Orally active exchange protein directly activated by cAMP (Epac) inhibitor produces natriuresis by decreasing NHE-3 expression and brush-border localization in the proximal tubule

In recent years, Epac signaling cascade has emerged as a potent physiologically relevant modulator of water-solute transport in the renal tubule. Mice with deleted Epac1 or Epac2 isoforms exhibit moderate diuresis, urinary Na + wasting, and perturbed acid-base balance due to compromised Na + reabsorption via the Na + /H+ exchanger (NHE-3) and the epithelial Na + channel (ENaC) in the proximal tubule (PT) and the collecting duct (CD), respectively. However, the pharmacological potential of Epac inhibition on renal function remains unexplored. To this end, we combined metabolic cage balance studies in 3-month-old C57Bl/6 mice of both sexes with super-resolution STED fluorescent microscopy and biochemical tools to detect changes in abundance and subcellular localization of the major transport proteins in the PT and CD upon systemic administration of the first-in-class orally active pan-isoform selective Epac blocker ESI-09. ESI-09 was mixed with a standard rodent diet (0.32% Na + ) at 80 mg/kg, which led to steady-state levels in the plasma of approximately 20 µM after 7 days similarly in both males and females, as was assessed with mass spectrometry. There were no significant changes in 24-hour food intake animal bodyweight during the ESI-09 administration for both sexes. Interestingly, ESI-09 treatment modestly increased 24-hour urinary volume in females but not in males. Inhibition of Epac with ESI-09 also produced a sustained natriuresis and kaliuresis, which was more apparent in females than in males. Moreover, treatment with ESI-09 led to a transient decrease in urinary pH during the first two days, which returned to the baseline at day 7. Western blot analysis detected a significant reduction in NHE-3 levels upon ESI-09 treatment, which was surprisingly more pronounced in males than in females. However, super-resolution STED microscopy reveal marked translocation of NHE-3 to the base of brush border of PT cells rendering its inactive in females but not in males. In contrast to the PT, we did not detect significant changes in γENaC and the apical Cl - /HCO 3 - exchanger pendrin in the CD in the ESI-09-treated animals suggesting a minor role of the CD in the observed natriuresis in response to Epac blockade in this experimental setting. In summary, our results demonstrate that pharmacological Epac blockade with ESI-09 causes a notable natriuresis in mice kept on a regular Na + intake by decreasing NHE-3-dependent Na + reabsorption in the PT. While NHE-3 levels were more strongly reduced in males, ESI-09 also led to a redistribution of NHE-3 to the base of brush border to account for the observed greater magnitude of natriuresis and kaliuresis in females. This abstract was presented at the American Physiology Summit 2026 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.