Fig. 6: Impact of 24-hour social isolation on Insula^Ins neuronal activity.

a Top. Experimental schematic. Bottom. Representative traces of Insula^Ins neurons in response to depolarizing current pulses (50 pA, 90 pA) in current clamp in group-housed and 24-hour isolated. b Representative example of a histological control showing Insula^Ins neurons (tomato) and Insula recorded neurons filled with biocytin (gray) and the overlay. c–e Group mean of the rheobase (c, Mann–Whitney test, U = 183.5, p = 0.045), resting membrane potential (d, two-tailed unpaired t-test, t(46) = 2.549, p = 0.0142), and resistance (e, Mann–Whitney test, U = 190, p = 0.0437) of Insula^Ins neurons in group-housed and 24-hour isolated mice (group-housed condition: 23 neurons in 3 mice; isolated conditions: 24 neurons in 3 mice). f, h Experimental schematic. g Representative epifluorescent image of a coronal brain slice injected with a rAAV2-Cre retrograde virus in the contralateral Insula, coupled with an AAV9-syn-flex-jCCaMP8f-WPRE virus injection in the ipsilateral Insula, along with the optic fiber placement. i, j Time spent around the juvenile social or the object enclosure in group-housed condition (i, two-tailed unpaired t-test, t(6) = 4.682, p = 0.0034) and after 24 h of social isolation (j, two-tailed unpaired t-test, t(6) = 5.321, p = 0.0018, white circle for female, black circle for male). k–m Example of fiber photometry recording in Insula^Ins neurons during three-chamber social interaction test in GFP control group (k, n), in GCaMP8f group-housed (l, o) and 24 h isolated mice (m, p). Blue and orange squares represent the episodes of interactions with the juvenile mouse or the object, respectively. Black arrows are the detected calcium events. q Quantification of ∆F/F mean fluorescence z-score of Insula^Ins neurons around social or object enclosure in both groups (N = 7 mice per group; two Way repeated measure Anova, Interaction F(1,6) = 8.89, p = 0.0246, main effect of housing F(1,6) = 16.17, p = 0.007, no main effect of zone F(1,6) = 2.428, p = 0.1702, followed by uncorrected Fisher’s LSD post hoc, group-housed social vs group-housed object p > 0.05, 24 h isolated social vs object p = 0.0073, group-housed social vs 24 h isolated social p > 0.05, group-housed object vs 24 h isolated object p = 0.012). dark blue/orange color circle: female, light blue/orange color: male. Data are presented as mean values ± SEM. *p < 0.05, **p < 0.01.