CHRONIC METABOLIC-ACIDOSIS ENHANCES NHE-3 PROTEIN ABUNDANCE AND TRANSPORT ACTIVITY IN THE RAT THICK ASCENDING LIMB BY INCREASING NHE-3 MESSENGER-RNA

Chronic metabolic acidosis (CMA) is associated with an adaptive increa se in the bicarbonate absorptive capacity of the rat medullary thick a scending limb (MTAL). To specify whether NHE-3, the apical MTAL Na/H: exchanger, is involved in this adaptation, NHE-3 mRNA was quantified b y a competitive RT-PCR using an internal standard which differed from the wild-type NHE-3 mRNA by an 80-bp deletion. CMA increased NHE-3 mRN A from 0.025 +/- 0.003 to 0.042 +/- 0.009 amol/ng total RNA (P < 0.005 ), NHE-3 transport activity was measured as the initial proton flux ra te calculated from the Na-dependent cell pH recovery of Na-depleted ac idified MTAL cells in the presence of 50 mu M HOE694 which specificall y blocks NHE-1, the basolateral MTAL NHE isoform, CMA caused a 68% inc rease in NHE-3 transport activity (P < 0.001), In addition, CMA was as sociated with a 71% increase in NHE-3 protein abundance (P < 0.05) as determined by Western blot analysis on MTAL membranes using a polyclon al antiserum directed against a cytoplasmic epitope of rat NHE-3. Thus , NHE-3 adapts to CMA in the rat MTAL via an increase in the mRNA tran script that enhances NHE-3 protein abundance and transport activity.