NHE3 is the predominant Na(+)/H(+) exchanger on the brush-border membrane (BBM) of the proximal tubule in adults. However, NHE3 null mice still have significant renal BBM Na(+)/H(+) activity. NHE8 has been localized to the BBM of proximal tubules and is more highly expressed in neonates than adult animals. The relative role of NHE8 in adult renal H(+) transport is unclear. This study examined whether there was compensation by NHE8 in NHE3(-/-) mice and by NHE3 in NHE8(-/-) mice. NHE3(-/-) mice had significant metabolic acidosis, and renal BBM NHE8 protein abundance was greater in NHE3(-/-) mice than control mice, indicating that there may be compensation by NHE8 in NHE3(-/-) mice. NHE8(-/-) mice had serum bicarbonate levels and pH that were not different from controls. NHE3 protein expression on the BBM was greater in NHE8(-/-) mice than in wild-type mice, indicating that there may be compensation by NHE3 in NHE8(-/-) mice. Both BBM NHE3 and NHE8 protein abundance increased in response to acidosis. Blood pressure and Na(+)/H(+) exchanger activity were comparable in NHE8(-/-) mice to that of controls, but both were significantly lower in NHE3(-/-) mice compared with control mice. Compared with NHE3(-/-) mice, NHE3(-/-)/NHE8(-/-) mice had lower blood pressures. While serum bicarbonate was comparable in NHE3(-/-) mice and NHE3(-/-)/NHE8(-/-) mice, proximal tubule Na(+)/H(+) exchange activity was less in NHE3(-/-)/NHE8(-/-) mice compared with NHE3(-/-) mice. In conclusion, NHE3 is the predominant Na(+)/H(+) exchanger in adult mice. NHE8 may play a compensatory role in renal acidification and blood pressure regulation in NHE3(-/-) mice.