Membrane-associated aquaporin-1 facilitates osmotically driven water flux across the basolateral membrane of the thick ascending limb

Abstract

Cabral PD, Herrera M. Membrane-associated aquaporin-1 facilitates osmotically driven water flux across the basolateral membrane of the thick ascending limb. Am J Physiol Renal Physiol 303: F621-F629, 2012. First published June 6, 2012 doi:10.1152/ajprenal.00268.2012.-The thick ascending limb of the loop of Henle (TAL) reabsorbs ϳ30% of filtered NaCl but is impermeable to water. The observation that little water traverses the TAL indicates an absence of water channels at the apical membrane. Yet TAL cells swell when peritubular osmolality decreases indicating that water channels must be present in the basolateral side. Consequently, we hypothesized that the water channel aquaporin-1 (AQP1) facilitates water flux across the basolateral membrane of TALs. Western blotting revealed AQP1 expression in microdissected rat and mouse TALs. Double immunofluorescence showed that 95 Ϯ 2% of tubules positive for the TAL-specific marker Tamm-Horsfall protein were also positive for AQP1 (n ϭ 6). RT-PCR was used to demonstrate presence of AQP1 mRNA and the TALspecific marker NKCC2 in microdissected TALs. Cell surface biotinylation assays showed that 23 Ϯ 3% of the total pool of AQP1 was present at the TAL basolateral membrane (n ϭ 7). To assess the functional importance of AQP1 in the basolateral membrane, we measured the rate of cell swelling initiated by decreasing peritubular osmolality as an indicator of water flux in microdissected TALs. Water flux was decreased by ϳ50% in Aqp1 knockout mice compared with wild-types (4.0 Ϯ 0.8 vs. 8.9 Ϯ 1.7 fluorescent U/s, P Ͻ 0.02; n ϭ 7). Furthermore, arginine vasopressin increased TAL AQP1 expression by 135 Ϯ 17% (glycosylated) and 41 Ϯ 11% (nonglycosylated; P Ͻ 0.01; n ϭ5). We conclude that 1) the TAL expresses AQP1, 2) ϳ23% of the total pool of AQP1 is localized to the basolateral membrane, 3) AQP1 mediates a significant portion of basolateral water flux, and 4) AQP1 is upregulated in TALs of rats infused with dDAVP. AQP1 could play an important role in regulation of TAL cell volume during changes in interstitial osmolality, such as during a high-salt diet or water deprivation. cell volume; loop of Henle; water channels; vasopressin; AQP-1 knockout THE THICK ASCENDING LIMB of the loop of Henle (TAL) reabsorbs ϳ30% of the filtered NaCl load. It is considered the "waterimpermeant" segment of the nephron because it reabsorbs Na but not water, thereby diluting the forming urine and establishing the corticomedullary gradient necessary for water reabsorption by the collecting duct. Based on measurements of transepithelial (lumen-bath) water permeability, net water flux is relatively low in isolated, perfused TALs, even in the presence of arginine vasopressin (44). Thus, it has long been assumed that the TAL does not contain water channels. While this may be true of the apical membrane, changes in basolateral osmolality result in rapid changes in cell volume (18 -20) strongly suggesting that water channels are present at the peritubular side of TAL cells to facilitate water flux. However, t...

Adaptable interaction between aquaporin‐1 and band 3 reveals a potential role of water channel in blood CO2transport

“…Nonand glycosylated AQP1 monomer, dimer, trimer, and tetramer (AQP1 oligomers) were labeled in an AQP1 Western blot. The molecular mass of the nonglycosylated AQP1 monomer was ;26 kDa, and the molecular masses of glycosylated AQP1 monomers ranged from 34 to 40 kDa (31)(32)(33). Middle: co-IP, with anti-band 3 mAb (BRIC170), pulled down much more AQP1 in Mi.III ghosts than in non-Mi.III ghosts.…”

Thick Ascending Limb of the Loop of Henle

“…Notably, however, the apical membrane of the TAL is completely water impermeable and TAL segments have an extremely low transepithelial water permeability. The basolateral membrane, however, expresses abundant Aquaporin-1 water channel protein, allowing for water flux across the basolateral membrane and changes in cell volume in response to changes in interstitial osmolality (48).…”

Comparative physiology and architecture associated with the mammalian urine concentrating mechanism: role of inner medullary water and urea transport pathways in the rodent medulla

“…The TAL cells undergo volume regulation (33,34,36), and AQP1 water channels in the basolateral membrane are believed to play a role in fluid flows associated with volume regulation (14). The rat and rabbit outer medullary TAL exhibits very low transepithelial urea permeability (58,118) and expresses no known urea transporters.…”

“…In addition, but not shown, AQP1 is expressed in the initial ϳ10% of the short-loop DTL(159) and has been reported to be expressed in the basolateral membrane of the thick ascending limb (TAL)(14). PT, proximal tubule; lDTL, long-loop descending thin limb; ATL, ascending thin limb; sDTL, short-loop descending thin limb; DCT, distal convoluted tubule; CCD, cortical collecting duct.…”