What Do Aquaporin Knockout Studies Tell Us about Fluid Transport in Epithelia?

Abstract

The investigation of near-isosmotic water transport in epithelia goes back over 100 years; however, debates over mechanism and pathway remain. Aquaporin (AQP) knockouts have been used by various research groups to test the hypothesis of an osmotic mechanism as well as to explore the paracellular versus transcellular pathway debate. Nonproportional reductions in the water permeability of a water-transporting epithelial cell (e.g., a reduction of around 80–90 %) compared to the reduction in overall water transport rate in the knockout animal (e.g., a reduction of 50–60 %) are commonly found. This nonproportionality has led to controversy over whether AQP knockout studies support or contradict the osmotic mechanism. Arguments raised for and against an interpretation supporting the osmotic mechanism typically have partially specified, implicit, or incorrect assumptions. We present a simple mathematical model of the osmotic mechanism with clear assumptions and, for models based on this mechanism, establish a baseline prediction of AQP knockout studies. We allow for deviations from isotonic/isosmotic conditions and utilize dimensional analysis to reduce the number of parameters that must be considered independently. This enables a single prediction curve to be used for multiple epithelial systems. We find that a simple, transcellular-only osmotic mechanism sufficiently predicts the results of knockout studies and find criticisms of this mechanism to be overstated. We note, however, that AQP knockout studies do not give sufficient information to definitively rule out an additional paracellular pathway.

Appendix: Wild-Type and AQP Knockout Data

The wild-type renal proximal tubule water flux we used is similar to that given by Whittembury and Reuss (1992) and that used by Mathias and Wang (2005). We took the water flux to be five times bigger than that in the proximal tubule in the case of the salivary glands and 100 times smaller than that in the proximal tubule in the case of the lung alveolus, similar to the rough estimates given by Ma et al. (1999, 2000) and Schnermann et al. (1998) [though Ma et al.(1999) gave an estimate of the maximum rate of saliva secretion of up to 20 times the rate of absorption in the proximal tubule, we used a more conservative estimate]. We assumed that the salivary glands and proximal tubule systems transport under near-isosmolar conditions, 5–10 % from isosmotic. The assumptions for water flux and osmolarity of transport are our main wild-type parameter assumptions, and the wild-type renal proximal tubule transport quantities derived from these assumptions—water permeability and salt transport—are also similar to those given by Whittembury and Reuss (1992) and those used by Mathias and Wang (2005). Because the lung alveolus has a high water permeability but low water transport and negligible measurable changes in transport rates and osmolarity in knockouts (Ma et al. 2000), we instead assumed a value of water permeability between the lower and upper values of salivary gland and renal proximal tubule water permeabilities and calculated the osmolarity of transport from this; the deviation from isosmotic transport was significantly lower than that assumed in the salivary glands and renal proximal tubule, as would be expected (Table 1).

Given the assumption on the degree of wild-type osmolarity, the solute transport rates were calculated assuming the validity of the collection boundary condition (Eq. 6). This condition was also used to estimate the measured solute transport rate given the measured osmolarity in knockouts as it is assumed to be valid independently of the osmotic mechanism (the osmotic mechanism predicts a possibly different osmolarity from that measured).

The knockout effects were based primarily on those given by Schnermann et al. (1998) and Ma et al. (1999, 2000) for the renal proximal tubule, salivary gland acini, and lung alveolus. There were no measured changes in transport in the lung alveolus. The estimate of the reduction in permeability for the proximal tubule was taken to be 90 %, as an upper-limit case, higher than the 78 % reduction found by Schnermann et al. (1998) but in line with the 89 % reduction in permeability found in proximal tubule vesicles by Ma et al. (1998) in another AQP1 knockout study. The reduction in permeability for the salivary glands was not measured by Ma et al. (1999), but the reduction was found to be 65 and 77 % for cells isolated from the parotid and sublingual glands, respectively, by Krane et al. (2001); we took the reduction to be 80 % as a representative upper value (Table 2).

Importantly, because the studies of Ma et al. (1998) and Krane et al. (2001) measured the reductions in isolated cells and vesicles, we have some confidence that these reductions are directly representative of the changes in the permeability of the transcellular-only pathway. We could not find measurements of the changes in permeability of single cells or vesicles for AQP5-deficient alveolar cells but made the assumption of a similar effect to that in salivary cells and proximal tubule vesicles.