The passage of dissolved gasses—including oxygen, carbon dioxide, carbon monoxide, nitrogen, and ammonia—into and out of cells is a universal biological transport process that underlies metabolism and pH regulation. While there is broad consensus that dissolved gasses passively cross lipid bilayer membranes, there is significant disagreement regarding the rate of transport of these gasses, with reported values spanning several orders of magnitude. An accurate measurement of gas transport rates is important to understanding how cells control cytosolic gas concentrations, particularly in terms of how protein gas channels or changes in lipid composition can modulate these concentrations. We have deployed a microfluidic system to observe the transport of carbon dioxide into giant unilamellar vesicles (GUVs) fabricated from phospholipids and cholesterol. GUVs containing a pH-sensitive fluorescent indicator were introduced into a serpentine channel where they mixed with a CO2-saturated buffer. Adapting the method of Cama and coworkers, we captured continuous video micrographs of these vesicles moving down the channel as the CO2 permeated the membrane and acidified the vesicle lumen. We modeled the process of CO2 transport as a set of coupled reactions involving the buffer species, dissolved CO2 and carbonic acid, and the fluorophore, with membrane permeability as a free parameter. The change in GUV fluorescence with time was fit to this model to quantify permeability. This method produced membrane permeabilities on the order of 10-3 to 10-2 cm/s, with high concentrations of cholesterol decreasing permeability by an order of magnitude. These relatively low permeability values suggest that gas transport in biological membranes is likely aided and modulated by transport channel proteins.

Measuring the CO2 permeabilities of lipid bilayers as a function of cholestrol concentration in a continuous flow microfluidic mixer

ACS Spring Meeting 2021

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.