Quantitation of aquaporin-2 abundance in microdissected collecting ducts: axial distribution and control by AVP.

Aquaporin-2 (AQP-2) is the arginine vasopressin-regulated water channel of the renal collecting ducts. Using an improved version of a fluorescence-based enzyme-linked immunosorbent assay (Y. Maeda, B. L. Smith, P. Agre, and M. A. Knepper. J. Clin. Invest. 95: 422-428, 1995), we quantified AQP-2 protein abundance in microdissected renal collecting ducts from normal Sprague-Dawley (SD) rats and vasopressin-deficient Brattleboro rats. Standard curves were linear in the range of 0-200 fmol/well and were highly reproducible from day to day (lower limit of detection 2.3 fmol; coefficient of variation 6-9%). In SD rats thirsted for 24 h, the measured quantities of AQP-2 were as follows (x 10(9) molecules/mm): cortical collecting ducts (CCD), 4.3 +/- 0.5; outer medullary collecting ducts (OMCD), 10.1 +/- 1.7; initial one-third of inner medullary collecting duct (IMCD-1), 9.2 +/- 1.1; middle one-third of the IMCD (IMCD-2), 7.5 +/- 0.8; terminal one-third of the IMCD (IMCD-3), 3.3 +/- 0.6; n = 7-12. In IMCD-2 this corresponds to 11.8 +/- 1.3 x 10(6) AQP-2 molecules per cell. Thus AQP-2 is extremely abundant in collecting duct cells. AQP-2 levels were decreased in untreated Brattleboro rats relative to the parent strain Long-Evans (LE) by 68% in IMCD-2 and 44% in CCD. Following vasopressin infusion by osmotic minipumps, AQP-2 levels in IMCD-2 of Brattleboro rats rose gradually, reaching levels equivalent to those seen in LE rats after 5 days. A similar rise was seen in the CCD, indicating that the vasopressin-induced increase was not dependent on a large increase in the interstitial tonicity. Thus a rise in circulating vasopressin concentration increases the level of AQP-2 protein expression in collecting ducts, presumably via a direct action of vasopressin.