A fluorescent dye uptake approach in cultured smooth muscle PD0332991 nmr cells was used to determine whether these cells have functional hemichannels. Results: We report for the first time that pannexins are expressed in the cerebral vasculature. We reveal that pannexin 1 is expressed in smooth muscle but not in endothelium and pannexin 2 is expressed in both endothelium and
smooth muscle. Fluorescent dye entered cultured smooth muscle cells in the absence of extracellular calcium or when the cells were depolarized, which was prevented by the putative hemichannel blocker carbenoxolone. Conclusions: The identification of pannexins in rat MCA indicates that pannexin expression is not restricted to neuronal cells. Dye uptake in cultured smooth muscle cells exhibited properties similar to those of connexin and pannexin hemichannels, which may represent another form of cell-to-cell communication within the vasculature. Copyright (C) 2012 S. Karger AG, Basel”
“Antagonists that are sufficiently selective to preferentially block GluN2A-containing N-methyl-D-aspartate receptors (NMDARs) over GluN2B-containing BAY 11-7082 in vitro NMDARs are few in number. In this study we describe a pharmacological characterization of 3-chloro-4-fluoro-N-[4-[[2-(phenylcarbonyl)hydrazino]carbonyl] benzyl]benzenesulphonamide (TCN 201), a sulphonamide derivative, that
was recently identified from a high-throughput screen as a potential GluN2A-selective selleck inhibitor antagonist. Using two-electrode
voltage-clamp (TEVC) recordings of NMDAR currents from Xenopus laevis oocytes expressing either GluN1/GluN2A or GluN1/GluN2B NMDARs we demonstrate the selective antagonism by TCN 201 of GluN2A-containing NMDARs. The degree of inhibition produced by TCN 201 is dependent on the concentration of the GluN1-site co-agonist, glycine (or D-serine), and is independent of the glutamate concentration. This GluN1 agonist-dependency is similar to that observed for a related GluN2A-selective antagonist, N-(cyclohexylmethyl)-2[5-[(phenylmethyl)amino]-1,3,4-thiadiazol-2-ylthio]acetamide (TCN 213). Schild analysis of TCN 201 antagonism indicates that it acts in a non-competitive manner but its equilibrium constant at GluN1/GluN2A NMDARs indicates TCN 201 is around 30-times more potent than TCN 213. In cortical neurones TCN 201 shows only modest antagonism of NMDAR-mediated currents recorded from young (DIV 9-10) neurones where GluN2B expression predominates. In older cultures (DIV 15-18) or in cultures where GluN2A subunits have been over-expressed TCN 201 gives a strong block that is negatively correlated with the degree of block produced by the GluN2B-selective antagonist, ifenprodil. Nevertheless, while TCN 201 is a potent antagonist it must be borne in mind that its ability to block GluN2A-containing NMDARs is dependent on the GluN1-agonist concentration and is limited by its low solubility.