CO₂ permeability of cell membranes is regulated by membrane cholesterol and protein gas channels

Recent observations that some membrane proteins act as gas channels seem surprising in view of the classical concept that membranes generally are highly permeable to gases. Here, we study the gas permeability of membranes for the case of CO₂, using a previously established mass spectrometric technique. We first show that biological membranes lacking protein gas channels but containing normal amounts of cholesterol (30-50 mol% of total lipid), e.g., MDCK and tsA201 cells, in fact possess an unexpectedly low CO₂ permeability (P_CO2) of ∼0.01 cm/s, which is 2 orders of magnitude lower than the P_CO2 of pure planar phospholipid bilayers (∼1 cm/s). Phospholipid vesicles enriched with similar amounts of cholesterol also exhibit P_CO2 ≈ 0.01 cm/s, identifying cholesterol as the major determinant of membrane P_CO2. This is confirmed by the demonstration that MDCK cells depleted of or enriched with membrane cholesterol show dramatic increases or decreases in P_CO2, respectively. We demonstrate, furthermore, that reconstitution of human AQP-1 into cholesterol-containing vesicles, as well as expression of human AQP-1 in MDCK cells, leads to drastic increases in P_CO2, indicating that gas channels are of high functional significance for gas transfer across membranes of low intrinsic gas permeability.