Plasma membrane depolarization: a possible coupling factor between epithelial transcellular and paracellular transport

Abstract

The coordinated transport through transcellular and paracellular pathways ensures efficient transepithelial flux of water and solutes. Despite the fact that the coupling between these processes is of vital importance, their link is not well understood. To gain insight into the mechanism we asked whether 1) depolarization, induced by apical electrogenic transport processes, can serve as a signal coupling transcellular and paracellular transport; 2) increased cell contractility might play a role in this process. Paracellular permeability across confluent monolayer of proximal tubular (LLC-PK1) cells was followed as apical-to-basolateral transport of 4 kDa FITC-dextran. We show that depolarization, elicited either by high extracellular [K+], or by glucose or alanine, substrates of apical electrogenic transporters, substantially increased paracellular permeability. The effect of glucose required Na+-dependent cotransport, since 2-deoxy-glucose, which is not a substrate of the Na+/glucose cotransporter, failed to increase paracellular permeability. Importantly, depolarization was accompanied with Rho activation and Rho kinase (ROK)-dependent MLC phosphorylation. Inhibitors of ROK and myosin ATPase (Y27632, and blebbistatin) partially prevented the depolarization-induced rise in paracellular permeability, suggesting that the Rho-ROK-MLC pathway and the consequent increase in contractility contribute to the effect. Thus, plasma membrane depolarization is a good candidate as a coupling signal between apical and paracellular transport. Supported by Canadian Institute of Health Research.