Carbon dioxide permeability of aquaporin-1 measured in erythrocytes and lung of aquaporin-1 null mice and in reconstituted proteoliposomes

Measurements of CO2 permeability in oocytes and liposomes containing water channel aquaporin-1 (AQP1) have suggested that AQP1 is able to transport both water and CO2. We studied the physiological consequences of CO2 transport by AQP1 by comparing CO2 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. CO2 permeability was measured from the rate of intracellular acidification in response to addition of CO2/HCO3- 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 (t(1/2), 7.3 +/- 0.4 ms, S.E., n = 11 mice) and blocked by acetazolamide and increasing external pH (to decrease CO2/HCO3- ratio). Apparent CO2 permeability (P-CO2) was not different in erythrocytes from wild-type (0.012 +/- 0.0008 cm/s) versus null (0.011 +/- 0.001 cm/s) mice. Lung CO2 transport was measured in anesthetized, ventilated mice subjected to a decrease in inspired CO2 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 t(1/2) 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 CO2 transport in erythrocytes and lung we re-examined CO2 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-CO2 (similar to 10(-3) cm/s) did not differ. Measurements using different CA concentrations and HgCl2 indicated that liposome P-CO2 is unstirred layer-limited and that HgCl2 slows acidification because of inhibition of CA rather than AQP1, These results provide direct evidence against physiologically significant AQP1-mediated CO2 transport and establish an upper limit to the CO2 permeability through single AQP1 water channels.