title: A Cell-Based Assay to Assess Hemichannel Function.
abstract: Activation of connexin hemichannels is involved in the pathophysiology of disorders that include deafness, stroke, and cardiac infarct. This aspect makes hemichannels an attractive therapeutic target. Unfortunately, most available inhibitors are not selective or isoform specific, which hampers their translational application. The absence of a battery of useful inhibitors is due in part to the absence of simple screening assays for the discovery of hemichannel-active drugs. Here, we present an assay that we have recently developed to assess hemichannel function. The assay is based on the expression of functional human connexins in a genetically modified bacterial strain deficient in K+uptake. These modified cells do not grow in low-K+medium, but functional expression of connexin hemichannels allows K+uptake and growth. This cell-growth-based assay is simple, robust, and easily scalable to high-throughput multi-well platforms.
authors.affiliation: Department of Cell Physiology and Molecular Biophysics, and Center for Membrane Protein Research, Texas Tech University Health Sciences Center, Lubbock, TX.
authors.forename: Srinivasan
authors.initials: S
authors.lastname: Krishnan
authors.affiliation: Department of Cell Physiology and Molecular Biophysics, and Center for Membrane Protein Research, Texas Tech University Health Sciences Center, Lubbock, TX.
authors.forename: Mariana C
authors.initials: MC
authors.lastname: Fiori
authors.affiliation: Department of Cell Physiology and Molecular Biophysics, and Center for Membrane Protein Research, Texas Tech University Health Sciences Center, Lubbock, TX.
authors.forename: Luis G
authors.initials: LG
authors.lastname: Cuello
authors.affiliation: Department of Cell Physiology and Molecular Biophysics, and Center for Membrane Protein Research, Texas Tech University Health Sciences Center, Lubbock, TX.
authors.forename: Guillermo A
authors.initials: GA
authors.lastname: Altenberg
mesh_terms: D000818:Animals; D001681:Biological Assay / Q000379:methods*; D017630:Connexins / Q000378:metabolism; D006801:Humans; D011188:Potassium / Q000378:metabolism; D020033:Protein Isoforms / Q000378:metabolism
chemical_list: D017630:Connexins; D020033:Protein Isoforms; D011188:Potassium
keywords: aminoglycoside; cerebrovascular accident; connexin; gap junction; high-throughput screening; infarct; ion channel