Carbon Dioxide Transport through Membranes

Andreas Missner,Philipp Kügler,Sapar M Saparov,Klaus Sommer,John C Mathai,Mark L Zeidel,Peter Pohl

doi:10.1074/jbc.m800096200

Abstract

Several membrane channels, like aquaporin-1 (AQP1) and the RhAG protein of the rhesus complex, were hypothesized to be of physiological relevance for CO(2) transport. However, the underlying assumption that the lipid matrix imposes a significant barrier to CO(2) diffusion was never confirmed experimentally. Here we have monitored transmembrane CO(2) flux (J(CO2)) by imposing a CO(2) concentration gradient across planar lipid bilayers and detecting the resulting small pH shift in the immediate membrane vicinity. An analytical model, which accounts for the presence of both carbonic anhydrase and buffer molecules, was fitted to the experimental pH profiles using inverse problems techniques. At pH 7.4, the model revealed that J(CO2) was entirely rate-limited by near-membrane unstirred layers (USL), which act as diffusional barriers in series with the membrane. Membrane tightening by sphingomyelin and cholesterol did not alter J(CO2) confirming that membrane resistance was comparatively small. In contrast, a pH-induced shift of the CO(2) hydration-dehydration equilibrium resulted in a relative membrane contribution of about 15% to the total resistance (pH 9.6). Under these conditions, a membrane CO(2) permeability (3.2 +/- 1.6 cm/s) was estimated. It indicates that cellular CO(2) uptake (pH 7.4) is always USL-limited, because the USL size always exceeds 1 mum. Consequently, facilitation of CO(2) transport by AQP1, RhAG, or any other protein is highly unlikely. The conclusion was confirmed by the observation that CO(2) permeability of epithelial cell monolayers was always the same whether AQP1 was overexpressed in both the apical and basolateral membranes or not.

Highlights

The widely accepted model that gases like NH3, CO2, and O2 pass biological membranes by diffusion through the lipid matrix has been recently called into question
To test whether the lipid bilayer constitutes the main diffusion barrier for CO2 transport under these conditions, we imposed a CO2 concentration gradient across the membrane and measured resulting pH shifts in the immediate membrane vicinity on the trans side at various CO2 concentrations (Fig. 2). pH was detected by scanning microelectrodes that were moved stepwise in the direction normal to the planar bilayer [17, 19]
The system converged with a zero penalty term. Because this term penalizes large PM,CO2 values, which may be computed in case flux limitations by unstirred layers (USL), the observation indicated that bilayer resistance to CO2 flux (JCO2) was not negligible

Summary

Introduction

The widely accepted model that gases like NH3, CO2, and O2 pass biological membranes by diffusion through the lipid matrix has been recently called into question. The membrane protein channels AmtB and aquaporin-8 have been identified to transport NH3 [1, 2]. Lowering membrane resistance by insertion of CO2 conducting channels seems to be of questionable physiological relevance This is exactly what has been concluded from a study where red blood cell aquaporin-1 has been knocked out without any effect on PM,CO2 [11]. This view is further supported by molecular dynamics simulations of CO2 transport through AQP1, which revealed that such a transport process would be physiologically meaningless in phospholipid membranes of common composition [12, 13]. In case of applicability of Overtone’s rule to CO2 transport, neither AQP1 nor Rh proteins would be able to facilitate CO2 membrane diffusion

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Results

Conclusion