In rats, urine concentrating ability develops progressively from birth and reaches the adult level at weaning in the fourth postnatal week. We addressed intrarenal mechanisms for this phenomenon. We determined the abundance and cellular localization of Na and Cl-transporters in the thick ascending limb of Henle′s loop (TAL) during postnatal development and tested whether their expression was stimulated by glucocorticoid, which exhibits a marked surge at weaning. Moreover, we determined whether proliferation in TAL cells during postnatal development excludes expression of NaCl transport molecules. Rat pups were humanely killed and for immunohistochemistry they were anaesthetized with sodium pentobarbitone (0.5 mg/10 g rat) and perfused with 4% paraformaldehyde through the left cardiac ventricle. Expression level and localization of NaCl transporters NKCC-2, NHE-3, ROMK, CLCK2 and Na⁺-K⁺-ATPase-α1 was determined by ribonuclease protection, Western blotting and immunohistochemistry in a developmental series of kidneys. The effect of dexamethasone (DEX) at the nadir of endogenous glucocorticoid (postnatal day (P)8-P12, 100 μg/kg) on transporter proteins and urinary concentrating ability was tested (n=5-8). Cell proliferation was assessed by immunostaining and Western blot for proliferating cell nuclear antigen (PCNA). Messenger RNAs for all tested NaCl transporters were significantly increased with postnatal development with a peak at weaning (P21). There were significant increase of Na⁺-K⁺-ATPase-α1, NHE-3 and ROMK between P7 and 21 (2234 ± 55 vs 2986 ± 84 cpm, 210 ± 20 vs 320 ± 10 cpm, and 88 ± 4 vs 129 ± 7 cpm, n = 4-5, P<0.01, respectively). For NKCC2 there was a significant increase between P0 and P21 (201 ± 12 vs 356 ± 22 cpm, n = 4-5, P<0.001). After P21, mRNA levels stabilized or decreased. Immunostaining for NKCC-2, ROMK and Na⁺-K⁺-ATPase-α1 confirmed a marked increase in labelling intensity and more wide distribution along loops of Henle with development. DEX injections increased significantly the abundance of NHE-3, NKCC2, Na⁺-K⁺-ATPase-α1 and ROMK (201 ± 19 vs 282 ± 33 cpm, 495 ± 31 vs 707 ± 16 cpm, 5451 ± 109 vs 6951 ± 228 cpm, and 109 ± 8 vs 175 ± 7 cpm, n = 7-8, P<0.001, respectively) mRNAs. Immunostaining confirmed up-regulation of NKCC2 and Na⁺-K⁺-ATPase-α1 by DEX. DEX treatment led to a significant increase of urinary concentrating ability (871 ± 17 vs 643 ± 77 mosm/kg, thirst 12 h) at P12. Kidney outer medulla was increasingly labelled for PCNA with development with a peak around P10-14. PCNA co-localized with the TAL marker Tamm-Horsfall glycoprotein. PCNA labelling was almost exclusively restricted to TAL cells not expressing NKCC-2. PCNA labelling was decreased in response to DEX. We conclude that development of urinary concentrating capacity is associated with increased expression and more wide distribution of crucial NaCl transporter proteins and fewer proliferating cells in TAL. As proliferating TAL cells differentiate they acquire NaCl transporters and this process is accelerated by glucocorticoid during postnatal kidney development.