Extended Data Fig. 1: Behavioural paradigms for sodium-appetite induction and histological analysis of the pre-LC.

a, Experimental protocols for inducing thirst and sodium appetite. Intraperitoneal injection of furosemide (FURO) (50 mg per kg body weight) was used to induce sodium appetite. b, Sodium-depleted animals showed a strong preference for sodium whereas water-deprived animals preferred water over sodium (n = 9). c, Water-deprivation for 48 h induced robust c-FOS expression in the subfornical organ. However, it did not activate the pre-LC (representative image for one out of four mice that were tested). d, FISH showing that PDYN–Cre expression (visualized in the Ai3 transgenic line, green) overlaps with endogenous PDYN transcripts in the pre-LC (red, one out of two mice). e, Pre-LC^PDYN neurons also overlap with FOXP2 expression, which is a known marker in the pre-LC^43,44 (93.8 ± 1.1% (mean ±s.e.m.); n = 3). Scale bars, 50 μm. **P < 0.01 by two-tailed Wilcoxon test. Data are mean ± s.e.m.