Assay for molecular transport across gap junction channels in one-dimensional cell arrays

Abstract

Transport across gap junction channels (GJCs) between neighboring cells is critical to synchronizing cell's electrical and metabolic activities and maintaining cell homeostasis. Here we present a non-invasive microfluidic method to measure molecular diffusion across GJCs in multiple 1D cell arrays in real time. Using the chip, selective loading of a membrane permeant fluorescence dye (carboxyfluorescein) in Normal Rat Kidney (NRK) cells shows that the dye was able to diffuse through three cells along single cell chains in ∼35 minutes. Application of 100 µM 2-aminoethoxydiphenyl borate (2-APB) reversibly inhibits connexin-43 gap junctions in NRK cells; 0.8 mM 1-heptanol inhibits the diffusion partially. The method offers rapid exchange of reagents, enabling sequential screening of multiple gap junction specific drugs with only one preparation of cells. It is capable of measuring gap junction mediated diffusion between single cells.