title: New selective inhibitors of calcium-activated chloride channels - T16A(inh) -A01, CaCC(inh) -A01 and MONNA - what do they inhibit?
abstract: OBJECTIVE
  T16A(inh)-A01, CaCC(inh)-A01 and MONNA are identified as selective inhibitors of the TMEM16A calcium-activated chloride channel (CaCC). The aim of this study was to examine the chloride-specificity of these compounds on isolated resistance arteries in the presence and absence (±) of extracellular chloride.
  
  
  METHODS
  Isolated resistance arteries were maintained in a myograph and tension recorded, in some instances combined with microelectrode impalement for membrane potential measurements or intracellular calcium monitoring using fura-2. Voltage-dependent calcium currents (VDCC) were measured in A7r5 cells with voltage-clamp electrophysiology using barium as a charge carrier.
  
  
  RESULTS
  Rodent arteries preconstricted with noradrenaline or U46619 were concentration-dependently relaxed by T16A(inh) -A01 (0.1-10 μM): IC50 and maximum relaxation were equivalent in ±chloride (30 min aspartate substitution) and the T16A(inh) -A01-induced vasorelaxation ±chloride were accompanied by membrane hyperpolarization and lowering of intracellular calcium. However, agonist concentration-response curves ±chloride, with 10 μM T16A(inh) -A01 present, achieved similar maximum constrictions although agonist-sensitivity decreased. Contractions induced by elevated extracellular potassium were concentration-dependently relaxed by T16A(inh)-A01 ±chloride. Moreover, T16A(inh) -A01 inhibited VDCCs in A7r5 cells in a concentration-dependent manner. CaCC(inh) -A01 and MONNA (0.1-10 μM) induced vasorelaxation ±chloride and both compounds lowered maximum contractility. MONNA, 10 μM, induced substantial membrane hyperpolarization under resting conditions.
  
  
  CONCLUSIONS
  T16A(inh) -A01, CaCC(inh) -A01 and MONNA concentration-dependently relax rodent resistance arteries, but an equivalent vasorelaxation occurs when the transmembrane chloride gradient is abolished with an impermeant anion. These compounds therefore display poor selectivity for TMEM16A and inhibition of CaCC in vascular tissue in the concentration range that inhibits the isolated conductance.
authors.affiliation: Department of Biomedicine, Aarhus University, Aarhus, Denmark.|Department of Cardiothoracic and Vascular Surgery, Aarhus University Hospital, Skejby, Denmark.|Department of Gynaecology and Obstetrics, Aarhus University Hospital, Skejby, Denmark.
authors.forename: D M B
authors.initials: DM
authors.lastname: Boedtkjer
authors.affiliation: Department of Biomedicine, Aarhus University, Aarhus, Denmark.
authors.forename: S
authors.initials: S
authors.lastname: Kim
authors.affiliation: Department of Biomedicine, Aarhus University, Aarhus, Denmark.
authors.forename: A B
authors.initials: AB
authors.lastname: Jensen
authors.affiliation: Department of Biomedicine, Aarhus University, Aarhus, Denmark.
authors.forename: V M
authors.initials: VM
authors.lastname: Matchkov
authors.affiliation: Department of Biomedicine, Aarhus University, Aarhus, Denmark.|Department of Gynaecology and Obstetrics, Aarhus University Hospital, Skejby, Denmark.|Aarhus Institute of Advanced Studies, Aarhus University, Aarhus, Denmark.
authors.forename: K E
authors.initials: KE
authors.lastname: Andersson
mesh_terms: D000818:Animals; D002460:Cell Line; D002536:Cerebral Arteries / Q000187:drug effects / Q000502:physiology; D018118:Chloride Channels / Q000037:antagonists & inhibitors* / Q000502:physiology; D005260:Female; D066298:In Vitro Techniques; D008297:Male; D008638:Mesenteric Arteries / Q000187:drug effects / Q000502:physiology; D008810:Mice, Inbred C57BL; D011743:Pyrimidines / Q000494:pharmacology*; D017208:Rats, Wistar; D013844:Thiazoles / Q000494:pharmacology*; D013876:Thiophenes / Q000494:pharmacology*; D014665:Vasodilator Agents / Q000494:pharmacology*; D062367:ortho-Aminobenzoates / Q000494:pharmacology*
chemical_list: C000607369:6-t-butyl-2-(furan-2-carboxamido)-4,5,6,7-tetrahydrobenzo(b)thiophene-3-carboxylic acid; D018118:Chloride Channels; C585066:N-((4-methoxy)-2-naphthyl)-5-nitroanthranilic acid; D011743:Pyrimidines; C578466:T16AInh-A01; D013844:Thiazoles; D013876:Thiophenes; D014665:Vasodilator Agents; D062367:ortho-Aminobenzoates