Fig. 2: Effects of chronic social stress on BA-NAc neural activity during reward-directed behaviour.

a Schematic showing unilateral injection site of FLEX Cre-On GCaMP6 AAV vector in BA, fibre-optic probe implantation dorsal to the injection site, and ipsilateral injection of retrograde Cre AAV vector in NAc (relative to bregma). b Experimental design. BBW + FC: measurement of baseline body weight and food consumption; 90–95% BBW: conditioning under food restriction that reduced BW to 90–95% BBW; S BA + NAc: stereotactic surgery; Recovery + AAVV: recovery from surgery and AAV vector expression; C + OF: Conditioning sessions with patch cord attached to optic fibre; CSS/CON: CSS protocol or control handling; re-BBW + FC: BW and food consumption under ad libitum feeding on days 1–12 of CSS/CON provided re-baseline values; 95–100% re-BBW: mice were mildly food restricted to be tested at 95–100% re-BBW; SIG: fibre photometry signal test; BP: brains were perfused for histology. c–h DRLM test with fibre photometry. Mice received 3 daily tests of 31 trials each and trials 1–20 per test were used for data analysis. Data are shown as group mean ± SEM and per mouse scores. Behaviour: c Number of chocolate pellets obtained, i.e. DS trials with a response. d Median DS response latency. e Median ITI response interval. f Median learning ratio. Statistical analysis was conducted using 2-way mixed-model ANOVA. BA-NAc neural activity (z-scored Ca²⁺ signal) relative to baseline, in tests 1 and 3: g For trials with a DS response, Ca²⁺ activity during the DS-on phase (time-normalized and subdivided into 10 equal intervals) from DS-onset to feeder response for trials 1–20 (left) and trials 11–20 (centre), and non-time normalized mean values (right). h For trials with a DS response, Ca²⁺ activity during the post-DS feeder phase which started with a feeder response and had a duration of 5 s (subdivided into 0.5 s intervals), for trials 1–20 (left) and trials 11–20 (right). Statistical analysis was conducted using 3-way mixed-model ANOVA. Test days and intervals indicated by different letters were significantly different in Tukey’s multiple comparisons tests: e.g. a vs. b, a vs. c, b vs. c, p < 0.05 or lower. i–p REV test with fibre photometry. Mice received 3 daily tests and the mean of data for tests 2 and 3 are given and were used for statistical analysis. Data are shown as individual values or group mean ± SEM. Behaviour: i Number of chocolate pellets earned. j Final ratio attained. k At progressive ratio 5 (PR 5), operant phase duration i.e. time from 1st until 5th nosepoke. l At PR 5, post-operant phase duration i.e. time from DS onset until feeder response. Statistical analysis was conducted using t-tests. BA-NAc neuron population activity (z-scored Ca²⁺ signal) relative to baseline: m In CON mice specifically, comparison of Ca²⁺ activity during the post-operant phase—time from DS onset to feeder response (time-normalized and subdivided into 10 equal intervals)—at PRs 5, 9 and 13. n Ca²⁺ activity during the operant phase at PR 5. o Ca²⁺ activity during the post-operant phase at PR 5. p Ca²⁺ activity during the post-feeder phase at PR 5. Statistical analysis was conducted using 2-way mixed-model ANOVA. Test days and intervals indicated by different letters were significantly different in Tukey’s multiple comparisons test: e.g. a vs. b, a vs. c, b vs. c, p < 0.05 or lower. Images a, DRLM test and REV test were created with BioRender.com.