Aminopeptidase N/CD13 (Anpep) is a membrane-bound protein that catalyzes the formation of natriuretic hexapeptide angiotensin IV (Ang IV) from Ang III. We have previously reported that Anpep is more highly expressed in the kidneys of Dahl salt-resistant (SR/Jr) than salt-sensitive (SS/Jr) rats; Anpep maps to a quantitative trait locus (QTL) for hypertension; and that the Dahl SR/Jr rat contains a functional polymorphism of the gene. This suggests that renal Anpep may be linked to salt-sensitivity, however, its effect on renal Na handling has not been determined. Here, we have examined regulation of basolateral Na⁺/K⁺ ATPase, a preeminent basolateral Na⁺ transporter in proximal tubule cells, by Anpep in LLC-PK1 cells. Treatment of the cells with Anpep siRNA increased total cellular Na⁺/K⁺ ATPase activity and basolateral Na⁺/K⁺ ATPase abundance by approximately 2-fold. Conversely, Anpep over-expression reduced Na⁺/K⁺ ATPase activity and basolateral abundance by about 50%. Similar effects were observed after treatment with Ang IV (10 nM, x 30 min and 12 hours). Ang IV receptor (AGTRIV) knockdown via specific siRNA relieved the decreases in basolateral Na⁺/K⁺ ATPase levels and activity induced by Anpep over-expression. In sum, these results demonstrate that Anpep reduces basolateral Na⁺/K⁺ ATPase levels via Ang IV/AGTRIV signaling. This novel pathway may be important in renal adaptation to high salt.