Generation and characterization of thick ascending limb specific knockout of NHE3 mouse model

The Na+/H+ exchanger 3 (NHE3) mediates Na+ (re)absorption in the intestine and the kidney, thereby playing a key role in fluid homeostasis and blood pressure (BP) regulation. NHE3 is expressed in the luminal membrane of the renal proximal tubule, where the majority of renal Na+ reabsorption (60% to 75%) occurs. Whole body NHE3 knockout mice exhibit hypovolemia, hypotension, diarrhea, metabolic acidosis and renal Na+ wasting. Study in kidney-specific (proximal tubule [PT] and thick ascending limb [TAL]) NHE3 knockout mice has reported a rather mild phenotype in terms of Na+ homeostasis and no phenotype for acid-base homeostasis (Kidney International; 2017, 92:397-414). NHE3 is also substantially expressed in the TAL and our data point to a novel role in this nephron segment for urinary concentrating ability. To determine the physiological role of NHE3 in the TAL for salt homeostasis, urinary concentrating ability and BP regulation, we generated a novel tamoxifen (Tam)-inducible TAL-specific NHE3 knockout (NHE3TAL-KO) mice by crossing floxed NHE3 (Control, Con) with UmodCreERT2 mice. Immunofluorescent staining showed that application of Tam (3 days, 67 mg/kg via oral gavage) selectively deleted NHE3 from TAL but not PT. Before Tam, in Con and NHE3TAL-KO mice had comparable food (4.9±0.1 vs 5.3±0.2 g/(day*30g)) and water intake (5.6±0.3 vs 5.9±0.4 g/(day*30g)), urinary osmolality (2435±107.4 vs 2220±111.9 mOsm/kg), urinary pH (6.02±0.06 vs 6.02±0.06), urinary Na+/creatinine (46.3±6.2 vs 52.0±5.2 mmol/mmol) and K+/creatinine (109.0±9.5 vs 97.2±9.2 mmol/mmol). Two weeks after Tam, compared to Con mice, NHE3TAL-KO mice showed lower urinary osmolality (2224±90.1 vs 1826±92.8 mOsm/kg, P<0.01) associated with a strong tendency of higher water intake (5.3±0.2 vs 6.1±0.3 g/(day*30g), P=0.08). However, no difference was found in urinary pH (6.07±0.05 vs 6.17±0.05, NS), urinary Na+/creatinine (37.4±4.6 vs 40.9±4.5 mmol/mmol, NS) and K+/creatinine (80.9±5.7 vs 94.1±6.8 mmol/mmol, NS). In conclusion, we successfully generated TAL-specific NHE3 knockout mouse model. This mouse model will be used to study the isolated role of NHE3 in the TAL regarding to salt homeostasis, urinary concentrating ability and BP regulation to distinguish the contribution of TAL from PT. © FASEB.