A Preliminary Spatial Transcriptomics Map of Renin-Angiotensin System Genes in the Mouse Nucleus Ambiguus

The brain-specific renin-angiotensin system (RAS) regulates blood pressure (BP) independently of the systemic RAS. For decades, the source of renin, the rate-limiting enzyme for this cascade within the brain was unclear. Recently, we utilized in situ hybridization to provide definitive evidence for renin within the brainstem, identifying renin-expressing cholinergic neurons within the Nucleus Ambiguus (NuAm). Here we sought to investigate the effect of a hypertensive stimulus on gene expression in the NuAm. We hypothesized that an elevation in BP would decrease the expression of pro-hypertensive RAS elements within the NuAm in a sex-independent manner. To test this hypothesis, we utilized an established protocol for hypertension with low systemic renin circulation. Male and female C57BL6/J mice were randomly assigned to either a control or deoxycorticosterone acetate (DOCA)-salt group. After four weeks brain tissues were fixed, sectioned, and stained to localize the NuAm. A spatial transcriptomic map of the NuAm was generated using Xenium Ranger technology and a set of 347 gene probes. Angiotensinogen expression increased by 0.4-fold in males and 0.9-fold in females in response to DOCA. Some RAS components demonstrated a sex-specific response to DOCA. Renin expression doubled in response to DOCA in males, but was halved in females. Agtr1a, which codes for the angiotensin II type 1 receptor, was largely unchanged in males, but was decreased by 50% in females. Even still, some RAS components, such as angiotensin converting enzyme ( Ace ), remained unaffected by DOCA treatment regardless of sex. Notably, in female mice, DOCA resulted in an approximately 4-fold increase in Agtr2 and Ace2, components of the anti-hypertensive axis of the RAS, suggesting compensatory mechanisms. Unbiased differential gene expression analysis of cholinergic NuAm neurons found that compared to sex-matched shams, DOCA treated males significantly down-regulated just one gene: Sncg. In contrast, DOCA treated females down-regulated Sncg, Calca, and Meis2 while up-regulating Adamts2, Bche, and Npy2r. Additionally, DOCA treatment differentially affected males and females, with males showing down-regulation of Gfap, Wfs1, Grk5, Calca, Syt2, and Chodl, and upregulation of Rgs2, Vat1l, and Rprm. Contrary to our hypothesis, this preliminary analysis suggests that brain RAS regulation may involve complex, sex-dependent regulatory processes. The identification of differentially expressed genes provides a starting point to elucidate a molecular mechanism underpinning these responses. Further analysis will refine our understanding of cell-cell interactions and the spatial organization of renin-expressing neurons in various physiological and pathological states. HL153101 from NIH, 23CDA1048244 from AHA, 5520639 from AHW Michael Keelan Award from the MCW Cardiovascular Center 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.