a Nissl-stained coronal brain sections showing location of the optic fibre implant including its tip in the BA at bregma -1.8 mm, and colocalized AAV vector-expressed GcaMP6 fluorescence. Right image: BA at high magnification with estimated coordinates of optic fibre tip placement (mean ± SD, n=10). (D, E) Peri-event BA-NAc neuron Ca2+ activity during (D) distal-SR and (E) proximal-SR. Left: Representative traces from individual mice showing 150 s of z-scored Ca2+activity and concomitant social events, the latter depicted in pink shading. Right: Peri-event histograms of z-scored Ca2+activity relative to social event onset (time = 0 s). The horizontal bars depict social event duration. Data are overall mean ± SEM values, n=10 mice. There was a change in Ca2+ activity after the onset of social episodes during both distal-SR and proximal-SR, with activity increasing at sec 3-4 in distal-SR and at sec 2 in proximal-SR (phase x time interaction effect: F(2.31, 20.77)=4.40, p=0.02, followed by Dunnett’s test); there was no time point at which peri-event Ca2+ activity was significantly different between proximal and distal SR phases (Sidak’s multiple comparisons test).