To examine the role of Na-K-ATPase in the natriuresis that occurs after acute extracellular volume expansion, we performed acute clearance experiments and in vitro analysis of renal microsomal ATPase activity in rats receiving intravenous 0.9% sodium chloride (0.1 ml/100 g bw/min). Despite increased absolute reabsorption of filtered sodium (196 +/- 8.1 vs. 165 +/- 11.4 uEq/min, p less than 0.05), renal medullary microsomal Na-K-ATPase activity was decreased from 134 +/- 5.9 to 110 +/- 6.3 pmol Pi/mg protein/hour (p less than 0.02). No changes occurred in cortical or papillary regions and Mg-ATPase was unaffected. Similar results were obtained after adding 4 mEq/l potassium chloride to the infusion to prevent any fall in serum K+. These data suggest that a considerable percentage of sodium reabsorption is suppressed in acutely volume expanded animals and it is proposed that this is mediated by inhibition of medullary Na-K-ATPase.