Carbon Dioxide Permeability of Aquaporin-1 Measured in Erythrocytes and Lung of Aquaporin-1 Null Mice and in Reconstituted Proteoliposomes

Abstract

Measurements of CO 2 permeability in oocytes and liposomes containing water channel aquaporin-1 (AQP1) have suggested that AQP1 is able to transport both water and CO 2 . We studied the physiological consequences of CO 2 transport by AQP1 by comparing CO 2 permeabilities in erythrocytes and intact lung of wild-type and AQP1 null mice. Erythrocytes from wild-type mice strongly expressed AQP1 protein and had 7-fold greater osmotic water permeability than did erythrocytes from null mice. CO 2 permeability was measured from the rate of intracellular acidification in response to addition of CO 2 /HCO 3 ؊ in a stopped-flow fluorometer using 2,7-bis-(2-carboxyethyl)-5-(and -6)-carboxyfluorescein (BCECF) as a cytoplasmic pH indicator. In erythrocytes from wild-type mice, acidification was rapid (t1 ⁄2 , 7.3 ؎ 0.4 ms, S.E., n ‫؍‬ 11 mice) and blocked by acetazolamide and increasing external pH (to decrease CO 2 /HCO 3 ؊ ratio). Apparent CO 2 permeability (P CO 2 ) was not different in erythrocytes from wild-type (0.012 ؎ 0.0008 cm/s) versus null (0.011 ؎ 0.001 cm/s) mice. Lung CO 2 transport was measured in anesthetized, ventilated mice subjected to a decrease in inspired CO 2 content from 5% to 0%, producing an average decrease in arterial blood pCO 2 from 77 ؎ 4 to 39 ؎ 3 mm Hg (14 mice) with a t1 ⁄2 of 1.4 min. The pCO 2 values and kinetics of decreasing pCO 2 were not different in wild-type versus null mice. Because AQP1 deletion did not affect CO 2 transport in erythrocytes and lung, we re-examined CO 2 permeability in AQP1-reconstituted liposomes containing carbonic anhydrase (CA) and a fluorescent pH indicator. Whereas osmotic water permeability in AQP1-reconstituted liposomes was >100-fold greater than that in control liposomes, apparent P CO 2 (ϳ10 ؊3 cm/s) did not differ. Measurements using different CA concentrations and HgCl 2 indicated that liposome P CO 2 is unstirred layer-limited and that HgCl 2 slows acidification because of inhibition of CA rather than AQP1. These results provide direct evidence against physiologically significant AQP1-mediated CO 2 transport and establish an upper limit to the CO 2 permeability through single AQP1 water channels.A family of related water-transporting proteins (aquaporins, AQP) 1 has been identified in which individual members are expressed in many fluid-transporting epithelia and endothelia. Many of the mammalian aquaporins appear to transport only water, whereas others (AQP3 and AQP7) transport small polar solutes such as glycerol and urea, and even larger solutes including monosaccharides (AQP9) (1-4). Structural information is available for AQP1, the water channel expressed in erythrocytes, kidney tubules and microvessels, alveolar endothelia, choroid plexus, ciliary body, and other tissues. AQP1 molecules associate in tetramers (5) in which the monomeric subunits are functionally independent with respect to water transport (6). Electron crystallography revealed that each AQP1 monomer contains six membrane-spanning, tilted helical domains (7-9); however, the resolut...

110 Years of the Meyer–Overton Rule: Predicting Membrane Permeability of Gases and Other Small Compounds

“…[58] Consequently, the change in the intravesicular acidification rate observed upon aquaporin reconstitution [47] cannot be interpreted in terms of a facilitated CO 2 transport by these channels. [61] The lack of an effect of carbonic anhydrase may indicate that CO 2 transport into vesicles is generally limited by CO 2 diffusion towards the vesicles through near-membrane ULs. [61] However, a realistic estimation of the UL effects reveals that the combined permeabilities of the external and internal ULs, δ out and δ in in size, is equal to [Eq.…”

“…[61] The lack of an effect of carbonic anhydrase may indicate that CO 2 transport into vesicles is generally limited by CO 2 diffusion towards the vesicles through near-membrane ULs. [61] However, a realistic estimation of the UL effects reveals that the combined permeabilities of the external and internal ULs, δ out and δ in in size, is equal to [Eq. (15)]: (15) P UL is, thus, much larger than the estimates given for P M,CO2 ≈1.2×10 −3 cm s −1 in ref.…”

“…(15)]: (15) P UL is, thus, much larger than the estimates given for P M,CO2 ≈1.2×10 −3 cm s −1 in ref. [61] and ≈1.9×10 −3 cm s −1 in ref. [47].…”

“…However, CO 2 flux through aquaporin-containing vesicles was independent on the concentration of external carbonic anhydrase. [61] In the absence of carbonic anhydrase, the reactions [Eqs. (14)]: (14) would proceed with k 1 ≈3.7×10 −2 s −1 and k 2 ≈2×10 −4 s −1 , that is, CO 2 would form much slower than it is transported through the membrane.…”

Barley plasma membrane intrinsic proteins (PIP Aquaporins) as water and CO2 transporters

“…In mammalian cells, there were some indications that CO 2 could permeate AQP1 [5,20], but Yang et al raised a serious question over this interpretation [30].…”