Characteristics of hemichannel-mediated substrate transport in human retinal pigment epithelial cells under deprivation of extracellular Ca²+

Retinal pigment epithelial (RPE) cells form the outer blood-retinal barrier (BRB) and regulate drug/ compound exchange between the neural retina and blood in the fenestrated blood vessels of retinal choroid via membrane transporters. Recent studies have elucidated that RPE cells express hemichannels, which are opened by extracellular Ca²⁺ depletion and accept several drugs/compounds as a transporting substrate. The objective of this study was to elucidate the hemichannel-mediated compound transport properties of the outer BRB. In human RPE cells, namely ARPE-19 cells, time-dependent uptake of fluorescent hemichannel substrates, such as Lucifer Yellow, sulforhodamine-101 (SR-101), and propidium iodide (PI) was promoted under Ca²⁺-depleted conditions. The uptake of these substrates under Ca²⁺-depleted conditions exhibited saturable kinetics with a Michaelis-Menten constant (K_m) of 87-109µM. In addition, SR-101 and PI uptake by ARPE-19 cells was dependent of extracellular Ca²⁺ concentration, and that under Ca²⁺-depleted conditions was significantly decreased by typical substrates and/or inhibitors for hemichannels. Moreover, Ca²⁺depleted conditions promoted the efflux transport of calcein from ARPE-19 cells, and the promoted calcein efflux transport was significantly inhibited by a typical hemichannel inhibitor. These results suggested that hemichannels at the outer BRB were involved in the influx and efflux transport of drugs/compounds.