Evidence that aquaporin 1 is a major pathway for CO₂transport across the human erythrocyte membrane

Abstract

We report here the application of a previously described method to directly determine the CO2 permeability (P(CO2)) of the cell membranes of normal human red blood cells (RBCs) vs. those deficient in aquaporin 1 (AQP1), as well as AQP1-expressing Xenopus laevis oocytes. This method measures the exchange of (18)O between CO2, HCO3(-), and H2O in cell suspensions. In addition, we measure the alkaline surface pH (pH(S)) transients caused by the dominant effect of entry of CO2 vs. HCO3(-) into oocytes exposed to step increases in [CO2]. We report that 1) AQP1 constitutes the major pathway for molecular CO2 in human RBCs; lack of AQP1 reduces P(CO2) from the normal value of 0.15 +/- 0.08 (SD; n=85) cm/s by 60% to 0.06 cm/s. Expression of AQP1 in oocytes increases P(CO2) 2-fold and doubles the alkaline pH(S) gradient. 2) pCMBS, an inhibitor of the AQP1 water channel, reduces P(CO2) of RBCs solely by action on AQP1 as it has no effect in AQP1-deficient RBCs. 3) P(CO2) determinations of RBCs and pH(S) measurements of oocytes indicate that DIDS inhibits the CO2 pathway of AQP1 by half. 4) RBCs have at least one other DIDS-sensitive pathway for CO2. We conclude that AQP1 is responsible for 60% of the high P(CO2) of red cells and that another, so far unidentified, CO2 pathway is present in this membrane that may account for at least 30% of total P(CO2).

Relative CO 2 /NH 3 selectivities of AQP1, AQP4, AQP5, AmtB, and RhAG

“…As described in ref. 6, exposing an AQP1-expressing oocyte to a solution containing 5% CO 2 /33 mM HCO 3 Ϫ at a constant pH of 7.50 causes a transient rise in pH S , followed by an exponential decay (Fig. 1C Left, green record).…”

“…We used microelectrodes to measure pH S (6,50). Briefly, the pH electrode had a tip diameter of 15 m, was filled at its tip with H ϩ ionophore mixture B (Catalog no.…”

Relative CO2/NH3 selectivities of mammalian aquaporins 0–9

“…To monitor pHS, the tips (diameter ϭ 15 m) of microelectrodes were filled with H ϩ ionophore mixture B (catalog no. 95293; Fluka Chemical, Ronkonkoma, NY), and the electrodes were connected to a model FD223 electrometer (World Precision Instruments, Sarasota, FL) (16,42,43). The extracellular reference electrode was a glass micropipette filled with 3 M KCl, connected to a model 750 electrometer (World Precision Instruments), via a calomel half-cell.…”

“…Figure 2 shows representative traces. An alkalinization (rise in pH S ) is indicative of CO 2 influx (16,41,42,44). Conversely, an acidification (decrease in pH S ) indicates an influx of NH 3 .…”

Dynamics and Energetics of Permeation Through Aquaporins. What Do We Learn from Molecular Dynamics Simulations?

“…In particular, the role of AQP1 as a CO 2 channel has been a matter of lively debate ever since it has been reported that AQP1 increases the CO 2 permeability of Xenopus oocytes membranes (Nakhoul et al 1998). By now, aquaporins have been reported to increase the CO 2 permeability of membranes of oocytes (Cooper and Boron 1998;Nakhoul et al 1998), liposomes (Prasad et al 1998), and red blood cells (Blank and Ehmke 2003; Endeward et al 2006). Other studies did not report any impact of AQP1 on membrane CO 2 permeability (Yang et al 2000).…”