Spatial transcriptomics identifies programmed changes in angiotensin AT₁R-expressing Agouti-related peptide neurons following early-life sodium restriction

Preterm birth is associated with high risk of short-term sodium (Na) depletion due to renal immaturity, and subsequently, with life-long increased risk for cardiometabolic diseases. We hypothesize that Na depletion during specific developmental stages contributes to programmed cardiometabolic risks via altered development of regulatory networks within the hypothalamus. We recently published that Na restriction during translationally relevant developmental stages in mice (e.g., 3-6 weeks of age) programs long-lasting changes in autonomic control of metabolism and blood pressure, and these effects are maintained in adulthood via an angiotensin II type 1 (AT 1 R/ Agtr1a )-dependent mechanism. We also previously discovered a major role for inhibitory AT 1 R-Gai signaling within a subset (“Type 1”) of Agouti-related peptide (AgRP/ Agrp ) neurons in the arcuate nucleus of the hypothalamus in cardiometabolic control. Here we used spatial transcriptomics to examine the hypothesis that early-life Na depletion programs lasting changes in the transcriptomes of Type 1 AgRP neurons. C57BL/6J mice were supplied low (0.04%) or control (0.30%) Na diets from 3-6 weeks of age, then returned to standard (0.15% Na) diet. At 8 weeks of age, mice were euthanized following a 5h fast. Paraffin-embedded coronal sections (5 µm) of the hypothalamus were analyzed using the 10X Genomics Xenium Mouse Brain probe set (247 genes) plus 100 custom add-on genes (n=2 technical replicates each from 1 male + 1 female per diet). Type 1 AgRP neurons were identified as both Agrp + and Agtr1a + , versus other subtypes that were Agrp + but Agtr1a — . In control mice, the AT 1 R + Type 1 AgRP neurons (n=89) expressed more Agrp (median=14 vs 5 transcripts/cell; distribution p<0.01 by two-sample Kolmogorov-Smirnov test), Npy (22 vs 9, p<0.01), and genes involved with γ-aminobutyric acid (GABA) synthesis ( Slc32a1 : 4.5 vs 3, p<0.01; Gad1 : 11.5 vs 7, p<0.01; and Gad2 : 5 vs 3, p<0.01) compared to other subtypes of AgRP neurons (n=385). Within Type 1 cells (n=69), early-life Na restriction programmed large reductions in Agrp (8.5 vs 14, p<0.01) and Npy (16.25 vs 22, p=0.02) but had variable effects on GABA synthetic genes ( Slc32a1 : 3.75 vs 4.5, p=0.49; Gad1 : 8.25 vs 11.5, p=0.01; Gad2 : 4 vs 5, p=0.06). Notably another cAMP-responsive gene, regulator of G protein signaling-2 was reduced ( Rgs2 : 6 vs 10.5, p<0.01), potentially indicating increased inhibitory AT 1 R-Gai signaling in these cells. These findings support the working hypothesis that early-life Na depletion during the developmental window relevant to hypothalamic maturation programs lasting changes in the transcriptome of the AT 1 R + Type 1 AgRP neuron that may contribute to programmed changes in cardiometabolic control. HL134850 DK133121 GM080202 This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.