H,K‐ATPase type 2 Knock‐Out Mice Have A Defect In Na + Balance Maintenance During Mineralocorticoid Escape

Mineralocorticoids induce an increase in renal Na + retention, leading to an increase in plasma volume which contributes to the elevation of blood pressure. Mineralocorticoid excess also induces hypokalemia and, as a consequence an increase in the expression of the renal H,K‐ATPase type 2 (HKa2). This pump avidly reabsorbs K + and extrudes protons in the colon and the collecting duct (CD). Increasing evidence suggest this transporter plays an important role in Na + homeostasis. We asked whether HKa2 plays a role in the renal response to mineralocorticoid excess. WT and HKa2 KO mice where fed a normal diet (0.2 % Na + ) and were treated with deoxycorticosterone pivalate (DOCP). Daily urinary and fecal Na + and K + excretions were analyzed and compared to intake. We used Time Domain Nuclear Magnetic Resonance (TDNMR) to measure three body compartments: total fat, lean mass (muscle) and free fluid (unbound H 2 O). At baseline, WT and HKa2 KO mice showed similar Na + balance, fat (~6% BW), lean (~73% BW) and free fluid (WT 2.86 ± 0.12 ml or 9.8 ± 0.14 % BW; KO 2.89 ± 0.09 ml or 9.93 ± 0.28 % BW ± SEM). One day after DOCP administration, both WT and HKa2 KO mice showed a similar significant transient increase in free fluid (WT 21.99 ± 2.36 % increase p<0.001 n=5; KO 15.91 ± 3.69 % increase p< 0.005 ± SEM n=8), but fat and lean mass were unchanged. This increase correlated with a doubling in Na + urinary retention as well as a significant increase in Na + balance (WT 48.71 ± 17.37 to 119.87 ± 13.22 μEq ± SEM p<0.05; HKA2 KO 11.55 ± 12.22 to 116.84 ± 16.18 μEq ± SEM, p<0.005). Na + balance decreased rapidly in both groups. Interestingly HKa2 KO mice had a smaller Na + balance on day 3 of DOCP treatment (WT 36.63 ± 8.11 μEq ± SEM; HKA2 KO −23.34 ± 15.55 μEq ± SEM, p<0.05). This “mineralocorticoid escape” phase, was due to a decrease in urinary Na + retention that was significantly more rapid in HKa2 KO mice. On day 4, urinary Na + retention and Na + balance had returned to normal in both genotypes. Interestingly, free fluid stayed significantly greater than baseline until day 4 in both groups. We also examined how DOCP affected K + balance. At baseline, HKa2 showed a normal K + balance although their fecal K + excretion was 4 times that of WT mice and their urinary K + retention was not increased. HKa2 KO mice ate significantly more than WT mice. Increasing their K + intake may have allowed them to compensate for their fecal loss. DOCP induced a striking increase in HKa2 KO mice fecal K + excretion that peaked on day 3 (control: 42.75 ± 2.55 μEq, day 3: 119,03 ± 6,81 μEq ± SEM) but had no significant effect in WT mice. DOCP did not induce an increase in urinary K + excretion in either mice group. In fact, urinary K + retention increased and K + balance was preserved in both genotypes throughout the experiment. Nevertheless, after 8 days of DOCP treatment, both WT and HKa2 KO mice where hypokalemic to the same extend (WT 3.67 ± 0.08 mM vs 3.00 ± 0.08 mM p<0.01, KO 3.69 ± 0.07 mM vs 2.95 ± 0.08 mM p<0.0005 ± SEM). In conclusion, we show for the first time, that mineralocorticoids induce a rapid and transient increase in free fluid by TDNMR that follows the increase in Na + balance. Moreover, our results show that HKa2 KO mice keep a normal K + balance under mineralocorticoid excess but have a defect in renal Na + handling during the escape phase. Support or Funding Information VA Merit Review Award IO1BX001472‐01A1 to C. S. WingoNa balanceFree fluid