It is generally accepted that Na(HCO3)n cotransport is the most important mechanism mediating basolateral bicarbonate efflux in the early proximal tubule. The presence of basolateral Na(HCO3)n cotransport in the late proximal tubule (S3 segment) and in the juxtamedullary S1 and S2 segments has been controversial. The renal sodium-bicarbonate cotransporter (NBC) has been recently cloned from rat (M. F. Romero, M. A. Hediger, E. L. Boulpaep, and W. F. Boron. J. Am. Soc. Nephrol. 7: 1259, 1996), salamander (M. F. Romero, M. A. Hediger, E. L. Boulpaep, and W. F. Boron. Nature 387: 409-413, 1997), and human (C. E. Burnham, H. Amlal, Z. Wang, G. E. Shull, and M. Soleimani. J. Biol. Chem. 272: 19111-19114, 1997). The localization of NBC in the kidney is unknown. The present study was designed to localize NBC mRNA expression in the rabbit proximal tubule. In situ hybridization studies were combined with functional studies of basolateral Na(HCO3)n cotransport in superficial and juxtamedullary S1, S2, and S3 segments of the rabbit proximal tubule. The results demonstrate that NBC mRNA is localized predominantly to the cortex, with less expression in the outer medulla. NBC expression was not detected in the inner medulla. The highest level of NBC mRNA is in the S1 proximal tubule. NBC is expressed at a low levels in the S3 segment, with intermediate expression in the S2 segment. In bicarbonate-buffered solutions, the rate of base efflux mediated by Na(HCO3)n cotransport followed a similar pattern in superficial and juxtamedullary proximal tubule segments, i.e., S1 > S2 > S3. The juxtamedullary S1 segment had the greatest rate of basolateral Na(HCO3)n cotransport and the highest level of NBC expression in the proximal tubule.