VAMP3 mediates NKCC2 trafficking in thick ascending limbs and is required for normal renal function and blood pressure

Abstract

In the thick ascending limb of Henle's loop (TAL), trafficking of NKCC2 into the apical membrane is essential for NaCl absorption and blood pressure control. The vesicle associated membrane protein 3 (VAMP3), a membrane fusion protein of the SNARE family, is expressed in the kidney. VAMP3 is localized apically in TALs and co‐immunoprecipitates with NKCC2. The role of VAMP3 on Na excretion and NKCC2 trafficking is unknown. We hypothesized that VAMP3 mediates NKCC2 exocytic delivery and maintains normal TAL function and renal Na excretion. We used adenovirus‐mediated in vivo gene silencing to decrease VAMP3 expression in rat outer medullas and measured NKCC2 exocytic delivery by surface biotinylation in TAL suspensions. Silencing VAMP3 by 68 ± 10% did not affect VAMP2/7/8 expression in TALs but blocked NKCC2 exocytic delivery rate by 91 ± 6% (p<0.05) and decreased total NKCC2 expression by 59 ± 8% (p<0.05). We studied the role of VAMP3 in urinary Na (UNa) excretion and blood pressure in VAMP3 knockout (KO) mice. VAMP3 KOs exhibited reduced NKCC2 expression by 44 ± 12% (p<0.05), elevated UNa excretion (wild type: 251 ± 11 vs VAMP3 KO: 294 ± 13 μmol/day, p<0.05) and lower systolic arterial pressure (WT: 114 ± 2 vs VAMP3 KO: 95 ± 2 mmHg, p<0.05). We conclude that VAMP3 mediates NKCC2 trafficking to the apical membrane and maintains NKCC2 expression. VAMP3 is required for normal Na excretion and blood pressure control. Funding: NIH, AHA