2.3.2. Intervention 2: Spironolactone

Mice were anesthetized with inhaled 2% isoflurane and they received a subcutaneous implant of either spironolactone (25 mg, 21‐day slow‐release) or a vehicle pellet (Innovative Research of America). After surgery, mice were maintained in standard cages on a NS gel diet (TestDiets, microstabilized rodent diet LD101 with 0.16% Na⁺/0.2% K⁺) for 3 days, then switched to a HS diet for 7 days (LD101 with 1.6% Na⁺/0.2 K⁺). A subset of these mice were individually acclimated to metabolic cages for 2 days before 24 h urine collections (under water‐saturated mineral oil) occurred on day 7 of the HS diet. Urine was spun at 1000 g for 5 min to pellet any debris and the supernatant stored at −80°C until analysis. At this point, the animals were euthanized and the blood and kidneys were collected. Blood electrolytes were immediately analyzed with EC8+ cartridges and the i‐STAT (Abbot Labs). Urine electrolytes were measured by atomic absorption spectrophotometry (Thermo Fisher). Excretion was calculated by multiplying the concentrations by urine flow.