Quantitative automated assays in living cells to screen for inhibitors
of hemichannel function
Abstract
In vertebrates, intercellular communication is largely mediated by
connexins, a family of structurally related transmembrane proteins that
assemble to form hemichannels (HCs) at the plasma membrane. HCs are
upregulated in different brain disorders; hence represent innovative
therapeutic targets and identifying modulators of Cx-based HCs is of
great interest for better understanding their function and defining new
treatments. In this study, we developed automated versions of two
different cell-based assays to identify new pharmacological modulators
of HCs. The first assay follows the incorporation of a fluorescent dye,
Yo-Pro, by real-time imaging while the second is based on the quenching
of a fluorescent protein, YFPQL, by iodide after iodide uptake. These
assays were then used to screen a collection of 2,242 approved drugs and
compounds under development. This study led to the identification of 11
new HC blockers, active in the two assays, with 5 drugs active on HC but
not on gap junction (GJ) activities. To our knowledge, this is the first
screening on HC activity and our results suggest the potential of a new
use of already approved drugs in CNS disorders with HC impairments.