Extended Data Fig. 7: Fiber photometry recording of PVN parvOT neurons and SON magnOT neurons with chemogenetic manipulation of parvOT neurons.

a-b, Schema of viral vectors injected (CAV2-Cre in the SON and OTp-DIO-GCaMP6s in the PVN) and implanted optic fiber for fiber photometry recording of PVN parvOT neurons (a). No Ca²⁺ transient nor changes in Ca²⁺ signal upon ‘airpuff’ stimulation were detected when recording parvOT neurons exclusively (b, solid line: average, shaded area: SEM, n = 3 rats). c-d, Schema of viral vectors injected and implanted optic fiber for fiber photometry recording of SON magnOT neurons (c). Examples of fiber photometry-based Ca²⁺ signals of SON magnOT population during airpuff stimulation (d); the graphic is an average of 33 airpuff responses (11 airpuffs per animal, n = 3 rats). e, Relative change in the area under the curve (AUC) 0-30 s after airpuffs with respect to 30 s before the stimuli, of SON magnOT vs. parvOT neurons (solid line: average, shaded area: SEM, n = 33 airpuffs from 3 rats, ** P = 0.008, unpaired two-sided t test). Data show mean + SEM. f-g-h, Schema of viral vectors injected and implanted optic fiber for fiber photometry recording (f) of SON OT neurons with concomitant DREADD-Gq activation of parvOT neurons. Normalized area under the curve (AUC) of GCaMP6s signal (g, solid line: average, shaded area: SEM, 1 min bin size) of SON OT neurons showing increase of cells activities after parvOT activation mediated by CNO i.p. injection (indicated by arrow). 30-min averaged AUC (h) showing a gradual increase in cellular activity and lasting at least 120 min (*P = 0.0406, *P = 0.0274, *P = 0.0107, *P = 0.0301, n = 3 rats, two-way ANOVA Tukey’s corrected post-hoc comparison). Data show mean + SEM. i-j-k, Schema of viral vectors injected and implanted optic fiber for fiber photometry recording (i) of SON OT neurons with concomitant DREADD-Gi inhibition of parvOT neurons. Normalized area under the curve (AUC) of GCaMP6s signal (j, solid line: average, shaded area: SEM, 1 min bin size) of PVN OT neurons showing a decrease of cellular activity after parvOT inhibition mediated by i.p. CNO injection (indicated by arrow). 30-min averaged AUC (k) shows a sharp decrease in cellular activity that lasts at least 60 min and then gradually recovers (***P = 0.0008, **P = 0.0051, *P = 0.0140, *P = 0.0168, n = 3 rats, two-way ANOVA Tukey’s corrected post-hoc comparison). Data show mean + SEM. l-m-n, Schema of viral vectors injected and implanted optic fiber for fiber photometry recording (l) of SON OT neurons in control animals (DREADD-free) expressing GFP in parvOT neurons. Normalized area under the curve (AUC) of GCaMP6s signal (m, solid line: average, shaded area: SEM, 1 min bin size) of PVN OT neurons showing no significant changes in Ca²⁺ signal upon CNO injection. No significant changes are detected in 30-min averaged AUC (n) up to 120 min (P = 0.5715, P = 0.2429, P = 0.4673, P = 0.2848, n = 2 rats, two-way ANOVA Tukey’s corrected post-hoc comparison). Data show mean values. o, Panels of an immunostained section of the SON (one of out of eight independent experiments) showing post-hoc verification of implanted optic fiber above the SON and co-localization of immunoreactive, GCaMP6s (green), oxytocin (blue), and merged channels. Scale bar 100 μm and 10 μm (inset).