NHE3 in the thick ascending limb is required for sustained but not acute furosemide-induced urinary acidification

Na⁺/H⁺ exchanger isoform 3 (NHE3) facilitates Na⁺ reabsorption and H⁺ secretion by the kidneys. Despite stronger NHE3 abundance in the thick ascending limb (TAL) compared with the S1 and S2 segments of the proximal tubule, the role of NHE3 in the TAL is poorly understood. To investigate the role of NHE3 in the TAL, we generated and phenotyped TAL-specific NHE3 knockout (NHE3^TAL-KO) mice. Compared with control mice, NHE3^TAL-KO mice did not show significant differences in body weight, blood pH, or plasma Na⁺, K⁺, or Cl^- levels. Fluid intake trended to be higher and urine osmolality was significantly lower in NHE3^TAL-KO mice. Despite a similar glomerular filtration rate, NHE3^TAL-KO mice had a greater urinary K⁺-to-creatinine ratio. One proposed role of NHE3 relates to furosemide-induced urinary acidification. Acute bolus treatment with furosemide under anesthesia did not result in differences in the dose dependence of urinary flow rate, Cl^- excretion, or maximal urinary acidification between genotypes; however, in contrast with control mice, urinary pH returned immediately toward baseline levels in NHE3^TAL-KO mice. Chronic furosemide treatment reduced urine osmolality similarly in both genotypes but metabolic alkalosis, hypokalemia, and calciuresis were absent in NHE3^TAL-KO mice. Compared with vehicle, chronic furosemide treatment resulted in greater Na⁺-K⁺-2Cl^- abundance regardless of genotype. Na⁺-phosphate cotransporter 2a abundance was also greater in furosemide-treated control mice compared with vehicle treatment, an effect that was absent in NHE3^TAL-KO mice. In summary, NHE3 in the TAL plays a role in the sustained acidification effect of furosemide. Consistent with this, long-term treatment with furosemide did not result in metabolic alkalosis in NHE3^TAL-KO mice.NEW & NOTEWORTHY Na⁺/H⁺ exchanger isoform 3 (NHE3) is very abundant in the thick ascending limb (TAL) compared with the proximal tubule. Much has been learned about the role of NHE3 in the proximal tubule; however, the function of NHE3 in the TAL remains elusive. A novel mouse model that lacks NHE3 selectively in the TAL not only shows a phenotype under baseline conditions but also identifies that NHE3 is required for sustained but not acute furosemide-induced urinary acidification.